Correspondence should be addressed to Klara Deacutegardin klaradegardinrochecom
Received 13 November 2016 Revised 25 January 2017 Accepted 15 February 2017 Published 5 March 2017
Academic Editor Christoph Krafft
Copyright copy 2017 Klara Deacutegardin et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited
The fight against medicine counterfeiting is a current focus of the pharmaceutical world Reliable analytical tools are needed topursue the counterfeiters Handheld devices present the advantage of providing quick results with analyses possibly performedon the field A large number of solid-dosage form medicines have been analyzed with a handheld Raman spectrometer 33 outof 39 product families could be successfully analysed The methods were validated with 100 of correct identification Eachproduct was additionally tested by the methods of the other products and successfully rejected A second validation wasperformed using counterfeits placebos and generics All the counterfeits were rejected with p values close to zero Somegenerics presented a similar formulation to the brand products and were then identified as such One placebo was positivelyidentified showing that low dosage products are difficult to analyze with Raman Robustness tests were carried out showing forinstance that the operator has no influence on the results and that the analyses might be performed through transparentpackaging The discovery mode was also investigated which proposes the chemical composition of the samples The resultsdemonstrated that the Raman handheld device is a reliable tool for the field analysis of counterfeits
1 Introduction
The pharmaceutical world is increasingly concerned with thecounterfeiting of medicines All types of medicines are tar-geted as well as all regions of the globe The quality of thecounterfeits reaching the patients is very variable In all casesthese products are dangerous for the patients since theyare not respecting the required hygiene for their goodmanufacturing [1ndash4] Quick tools are consequently requiredfor the analysis of suspect products in order to speed up theinvestigations and increase the chances to pursue the crimi-nals Several analytical tools have proved efficient for thereliable and fast analysis of counterfeit medicines Amongthem spectroscopy is one of the most widely used sinceit is mostly nondestructive and environment-friendly andrequires no sampling [5] Raman [6ndash12] near-infrared(NIR) [13ndash20] and midinfrared (MIR) [21ndash23] spectroscopyhave been especially on the focus for counterfeit analysis
Technological progress has enabled the development ofhandheld and portable spectrometers The aim of these
miniaturized devices is to be able to perform measurementson the field rather than having to send all the samples tothe lab Therefore time can be spared for the analysis byperforming a first screening of the samples Handheldspectrometers are for instance more and more used for thefield analysis of pharmaceuticals agriculture drugs explo-sives and mineral samples [24ndash33] Handheld Raman NIRand MIR spectrometers have been especially studied andcompared in the context of the analysis of counterfeit medi-cines [34ndash38] Several handheld Raman devices have beenevaluated in the literature A dual-laser handheld Ramanwas evaluated by Assi et al for the identification of threemedicines with chemometrics [39] and another handheldRaman spectrometer for the authentication of 9 productsand 22 test products using principal component analysis(PCA) [34] Counterfeit medicines of three products wereanalysed by Luczak and Kalyanaraman [40] Bate and Hesscompared the results obtainedwith a handheldRaman instru-ment and the Minilab for the analysis of several antimalarialmedicines [41] and for the authentication of antimalarial
HindawiJournal of SpectroscopyVolume 2017 Article ID 3154035 13 pageshttpsdoiorg10115520173154035
antibiotic and antimycobacterial medicines [42] A handheldRaman spectrometer was tested on 15 pharmaceutical prod-ucts by Bugay and Brush [43] and its specificity was evaluatedbyHajjou et al regarding the active pharmaceutical ingredient(API) content of six medicines [44] Kalyanaraman et alalso presented the general use of such a device for counterfeitdetection [45] Ricci et al compared the performance of ahandheld Raman instrument with a benchtop device anddesorption electrospray ionization (DESI) MS for the detec-tionof counterfeits of antimalarials [46]Theuseof ahandheldRaman device on the field in Nigeria was finally presented bySpink et al [47]
In this paper a handheld Raman spectrometer wasevaluated for the detection of counterfeits of a high numberof pharmaceutical medicines consisting in tablets capsulesand powders Robust methods were developed using a largedatabase of products In total there are several batches of33 product families and among them 62 formulations weremeasured including the different manufacturing sites of theproducts A detailed calibration and validation strategy wasproposed as well as some robustness tests an evaluation ofthe discovery mode results for the composition of the con-firmed counterfeits and a comparison with a lab instrument
2 Material and Methods
21 Instrument Characteristics and Settings A handheldRaman spectrometer was used to collect the spectra andperform the calibration The device is a direct dispersiveRamanusing a 785nm laser excitationwavelength Its spectralresolution varies between 8 and 105 cmminus1 in average and theRaman shift range is 250 to 2875 cmminus1 The integration timein automatic mode is of 12ms minimum
The tablet sample holder was used for the measurementof the tablets and capsules Some of the samples had to bemeasured directly against the cone of the Raman sincethey were too large for the holder While the tablets and thecapsules were not measured in their packaging the powderswere analysed through their original glass vial
22 Statistical Method The statistical method used for theidentification of the products was the one provided by themanufacturer The calibration consisted in taking spectraof the genuine products The device then computed auto-matically a ldquosignaturerdquo of the product with each spectrumrecorded Several signatures of the same product were thengathered into a method The signatures are based on a multi-variate test of equivalence This test is used to compare anewly acquired Raman spectrum and the reference spectraregistered in the database If the probabilistic value (p value)of the new spectrum is above or equal to 005 meaning thespectrum presents a match within the preset 95 percentconfidence limit the measurement is considered consistentwith the reference spectra and the device reports a ldquopassrdquoresult If the p value is inferior to 005 the device reports aldquofailrdquo result In the present method while the validation withthe genuine products should provide ldquopassrdquo results the coun-terfeits generics and placebos should provide a ldquofailrdquo result
The multivariate test of equivalence is computed using allthe channels across the detector array region and can beconsidered a multivariate version of a statistical t-test Theuncertainty is directly modeled by the instrument softwareand thus no modeling has to be performed by the user
23 Calibration and Validation Strategy The calibration ofthe methods consisted in taking several spectra of a productwhichwere automatically converted in signatures and gather-ing them into a single method One method per product wasideally created For products presenting different formula-tions between the dosages (different excipients) two or moremethods were created per product
The validation was performed with an independent set ofspectra The samples were tested on the methods in order tocheck if they were correctly recognized All tested sampleshad to deliver a ldquopassrdquo result with the appropriate method
Additionally the products were tested on all the othermethods to check that no mismatch could occur for theseproducts
A second validation was then performed using counter-feits placebos and generics in order to test the ability ofthe handheld Raman spectrometer to detect counterfeitsThe generics and the placebos were tested in order to mimicthe ldquobest case counterfeitsrdquo All these samples had to berejected (present a ldquofailrdquo result) by the instrument
24 Samples
241 Genuine Samples At least 5 batches per product weremeasured with the split of 3 batches for the calibration setand 2 batches for the validation set for each of the 62 studiedformulations of the 33 product families For the productsmanufactured in several sites more than 5 batches weremeasured in order to include more variability One sampleper batch and three spectra per sample were measured Thesamples were slightly moved (for the capsules and powders)or flipped (for the tablets) between each measurement 543signatures were acquired for the calibration while the valida-tion was performed with 392 spectra
242 Placebos Generics and Counterfeits 44 counterfeitsamples (from eight different seizures) were tested whichhad already been confirmed as counterfeits by previous anal-yses In order to enlarge the number of samples 3 placebosand 14 generics from 5 different batches were also testedagainst the methods On the whole 61 spectra were acquiredfor the second validation samples
25 Robustness Tests Once the methods were calibratedvalidated and tested against the counterfeits placebos andgenerics 4 additional tests were performed in order to checkthe robustness of the methods
The first robustness test (R1) consisted in measuring thesame sample 20 times at the same position in order to see ifthe laser degraded the sample Two samples (one tablet andone capsule) were tested
In the second robustness test (R2) two samples (onetablet and one capsule) were measured 10 times at differentpositions in the sample holder The objective was to evaluate
2 Journal of Spectroscopy
if the position of the measurement had an influence onthe result
The influence of the sampling was evaluated in the thirdrobustness test (R3) Three samples (one tablet and twocapsules) were measured either through a glass vial througha blister (transparent or white) or directly against the cone ofthe instrument
Finally the fourth robustness test (R4) consisted in mea-suring two samples (one tablet and one capsule) three timesby two operators with the aim of testing if the operatorshad an influence on the measurement
3 Results and Interpretation
31 Spectrum Interpretation The spectra obtained by thedevice were first evaluated Several challenges to the creationof the calibration could be pointed out
First of all it can be observed that some product fam-ilies present heterogeneous spectra Two or more dosagesof the same product can indeed generate different spectrawhen a different formulationmdashit means a different excipientprofilemdashwas used (see Figure 1) Additionally in rare casesthe formulation can differ for the same product and the samedosage depending on the manufacturing site of the prod-uct (see Figure 2) It is thus important to include all thedosages and if possible all the manufacturing sites of aproduct into the methods Different methods might thenhave to be created for one product if the spectra of theproduct family differ Products 19 and 23 are indeedconcerned by both the heterogeneity within the productfamily and the different formulations depending on themanufacturing site Depending on the dosage or themanufacturing site these products underwent changes inthe composition of the sugar excipients mainly goingfrom lactose to starch Since these excipients have a big
influence on the spectral pattern of these low-dosageproductsmdash03 06 and 1 API for product 19 and13 and 6 API for product 23mdashit might be difficultto perform the identification unless separate methods arecreated per formulation
Another challenge to the calibration is that similarexcipient profiles can be found between different productfamilies The resulting spectra will then be similar likethe ones presented in Figure 3 and thus the products willbe hard to differentiate This is the case for products 18and 23 which contain the same excipients and have lowdosage of APImdash2 for product 18 and 50 and 83 forproduct 23 However small spectral differences can beobserved between products 18 and 23 in the range of1500 and 1700 cmminus1 and are due to API bands of product18 The two other products concerned by the similarity ofexcipient patterns are products 31 and 32 which withtheir high content of APImdash83 and 50mdashshould how-ever be distinguishable
Some generics present formulations that are almostidentical to the ones of the brand products This will resultin similar spectra that are not possible to distinguish fromthe brand product like in the example in Figure 4 In thesecases a ldquopassrdquo result can be expected
Low-dosage products are difficult to analyse with Ramanspectroscopy since the API will be hardly observed in thespectrum of the medicine The limit of detection of API inRaman spectra is quite difficult to define especially becauseit depends on the excipient pattern of the medicine In thestudied products excipient patterns made of sugar likestarch cellulose or glucose are predominant in the spectraand might mask the API bands In a previous study a limitof detection of 06 was encountered while the authorsindicated that the limit of detection was usually locatedaround 1 or 2 of API [48] For this reason the placebos of
Product 19 XmgProduct 19 Ymg
3456789
10111213141516171819
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 1 Raman spectra of two dosages of the same product family (product 19) Due to differences in the formulation the product presentsheterogeneous spectra
3Journal of Spectroscopy
low-dosage products might provide spectra that are verysimilar to the genuine products This was confirmed forproduct 26 which with an API content of 06 and anexcipient pattern dominated by starch and lactose hasidentical spectra between the genuine product and itsplacebo
The quality of the spectra might also have an influence onthe calibration The measured spectra present a wide rangefrom 250 to 2875 cmminus1 The end of the range approximatelyfrom 1800 to 2875 cmminus1 is often noisy and not necessarilyuseful since it does not bring additional information aboutthe product Last but not least a few samples like product
32 underwent fluorescence (Figure 5) This phenomenonwas observed when special excipients were used in theformulation of either the tablet or the shell of the capsules Inthis study in which a laser wavelength of 765 nm was usedindigo carmine (for product 32) microcrystalline cellulosehydroxypropyl methylcellulose and croscarmellose werefound responsible for the fluorescence phenomenon Whilethe fluorescence made the identification harder most bandsof the API and the excipients could still be observed like inFigure 5 When measuring other products not presented inthis paper the fluorescence was however so strong that no sig-nature could even be taken and no method created
Product 19 Xmg manufacturing site AProduct 19 Xmg manufacturing site B
4
6
8
10
12
14
16
18
20
22
24
26
28
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 2 Raman spectra of two samples produced at different manufacturing sites and possessing the same dosage (product 19 Xmg) Due todifferences in the chosen formulation the product presents heterogeneous spectra
Product 23 XmgProduct 18
468
1012141618202224262830323436384042
500 1000 1500 2000 2500Raman shift (cm‒1)
Ram
an in
tens
ity
Figure 3 Raman spectra of two different products (product 18 and product 23 Xmg) The products present similar excipients profiles andthus close Raman spectra
4 Journal of Spectroscopy
32 Identification Results
321 Calibration and Validation Results There are 33product families and among them 62 formulations couldbe successfully measured and integrated in the database ofthe instrument In total 31 methods have been created fortablets 6 for capsules and one for powders Most methodsgather spectra of several dosages per product For the productfamilies that presented heterogeneous formulations severalmethods have been created
100 of the spectra from the validation set has beencorrectly identified according to the pVal limit for positive
identification (superior to 005) According to the manufac-turer a pVal lower than 01 is considered too low for a robustmethod For all the developed methods the minimum pValwas higher than 01 (see Table 1)
Additionally no confusion could be observed whentesting the products on all the other methods than theirdedicated method 100 of all other products were rejectedby the tested method Table 1 presents the minimum p valuesobtained for the validation samples of the correspondingmethod together with the maximum p values of the closestother products These values show that the methods weresuccessfully validated and are very robust
Figure 4 Raman spectra of a brand product and one of its generic (product 20 Zmg) Both samples present a similar formulation and thesame dosage and thus similar Raman spectra
Product 32
20
25
30
35
40
45
50
55
60
65
70
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 5 Raman spectrum of product 32 While a strong fluorescence phenomenon can be observed the characteristic bands of the productare still visible enabling a correct identification
5Journal of Spectroscopy
A selectivity test proposed by the device was evaluatedafter the validation This test consists in evaluating for eachmethod the risks of getting mismatches The option wastested for the products providing the lowest p values duringthe validation For product 26 for instance the selectivity testsuggested that the closest other products would deliver a pvalue of 00026 Tested physically during the validation theproduct actually delivered a p value of 00352 using the
method of product 26 While the selectivity test might bean option to see quickly if possible mismatches might occurthe results delivered are not real The validation performed inthis study is thus considered more completed than theselectivity test Raman can thus not be considered a universalmethod for counterfeit detection
For six other products not included in the 33 presentedproduct families no method could be created since the
Table 1 Validation results obtained with the genuine products The number of signatures measured for each product has been added Theminimum p values (pVal min) obtained for the validation samples of the corresponding method are presented in the table together with thecorresponding maximum p values (pVal max) of the closest other products The device provides a positive identification for p values superioror equal to 005
Sample type Product (API) Number of signaturesValidation
pVal min ofvalidation samples
pVal max of closestother products
Tablets
Product 1 (71) 9 01404 00000
Product 2 (36) 9 05973 00105
Product 2 (72) 9 03576 00013
Product 3 (80 95) 18 05455 00000
Product 4 (67 75) 12 04137 00000
Product 5 (25 33) 18 03439 00171
Product 6 (61) 9 04847 00000
Product 7 (57) 9 04490 00007
Product 7 (56) 9 04106 00042
Product 8 (17) 9 04031 00083
Product 9 (3 6 12 30) 36 01292 00000
Product 10 (8) 9 05329 00044
Product 10 (4) 9 04481 00010
Product 11 (3) 9 06458 00129
Product 12 (75) 9 05785 00000
Product 13 (04 08 1 2 3) 54 04089 00145
Product 14 (61) 9 06009 00000
Product 15 (12) 9 04619 00167
Product 16 (51) 9 04299 00000
Product 17 (25 45) 27 04416 00000
Product 18 (2) 9 06275 00015
Product 19 (03 06 1) 24 01763 00009
Product 20 (24 32) 27 03453 00000
Product 21 (75) 6 05781 00000
Product 22 (70) 9 01562 00000
Product 23 (1 3 6) 36 04444 00348
Product 24 (41) 9 05872 00000
Product 25 (77 78) 18 05822 00000
Product 26 (08) 24 01807 00352
Capsules
Product 27 (83) 9 06223 00000
Product 28 (49) 9 05439 00000
Product 29 (76) 6 05920 00000
Product 30 (49) 9 01669 00000
Product 31 (83) 18 01218 00000
Product 32 (50) 9 01165 00000
Powders Product 33 (81) 30 06139 00000
6 Journal of Spectroscopy
fluorescence prevented the measurement of the signaturesAmong these six products two were tablets two werecapsules and two were lyophilized products Since nosignatures could be taken with these samples the resultsare not displayed in this paper
322 Second Validation with Counterfeits Placebos andGenerics All the tested counterfeits consisting in 44 samplesfailed for identification The obtained p values were verylowmdashinferior to 00003mdashwhich comforted the robustnessof the methods for the detection of counterfeits The detailsof the results of the second validation are presented inTable 2
Nine of the tested generics have the same formulationas the genuine brand products and their spectra are identicalas previously presented in Figure 4 These samples wereconsequently identified by the device as genuine Since theobjective of the methods is to reject counterfeits and genericswere only used to simulate them these results are notconsidered critical The other five generics were rejected bythe methods The p values obtained are highly dependenton the tested generic
Two of the three tested placebos were rejected by themethods The third one the placebo of product 26 wasdetectedby the instrumentas genuinewithamaximumpvalueof 05257 which is very high The method for product 26 alow-dosage product was then considered not specific enoughand was then deleted from the database
The results of the validation and the tests performed forthe second validation are summarized in Table 3
33 Robustness Results The results of the four robustnesstests are presented in Table 4
According to the first test a capsule of product 31 and atablet of product 20 were measured 20 times at the sameposition to check if the samples were damaged by the laserThe capsule and the tablet were correctly identified even after20 measurements Additionally no important variation ofthe p value range was observed which confirms that the laserdid not damage the samples At this point it should be notedthat no soft-gel capsules should be measured with the Ramaninstrument since their shell is very sensitive to the heat ofthe laser
A capsule of product 31 and a tablet of product 20 wereeach measured 10 times for the robustness test R2 They weremoved between each measurement in order to check if theposition of the sample had an influence on the results Thesamples were correctly identified during each measurementwith little difference in the p values This means that theposition of the samples has low influence on their identifica-tion However it is recommended to measure the capsules onthe side where there is only one layer of gelatin Also careshould be taken in order not to break the samples with thesample holder For instance if a tablet is broken and its coatingremoveddue to thebreaking thena slightly different spectrumcan be obtained which might alter the results
The robustness test R3 deals with the sampling that isavailable for the studied medicines Three products weremeasured either directly with the sample holder throughtransparent glass vials or through blisters depending onthe available packaging Two types of blister were evaluatedtransparent (plastic) and white (plastic) The samples wereall correctly authenticated when measured directly throughtransparent glass vials and through transparent blisters Theproducts could not be correctly identified through whiteblister It is consequently recommended to use the tablet
Table 2 Second validation results obtained with the challenging samples The number of samples measured for each product has been addedThe maximum p values (pVal) obtained for each counterfeit are presented in the table together with the corresponding passfail result
Sample set Counterfeited product Number of samples pVal max Identification against the corresponding method
Counterfeits
Product 27 2 00000 Fail
Product 6 6 00000 Fail
Product 28 3 00000 Fail
Product 26 1 00000 Fail
Product 19 5 00001 Fail
Product 23 10 00000 Fail
Product 25 7 00000 Fail
Product 32 10 00003 Fail
Placebos
Product 4 1 00000 Fail
Product 32 1 00040 Fail
Product 26 1 05257 Pass
Generics
Product 20 1 04937 Pass
Product 25 batch 1 2 00224 Fail
Product 25 batch 2 3 06326 Pass
Product 32 batch 1 1 00042 Fail
Product 32 batch 2 4 01548 Pass
Product 5 2 00110 Fail
Product 33 1 01295 Pass
7Journal of Spectroscopy
holder to measure the samples However it is interesting tonote that according to this test a product measured on thefield through transparent blister or in transparent glass vialsmight be correctly authenticated
In order to evaluate the human influence (test R4) a tabletof product 20 and a capsule of product 31 have beenmeasured three times by two different operators The sampleswere all correctly identified and the obtained p values are veryclose between the operators It seems that the operators haveno influence on the identification which is an importantfactor for the use on the field It should be noted that bothoperators were properly trained before the robustness test
34 Composition of the Counterfeits As soon as a counterfeithas been confirmed it is necessary to determine its chemicalcomposition in order to estimate the impact on the patientsrsquosafety In this frame the discovery mode of the instrumenthas been evaluated with the studied counterfeits This modeproposes in case of a ldquofailrdquo result the chemical compositionof the tested sample This composition is suggested according
to the database furnished by the supplier and integrated inthe device The database contains more than 8000 referencesof excipients and active substances and was completed by thestudied products during the calibration
Several samples per counterfeit seizures have been ana-lysed with the device and interpreted with the discoverymode Their chemical composition was previously deter-mined by the lab instruments a Raman a near-infraredand a nuclear magnetic resonance spectrometers
For 14 samples no match could be found with theinstrument database This can be explained by the factthat for these samples the chemical composition was amixture of at least three components part of the databasebut difficult to detect all with an automatic Raman data-base search
The second point is that for 10 counterfeits a differentcompositionwas proposed by the device between the differentsamples of the same seizure Once again it can be explainedby the complex mixture of the counterfeits Some compo-nents could be identified while others not For instance for
Table 4 Results of the four robustness tests The conditions of the test like the sampling have been added together with the number ofspectra measured The results consist in the p value range and the passfail answers
Robustness test Product ConditionsNumber ofspectra
pVal resultsIdentification against
the corresponding method
R1 degradationwith the laser
Product 20 No change in sample position 20 [05745ndash06669] Pass
Product 31 No change in sample position 20 [05616ndash07335] Pass
R2 influence ofthe sample position
Product 20 Sample position changed 10 [05523ndash06746] Pass
Product 31 Sample position changed 10 [03316ndash06481] Pass
R3 influence ofthe sampling
Product 20
Glass vial 3 [05386ndash05950] Pass
Transparent blister 3 [05891ndash06534] Pass
Direct 3 [05344ndash05746] Pass
Product 32
Glass vial 3 [01131ndash02471] Pass
Transparent blister 3 [00814ndash03385] Pass
Direct 3 [01628ndash04482] Pass
Product 27
White blister 3 [00175ndash00259] Fail
White blister 3 [00272ndash01164] Fail
White blister 3 [00017ndash00731] Fail
R4 influence ofthe operator
Product 20Operator A 3 [06815ndash06871] Pass
Operator B 3 [06143ndash06526] Pass
Product 31Operator A 3 [06159ndash06779] Pass
Operator B 3 [06599ndash07381] Pass
Table 3 Overview of the results obtained for the validation and the tests of the second validation (lowast) In these tests ldquofailrdquo is actually theexpected answer since these products should not be identified by the method
Step of the method development Sample set Results of the identification
Validation
Validation with positive samples(eg product 1 tested on the method of product 1)
100 pass
Validation against all other products(eg product 1 tested on the method of product 2)
100 faillowast
Second validation
Counterfeits 100 faillowast
Placebos 67 faillowast
Generics 35 faillowast
8 Journal of Spectroscopy
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
antibiotic and antimycobacterial medicines [42] A handheldRaman spectrometer was tested on 15 pharmaceutical prod-ucts by Bugay and Brush [43] and its specificity was evaluatedbyHajjou et al regarding the active pharmaceutical ingredient(API) content of six medicines [44] Kalyanaraman et alalso presented the general use of such a device for counterfeitdetection [45] Ricci et al compared the performance of ahandheld Raman instrument with a benchtop device anddesorption electrospray ionization (DESI) MS for the detec-tionof counterfeits of antimalarials [46]Theuseof ahandheldRaman device on the field in Nigeria was finally presented bySpink et al [47]
In this paper a handheld Raman spectrometer wasevaluated for the detection of counterfeits of a high numberof pharmaceutical medicines consisting in tablets capsulesand powders Robust methods were developed using a largedatabase of products In total there are several batches of33 product families and among them 62 formulations weremeasured including the different manufacturing sites of theproducts A detailed calibration and validation strategy wasproposed as well as some robustness tests an evaluation ofthe discovery mode results for the composition of the con-firmed counterfeits and a comparison with a lab instrument
2 Material and Methods
21 Instrument Characteristics and Settings A handheldRaman spectrometer was used to collect the spectra andperform the calibration The device is a direct dispersiveRamanusing a 785nm laser excitationwavelength Its spectralresolution varies between 8 and 105 cmminus1 in average and theRaman shift range is 250 to 2875 cmminus1 The integration timein automatic mode is of 12ms minimum
The tablet sample holder was used for the measurementof the tablets and capsules Some of the samples had to bemeasured directly against the cone of the Raman sincethey were too large for the holder While the tablets and thecapsules were not measured in their packaging the powderswere analysed through their original glass vial
22 Statistical Method The statistical method used for theidentification of the products was the one provided by themanufacturer The calibration consisted in taking spectraof the genuine products The device then computed auto-matically a ldquosignaturerdquo of the product with each spectrumrecorded Several signatures of the same product were thengathered into a method The signatures are based on a multi-variate test of equivalence This test is used to compare anewly acquired Raman spectrum and the reference spectraregistered in the database If the probabilistic value (p value)of the new spectrum is above or equal to 005 meaning thespectrum presents a match within the preset 95 percentconfidence limit the measurement is considered consistentwith the reference spectra and the device reports a ldquopassrdquoresult If the p value is inferior to 005 the device reports aldquofailrdquo result In the present method while the validation withthe genuine products should provide ldquopassrdquo results the coun-terfeits generics and placebos should provide a ldquofailrdquo result
The multivariate test of equivalence is computed using allthe channels across the detector array region and can beconsidered a multivariate version of a statistical t-test Theuncertainty is directly modeled by the instrument softwareand thus no modeling has to be performed by the user
23 Calibration and Validation Strategy The calibration ofthe methods consisted in taking several spectra of a productwhichwere automatically converted in signatures and gather-ing them into a single method One method per product wasideally created For products presenting different formula-tions between the dosages (different excipients) two or moremethods were created per product
The validation was performed with an independent set ofspectra The samples were tested on the methods in order tocheck if they were correctly recognized All tested sampleshad to deliver a ldquopassrdquo result with the appropriate method
Additionally the products were tested on all the othermethods to check that no mismatch could occur for theseproducts
A second validation was then performed using counter-feits placebos and generics in order to test the ability ofthe handheld Raman spectrometer to detect counterfeitsThe generics and the placebos were tested in order to mimicthe ldquobest case counterfeitsrdquo All these samples had to berejected (present a ldquofailrdquo result) by the instrument
24 Samples
241 Genuine Samples At least 5 batches per product weremeasured with the split of 3 batches for the calibration setand 2 batches for the validation set for each of the 62 studiedformulations of the 33 product families For the productsmanufactured in several sites more than 5 batches weremeasured in order to include more variability One sampleper batch and three spectra per sample were measured Thesamples were slightly moved (for the capsules and powders)or flipped (for the tablets) between each measurement 543signatures were acquired for the calibration while the valida-tion was performed with 392 spectra
242 Placebos Generics and Counterfeits 44 counterfeitsamples (from eight different seizures) were tested whichhad already been confirmed as counterfeits by previous anal-yses In order to enlarge the number of samples 3 placebosand 14 generics from 5 different batches were also testedagainst the methods On the whole 61 spectra were acquiredfor the second validation samples
25 Robustness Tests Once the methods were calibratedvalidated and tested against the counterfeits placebos andgenerics 4 additional tests were performed in order to checkthe robustness of the methods
The first robustness test (R1) consisted in measuring thesame sample 20 times at the same position in order to see ifthe laser degraded the sample Two samples (one tablet andone capsule) were tested
In the second robustness test (R2) two samples (onetablet and one capsule) were measured 10 times at differentpositions in the sample holder The objective was to evaluate
2 Journal of Spectroscopy
if the position of the measurement had an influence onthe result
The influence of the sampling was evaluated in the thirdrobustness test (R3) Three samples (one tablet and twocapsules) were measured either through a glass vial througha blister (transparent or white) or directly against the cone ofthe instrument
Finally the fourth robustness test (R4) consisted in mea-suring two samples (one tablet and one capsule) three timesby two operators with the aim of testing if the operatorshad an influence on the measurement
3 Results and Interpretation
31 Spectrum Interpretation The spectra obtained by thedevice were first evaluated Several challenges to the creationof the calibration could be pointed out
First of all it can be observed that some product fam-ilies present heterogeneous spectra Two or more dosagesof the same product can indeed generate different spectrawhen a different formulationmdashit means a different excipientprofilemdashwas used (see Figure 1) Additionally in rare casesthe formulation can differ for the same product and the samedosage depending on the manufacturing site of the prod-uct (see Figure 2) It is thus important to include all thedosages and if possible all the manufacturing sites of aproduct into the methods Different methods might thenhave to be created for one product if the spectra of theproduct family differ Products 19 and 23 are indeedconcerned by both the heterogeneity within the productfamily and the different formulations depending on themanufacturing site Depending on the dosage or themanufacturing site these products underwent changes inthe composition of the sugar excipients mainly goingfrom lactose to starch Since these excipients have a big
influence on the spectral pattern of these low-dosageproductsmdash03 06 and 1 API for product 19 and13 and 6 API for product 23mdashit might be difficultto perform the identification unless separate methods arecreated per formulation
Another challenge to the calibration is that similarexcipient profiles can be found between different productfamilies The resulting spectra will then be similar likethe ones presented in Figure 3 and thus the products willbe hard to differentiate This is the case for products 18and 23 which contain the same excipients and have lowdosage of APImdash2 for product 18 and 50 and 83 forproduct 23 However small spectral differences can beobserved between products 18 and 23 in the range of1500 and 1700 cmminus1 and are due to API bands of product18 The two other products concerned by the similarity ofexcipient patterns are products 31 and 32 which withtheir high content of APImdash83 and 50mdashshould how-ever be distinguishable
Some generics present formulations that are almostidentical to the ones of the brand products This will resultin similar spectra that are not possible to distinguish fromthe brand product like in the example in Figure 4 In thesecases a ldquopassrdquo result can be expected
Low-dosage products are difficult to analyse with Ramanspectroscopy since the API will be hardly observed in thespectrum of the medicine The limit of detection of API inRaman spectra is quite difficult to define especially becauseit depends on the excipient pattern of the medicine In thestudied products excipient patterns made of sugar likestarch cellulose or glucose are predominant in the spectraand might mask the API bands In a previous study a limitof detection of 06 was encountered while the authorsindicated that the limit of detection was usually locatedaround 1 or 2 of API [48] For this reason the placebos of
Product 19 XmgProduct 19 Ymg
3456789
10111213141516171819
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 1 Raman spectra of two dosages of the same product family (product 19) Due to differences in the formulation the product presentsheterogeneous spectra
3Journal of Spectroscopy
low-dosage products might provide spectra that are verysimilar to the genuine products This was confirmed forproduct 26 which with an API content of 06 and anexcipient pattern dominated by starch and lactose hasidentical spectra between the genuine product and itsplacebo
The quality of the spectra might also have an influence onthe calibration The measured spectra present a wide rangefrom 250 to 2875 cmminus1 The end of the range approximatelyfrom 1800 to 2875 cmminus1 is often noisy and not necessarilyuseful since it does not bring additional information aboutthe product Last but not least a few samples like product
32 underwent fluorescence (Figure 5) This phenomenonwas observed when special excipients were used in theformulation of either the tablet or the shell of the capsules Inthis study in which a laser wavelength of 765 nm was usedindigo carmine (for product 32) microcrystalline cellulosehydroxypropyl methylcellulose and croscarmellose werefound responsible for the fluorescence phenomenon Whilethe fluorescence made the identification harder most bandsof the API and the excipients could still be observed like inFigure 5 When measuring other products not presented inthis paper the fluorescence was however so strong that no sig-nature could even be taken and no method created
Product 19 Xmg manufacturing site AProduct 19 Xmg manufacturing site B
4
6
8
10
12
14
16
18
20
22
24
26
28
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 2 Raman spectra of two samples produced at different manufacturing sites and possessing the same dosage (product 19 Xmg) Due todifferences in the chosen formulation the product presents heterogeneous spectra
Product 23 XmgProduct 18
468
1012141618202224262830323436384042
500 1000 1500 2000 2500Raman shift (cm‒1)
Ram
an in
tens
ity
Figure 3 Raman spectra of two different products (product 18 and product 23 Xmg) The products present similar excipients profiles andthus close Raman spectra
4 Journal of Spectroscopy
32 Identification Results
321 Calibration and Validation Results There are 33product families and among them 62 formulations couldbe successfully measured and integrated in the database ofthe instrument In total 31 methods have been created fortablets 6 for capsules and one for powders Most methodsgather spectra of several dosages per product For the productfamilies that presented heterogeneous formulations severalmethods have been created
100 of the spectra from the validation set has beencorrectly identified according to the pVal limit for positive
identification (superior to 005) According to the manufac-turer a pVal lower than 01 is considered too low for a robustmethod For all the developed methods the minimum pValwas higher than 01 (see Table 1)
Additionally no confusion could be observed whentesting the products on all the other methods than theirdedicated method 100 of all other products were rejectedby the tested method Table 1 presents the minimum p valuesobtained for the validation samples of the correspondingmethod together with the maximum p values of the closestother products These values show that the methods weresuccessfully validated and are very robust
Figure 4 Raman spectra of a brand product and one of its generic (product 20 Zmg) Both samples present a similar formulation and thesame dosage and thus similar Raman spectra
Product 32
20
25
30
35
40
45
50
55
60
65
70
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 5 Raman spectrum of product 32 While a strong fluorescence phenomenon can be observed the characteristic bands of the productare still visible enabling a correct identification
5Journal of Spectroscopy
A selectivity test proposed by the device was evaluatedafter the validation This test consists in evaluating for eachmethod the risks of getting mismatches The option wastested for the products providing the lowest p values duringthe validation For product 26 for instance the selectivity testsuggested that the closest other products would deliver a pvalue of 00026 Tested physically during the validation theproduct actually delivered a p value of 00352 using the
method of product 26 While the selectivity test might bean option to see quickly if possible mismatches might occurthe results delivered are not real The validation performed inthis study is thus considered more completed than theselectivity test Raman can thus not be considered a universalmethod for counterfeit detection
For six other products not included in the 33 presentedproduct families no method could be created since the
Table 1 Validation results obtained with the genuine products The number of signatures measured for each product has been added Theminimum p values (pVal min) obtained for the validation samples of the corresponding method are presented in the table together with thecorresponding maximum p values (pVal max) of the closest other products The device provides a positive identification for p values superioror equal to 005
Sample type Product (API) Number of signaturesValidation
pVal min ofvalidation samples
pVal max of closestother products
Tablets
Product 1 (71) 9 01404 00000
Product 2 (36) 9 05973 00105
Product 2 (72) 9 03576 00013
Product 3 (80 95) 18 05455 00000
Product 4 (67 75) 12 04137 00000
Product 5 (25 33) 18 03439 00171
Product 6 (61) 9 04847 00000
Product 7 (57) 9 04490 00007
Product 7 (56) 9 04106 00042
Product 8 (17) 9 04031 00083
Product 9 (3 6 12 30) 36 01292 00000
Product 10 (8) 9 05329 00044
Product 10 (4) 9 04481 00010
Product 11 (3) 9 06458 00129
Product 12 (75) 9 05785 00000
Product 13 (04 08 1 2 3) 54 04089 00145
Product 14 (61) 9 06009 00000
Product 15 (12) 9 04619 00167
Product 16 (51) 9 04299 00000
Product 17 (25 45) 27 04416 00000
Product 18 (2) 9 06275 00015
Product 19 (03 06 1) 24 01763 00009
Product 20 (24 32) 27 03453 00000
Product 21 (75) 6 05781 00000
Product 22 (70) 9 01562 00000
Product 23 (1 3 6) 36 04444 00348
Product 24 (41) 9 05872 00000
Product 25 (77 78) 18 05822 00000
Product 26 (08) 24 01807 00352
Capsules
Product 27 (83) 9 06223 00000
Product 28 (49) 9 05439 00000
Product 29 (76) 6 05920 00000
Product 30 (49) 9 01669 00000
Product 31 (83) 18 01218 00000
Product 32 (50) 9 01165 00000
Powders Product 33 (81) 30 06139 00000
6 Journal of Spectroscopy
fluorescence prevented the measurement of the signaturesAmong these six products two were tablets two werecapsules and two were lyophilized products Since nosignatures could be taken with these samples the resultsare not displayed in this paper
322 Second Validation with Counterfeits Placebos andGenerics All the tested counterfeits consisting in 44 samplesfailed for identification The obtained p values were verylowmdashinferior to 00003mdashwhich comforted the robustnessof the methods for the detection of counterfeits The detailsof the results of the second validation are presented inTable 2
Nine of the tested generics have the same formulationas the genuine brand products and their spectra are identicalas previously presented in Figure 4 These samples wereconsequently identified by the device as genuine Since theobjective of the methods is to reject counterfeits and genericswere only used to simulate them these results are notconsidered critical The other five generics were rejected bythe methods The p values obtained are highly dependenton the tested generic
Two of the three tested placebos were rejected by themethods The third one the placebo of product 26 wasdetectedby the instrumentas genuinewithamaximumpvalueof 05257 which is very high The method for product 26 alow-dosage product was then considered not specific enoughand was then deleted from the database
The results of the validation and the tests performed forthe second validation are summarized in Table 3
33 Robustness Results The results of the four robustnesstests are presented in Table 4
According to the first test a capsule of product 31 and atablet of product 20 were measured 20 times at the sameposition to check if the samples were damaged by the laserThe capsule and the tablet were correctly identified even after20 measurements Additionally no important variation ofthe p value range was observed which confirms that the laserdid not damage the samples At this point it should be notedthat no soft-gel capsules should be measured with the Ramaninstrument since their shell is very sensitive to the heat ofthe laser
A capsule of product 31 and a tablet of product 20 wereeach measured 10 times for the robustness test R2 They weremoved between each measurement in order to check if theposition of the sample had an influence on the results Thesamples were correctly identified during each measurementwith little difference in the p values This means that theposition of the samples has low influence on their identifica-tion However it is recommended to measure the capsules onthe side where there is only one layer of gelatin Also careshould be taken in order not to break the samples with thesample holder For instance if a tablet is broken and its coatingremoveddue to thebreaking thena slightly different spectrumcan be obtained which might alter the results
The robustness test R3 deals with the sampling that isavailable for the studied medicines Three products weremeasured either directly with the sample holder throughtransparent glass vials or through blisters depending onthe available packaging Two types of blister were evaluatedtransparent (plastic) and white (plastic) The samples wereall correctly authenticated when measured directly throughtransparent glass vials and through transparent blisters Theproducts could not be correctly identified through whiteblister It is consequently recommended to use the tablet
Table 2 Second validation results obtained with the challenging samples The number of samples measured for each product has been addedThe maximum p values (pVal) obtained for each counterfeit are presented in the table together with the corresponding passfail result
Sample set Counterfeited product Number of samples pVal max Identification against the corresponding method
Counterfeits
Product 27 2 00000 Fail
Product 6 6 00000 Fail
Product 28 3 00000 Fail
Product 26 1 00000 Fail
Product 19 5 00001 Fail
Product 23 10 00000 Fail
Product 25 7 00000 Fail
Product 32 10 00003 Fail
Placebos
Product 4 1 00000 Fail
Product 32 1 00040 Fail
Product 26 1 05257 Pass
Generics
Product 20 1 04937 Pass
Product 25 batch 1 2 00224 Fail
Product 25 batch 2 3 06326 Pass
Product 32 batch 1 1 00042 Fail
Product 32 batch 2 4 01548 Pass
Product 5 2 00110 Fail
Product 33 1 01295 Pass
7Journal of Spectroscopy
holder to measure the samples However it is interesting tonote that according to this test a product measured on thefield through transparent blister or in transparent glass vialsmight be correctly authenticated
In order to evaluate the human influence (test R4) a tabletof product 20 and a capsule of product 31 have beenmeasured three times by two different operators The sampleswere all correctly identified and the obtained p values are veryclose between the operators It seems that the operators haveno influence on the identification which is an importantfactor for the use on the field It should be noted that bothoperators were properly trained before the robustness test
34 Composition of the Counterfeits As soon as a counterfeithas been confirmed it is necessary to determine its chemicalcomposition in order to estimate the impact on the patientsrsquosafety In this frame the discovery mode of the instrumenthas been evaluated with the studied counterfeits This modeproposes in case of a ldquofailrdquo result the chemical compositionof the tested sample This composition is suggested according
to the database furnished by the supplier and integrated inthe device The database contains more than 8000 referencesof excipients and active substances and was completed by thestudied products during the calibration
Several samples per counterfeit seizures have been ana-lysed with the device and interpreted with the discoverymode Their chemical composition was previously deter-mined by the lab instruments a Raman a near-infraredand a nuclear magnetic resonance spectrometers
For 14 samples no match could be found with theinstrument database This can be explained by the factthat for these samples the chemical composition was amixture of at least three components part of the databasebut difficult to detect all with an automatic Raman data-base search
The second point is that for 10 counterfeits a differentcompositionwas proposed by the device between the differentsamples of the same seizure Once again it can be explainedby the complex mixture of the counterfeits Some compo-nents could be identified while others not For instance for
Table 4 Results of the four robustness tests The conditions of the test like the sampling have been added together with the number ofspectra measured The results consist in the p value range and the passfail answers
Robustness test Product ConditionsNumber ofspectra
pVal resultsIdentification against
the corresponding method
R1 degradationwith the laser
Product 20 No change in sample position 20 [05745ndash06669] Pass
Product 31 No change in sample position 20 [05616ndash07335] Pass
R2 influence ofthe sample position
Product 20 Sample position changed 10 [05523ndash06746] Pass
Product 31 Sample position changed 10 [03316ndash06481] Pass
R3 influence ofthe sampling
Product 20
Glass vial 3 [05386ndash05950] Pass
Transparent blister 3 [05891ndash06534] Pass
Direct 3 [05344ndash05746] Pass
Product 32
Glass vial 3 [01131ndash02471] Pass
Transparent blister 3 [00814ndash03385] Pass
Direct 3 [01628ndash04482] Pass
Product 27
White blister 3 [00175ndash00259] Fail
White blister 3 [00272ndash01164] Fail
White blister 3 [00017ndash00731] Fail
R4 influence ofthe operator
Product 20Operator A 3 [06815ndash06871] Pass
Operator B 3 [06143ndash06526] Pass
Product 31Operator A 3 [06159ndash06779] Pass
Operator B 3 [06599ndash07381] Pass
Table 3 Overview of the results obtained for the validation and the tests of the second validation (lowast) In these tests ldquofailrdquo is actually theexpected answer since these products should not be identified by the method
Step of the method development Sample set Results of the identification
Validation
Validation with positive samples(eg product 1 tested on the method of product 1)
100 pass
Validation against all other products(eg product 1 tested on the method of product 2)
100 faillowast
Second validation
Counterfeits 100 faillowast
Placebos 67 faillowast
Generics 35 faillowast
8 Journal of Spectroscopy
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
if the position of the measurement had an influence onthe result
The influence of the sampling was evaluated in the thirdrobustness test (R3) Three samples (one tablet and twocapsules) were measured either through a glass vial througha blister (transparent or white) or directly against the cone ofthe instrument
Finally the fourth robustness test (R4) consisted in mea-suring two samples (one tablet and one capsule) three timesby two operators with the aim of testing if the operatorshad an influence on the measurement
3 Results and Interpretation
31 Spectrum Interpretation The spectra obtained by thedevice were first evaluated Several challenges to the creationof the calibration could be pointed out
First of all it can be observed that some product fam-ilies present heterogeneous spectra Two or more dosagesof the same product can indeed generate different spectrawhen a different formulationmdashit means a different excipientprofilemdashwas used (see Figure 1) Additionally in rare casesthe formulation can differ for the same product and the samedosage depending on the manufacturing site of the prod-uct (see Figure 2) It is thus important to include all thedosages and if possible all the manufacturing sites of aproduct into the methods Different methods might thenhave to be created for one product if the spectra of theproduct family differ Products 19 and 23 are indeedconcerned by both the heterogeneity within the productfamily and the different formulations depending on themanufacturing site Depending on the dosage or themanufacturing site these products underwent changes inthe composition of the sugar excipients mainly goingfrom lactose to starch Since these excipients have a big
influence on the spectral pattern of these low-dosageproductsmdash03 06 and 1 API for product 19 and13 and 6 API for product 23mdashit might be difficultto perform the identification unless separate methods arecreated per formulation
Another challenge to the calibration is that similarexcipient profiles can be found between different productfamilies The resulting spectra will then be similar likethe ones presented in Figure 3 and thus the products willbe hard to differentiate This is the case for products 18and 23 which contain the same excipients and have lowdosage of APImdash2 for product 18 and 50 and 83 forproduct 23 However small spectral differences can beobserved between products 18 and 23 in the range of1500 and 1700 cmminus1 and are due to API bands of product18 The two other products concerned by the similarity ofexcipient patterns are products 31 and 32 which withtheir high content of APImdash83 and 50mdashshould how-ever be distinguishable
Some generics present formulations that are almostidentical to the ones of the brand products This will resultin similar spectra that are not possible to distinguish fromthe brand product like in the example in Figure 4 In thesecases a ldquopassrdquo result can be expected
Low-dosage products are difficult to analyse with Ramanspectroscopy since the API will be hardly observed in thespectrum of the medicine The limit of detection of API inRaman spectra is quite difficult to define especially becauseit depends on the excipient pattern of the medicine In thestudied products excipient patterns made of sugar likestarch cellulose or glucose are predominant in the spectraand might mask the API bands In a previous study a limitof detection of 06 was encountered while the authorsindicated that the limit of detection was usually locatedaround 1 or 2 of API [48] For this reason the placebos of
Product 19 XmgProduct 19 Ymg
3456789
10111213141516171819
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 1 Raman spectra of two dosages of the same product family (product 19) Due to differences in the formulation the product presentsheterogeneous spectra
3Journal of Spectroscopy
low-dosage products might provide spectra that are verysimilar to the genuine products This was confirmed forproduct 26 which with an API content of 06 and anexcipient pattern dominated by starch and lactose hasidentical spectra between the genuine product and itsplacebo
The quality of the spectra might also have an influence onthe calibration The measured spectra present a wide rangefrom 250 to 2875 cmminus1 The end of the range approximatelyfrom 1800 to 2875 cmminus1 is often noisy and not necessarilyuseful since it does not bring additional information aboutthe product Last but not least a few samples like product
32 underwent fluorescence (Figure 5) This phenomenonwas observed when special excipients were used in theformulation of either the tablet or the shell of the capsules Inthis study in which a laser wavelength of 765 nm was usedindigo carmine (for product 32) microcrystalline cellulosehydroxypropyl methylcellulose and croscarmellose werefound responsible for the fluorescence phenomenon Whilethe fluorescence made the identification harder most bandsof the API and the excipients could still be observed like inFigure 5 When measuring other products not presented inthis paper the fluorescence was however so strong that no sig-nature could even be taken and no method created
Product 19 Xmg manufacturing site AProduct 19 Xmg manufacturing site B
4
6
8
10
12
14
16
18
20
22
24
26
28
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 2 Raman spectra of two samples produced at different manufacturing sites and possessing the same dosage (product 19 Xmg) Due todifferences in the chosen formulation the product presents heterogeneous spectra
Product 23 XmgProduct 18
468
1012141618202224262830323436384042
500 1000 1500 2000 2500Raman shift (cm‒1)
Ram
an in
tens
ity
Figure 3 Raman spectra of two different products (product 18 and product 23 Xmg) The products present similar excipients profiles andthus close Raman spectra
4 Journal of Spectroscopy
32 Identification Results
321 Calibration and Validation Results There are 33product families and among them 62 formulations couldbe successfully measured and integrated in the database ofthe instrument In total 31 methods have been created fortablets 6 for capsules and one for powders Most methodsgather spectra of several dosages per product For the productfamilies that presented heterogeneous formulations severalmethods have been created
100 of the spectra from the validation set has beencorrectly identified according to the pVal limit for positive
identification (superior to 005) According to the manufac-turer a pVal lower than 01 is considered too low for a robustmethod For all the developed methods the minimum pValwas higher than 01 (see Table 1)
Additionally no confusion could be observed whentesting the products on all the other methods than theirdedicated method 100 of all other products were rejectedby the tested method Table 1 presents the minimum p valuesobtained for the validation samples of the correspondingmethod together with the maximum p values of the closestother products These values show that the methods weresuccessfully validated and are very robust
Figure 4 Raman spectra of a brand product and one of its generic (product 20 Zmg) Both samples present a similar formulation and thesame dosage and thus similar Raman spectra
Product 32
20
25
30
35
40
45
50
55
60
65
70
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 5 Raman spectrum of product 32 While a strong fluorescence phenomenon can be observed the characteristic bands of the productare still visible enabling a correct identification
5Journal of Spectroscopy
A selectivity test proposed by the device was evaluatedafter the validation This test consists in evaluating for eachmethod the risks of getting mismatches The option wastested for the products providing the lowest p values duringthe validation For product 26 for instance the selectivity testsuggested that the closest other products would deliver a pvalue of 00026 Tested physically during the validation theproduct actually delivered a p value of 00352 using the
method of product 26 While the selectivity test might bean option to see quickly if possible mismatches might occurthe results delivered are not real The validation performed inthis study is thus considered more completed than theselectivity test Raman can thus not be considered a universalmethod for counterfeit detection
For six other products not included in the 33 presentedproduct families no method could be created since the
Table 1 Validation results obtained with the genuine products The number of signatures measured for each product has been added Theminimum p values (pVal min) obtained for the validation samples of the corresponding method are presented in the table together with thecorresponding maximum p values (pVal max) of the closest other products The device provides a positive identification for p values superioror equal to 005
Sample type Product (API) Number of signaturesValidation
pVal min ofvalidation samples
pVal max of closestother products
Tablets
Product 1 (71) 9 01404 00000
Product 2 (36) 9 05973 00105
Product 2 (72) 9 03576 00013
Product 3 (80 95) 18 05455 00000
Product 4 (67 75) 12 04137 00000
Product 5 (25 33) 18 03439 00171
Product 6 (61) 9 04847 00000
Product 7 (57) 9 04490 00007
Product 7 (56) 9 04106 00042
Product 8 (17) 9 04031 00083
Product 9 (3 6 12 30) 36 01292 00000
Product 10 (8) 9 05329 00044
Product 10 (4) 9 04481 00010
Product 11 (3) 9 06458 00129
Product 12 (75) 9 05785 00000
Product 13 (04 08 1 2 3) 54 04089 00145
Product 14 (61) 9 06009 00000
Product 15 (12) 9 04619 00167
Product 16 (51) 9 04299 00000
Product 17 (25 45) 27 04416 00000
Product 18 (2) 9 06275 00015
Product 19 (03 06 1) 24 01763 00009
Product 20 (24 32) 27 03453 00000
Product 21 (75) 6 05781 00000
Product 22 (70) 9 01562 00000
Product 23 (1 3 6) 36 04444 00348
Product 24 (41) 9 05872 00000
Product 25 (77 78) 18 05822 00000
Product 26 (08) 24 01807 00352
Capsules
Product 27 (83) 9 06223 00000
Product 28 (49) 9 05439 00000
Product 29 (76) 6 05920 00000
Product 30 (49) 9 01669 00000
Product 31 (83) 18 01218 00000
Product 32 (50) 9 01165 00000
Powders Product 33 (81) 30 06139 00000
6 Journal of Spectroscopy
fluorescence prevented the measurement of the signaturesAmong these six products two were tablets two werecapsules and two were lyophilized products Since nosignatures could be taken with these samples the resultsare not displayed in this paper
322 Second Validation with Counterfeits Placebos andGenerics All the tested counterfeits consisting in 44 samplesfailed for identification The obtained p values were verylowmdashinferior to 00003mdashwhich comforted the robustnessof the methods for the detection of counterfeits The detailsof the results of the second validation are presented inTable 2
Nine of the tested generics have the same formulationas the genuine brand products and their spectra are identicalas previously presented in Figure 4 These samples wereconsequently identified by the device as genuine Since theobjective of the methods is to reject counterfeits and genericswere only used to simulate them these results are notconsidered critical The other five generics were rejected bythe methods The p values obtained are highly dependenton the tested generic
Two of the three tested placebos were rejected by themethods The third one the placebo of product 26 wasdetectedby the instrumentas genuinewithamaximumpvalueof 05257 which is very high The method for product 26 alow-dosage product was then considered not specific enoughand was then deleted from the database
The results of the validation and the tests performed forthe second validation are summarized in Table 3
33 Robustness Results The results of the four robustnesstests are presented in Table 4
According to the first test a capsule of product 31 and atablet of product 20 were measured 20 times at the sameposition to check if the samples were damaged by the laserThe capsule and the tablet were correctly identified even after20 measurements Additionally no important variation ofthe p value range was observed which confirms that the laserdid not damage the samples At this point it should be notedthat no soft-gel capsules should be measured with the Ramaninstrument since their shell is very sensitive to the heat ofthe laser
A capsule of product 31 and a tablet of product 20 wereeach measured 10 times for the robustness test R2 They weremoved between each measurement in order to check if theposition of the sample had an influence on the results Thesamples were correctly identified during each measurementwith little difference in the p values This means that theposition of the samples has low influence on their identifica-tion However it is recommended to measure the capsules onthe side where there is only one layer of gelatin Also careshould be taken in order not to break the samples with thesample holder For instance if a tablet is broken and its coatingremoveddue to thebreaking thena slightly different spectrumcan be obtained which might alter the results
The robustness test R3 deals with the sampling that isavailable for the studied medicines Three products weremeasured either directly with the sample holder throughtransparent glass vials or through blisters depending onthe available packaging Two types of blister were evaluatedtransparent (plastic) and white (plastic) The samples wereall correctly authenticated when measured directly throughtransparent glass vials and through transparent blisters Theproducts could not be correctly identified through whiteblister It is consequently recommended to use the tablet
Table 2 Second validation results obtained with the challenging samples The number of samples measured for each product has been addedThe maximum p values (pVal) obtained for each counterfeit are presented in the table together with the corresponding passfail result
Sample set Counterfeited product Number of samples pVal max Identification against the corresponding method
Counterfeits
Product 27 2 00000 Fail
Product 6 6 00000 Fail
Product 28 3 00000 Fail
Product 26 1 00000 Fail
Product 19 5 00001 Fail
Product 23 10 00000 Fail
Product 25 7 00000 Fail
Product 32 10 00003 Fail
Placebos
Product 4 1 00000 Fail
Product 32 1 00040 Fail
Product 26 1 05257 Pass
Generics
Product 20 1 04937 Pass
Product 25 batch 1 2 00224 Fail
Product 25 batch 2 3 06326 Pass
Product 32 batch 1 1 00042 Fail
Product 32 batch 2 4 01548 Pass
Product 5 2 00110 Fail
Product 33 1 01295 Pass
7Journal of Spectroscopy
holder to measure the samples However it is interesting tonote that according to this test a product measured on thefield through transparent blister or in transparent glass vialsmight be correctly authenticated
In order to evaluate the human influence (test R4) a tabletof product 20 and a capsule of product 31 have beenmeasured three times by two different operators The sampleswere all correctly identified and the obtained p values are veryclose between the operators It seems that the operators haveno influence on the identification which is an importantfactor for the use on the field It should be noted that bothoperators were properly trained before the robustness test
34 Composition of the Counterfeits As soon as a counterfeithas been confirmed it is necessary to determine its chemicalcomposition in order to estimate the impact on the patientsrsquosafety In this frame the discovery mode of the instrumenthas been evaluated with the studied counterfeits This modeproposes in case of a ldquofailrdquo result the chemical compositionof the tested sample This composition is suggested according
to the database furnished by the supplier and integrated inthe device The database contains more than 8000 referencesof excipients and active substances and was completed by thestudied products during the calibration
Several samples per counterfeit seizures have been ana-lysed with the device and interpreted with the discoverymode Their chemical composition was previously deter-mined by the lab instruments a Raman a near-infraredand a nuclear magnetic resonance spectrometers
For 14 samples no match could be found with theinstrument database This can be explained by the factthat for these samples the chemical composition was amixture of at least three components part of the databasebut difficult to detect all with an automatic Raman data-base search
The second point is that for 10 counterfeits a differentcompositionwas proposed by the device between the differentsamples of the same seizure Once again it can be explainedby the complex mixture of the counterfeits Some compo-nents could be identified while others not For instance for
Table 4 Results of the four robustness tests The conditions of the test like the sampling have been added together with the number ofspectra measured The results consist in the p value range and the passfail answers
Robustness test Product ConditionsNumber ofspectra
pVal resultsIdentification against
the corresponding method
R1 degradationwith the laser
Product 20 No change in sample position 20 [05745ndash06669] Pass
Product 31 No change in sample position 20 [05616ndash07335] Pass
R2 influence ofthe sample position
Product 20 Sample position changed 10 [05523ndash06746] Pass
Product 31 Sample position changed 10 [03316ndash06481] Pass
R3 influence ofthe sampling
Product 20
Glass vial 3 [05386ndash05950] Pass
Transparent blister 3 [05891ndash06534] Pass
Direct 3 [05344ndash05746] Pass
Product 32
Glass vial 3 [01131ndash02471] Pass
Transparent blister 3 [00814ndash03385] Pass
Direct 3 [01628ndash04482] Pass
Product 27
White blister 3 [00175ndash00259] Fail
White blister 3 [00272ndash01164] Fail
White blister 3 [00017ndash00731] Fail
R4 influence ofthe operator
Product 20Operator A 3 [06815ndash06871] Pass
Operator B 3 [06143ndash06526] Pass
Product 31Operator A 3 [06159ndash06779] Pass
Operator B 3 [06599ndash07381] Pass
Table 3 Overview of the results obtained for the validation and the tests of the second validation (lowast) In these tests ldquofailrdquo is actually theexpected answer since these products should not be identified by the method
Step of the method development Sample set Results of the identification
Validation
Validation with positive samples(eg product 1 tested on the method of product 1)
100 pass
Validation against all other products(eg product 1 tested on the method of product 2)
100 faillowast
Second validation
Counterfeits 100 faillowast
Placebos 67 faillowast
Generics 35 faillowast
8 Journal of Spectroscopy
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
low-dosage products might provide spectra that are verysimilar to the genuine products This was confirmed forproduct 26 which with an API content of 06 and anexcipient pattern dominated by starch and lactose hasidentical spectra between the genuine product and itsplacebo
The quality of the spectra might also have an influence onthe calibration The measured spectra present a wide rangefrom 250 to 2875 cmminus1 The end of the range approximatelyfrom 1800 to 2875 cmminus1 is often noisy and not necessarilyuseful since it does not bring additional information aboutthe product Last but not least a few samples like product
32 underwent fluorescence (Figure 5) This phenomenonwas observed when special excipients were used in theformulation of either the tablet or the shell of the capsules Inthis study in which a laser wavelength of 765 nm was usedindigo carmine (for product 32) microcrystalline cellulosehydroxypropyl methylcellulose and croscarmellose werefound responsible for the fluorescence phenomenon Whilethe fluorescence made the identification harder most bandsof the API and the excipients could still be observed like inFigure 5 When measuring other products not presented inthis paper the fluorescence was however so strong that no sig-nature could even be taken and no method created
Product 19 Xmg manufacturing site AProduct 19 Xmg manufacturing site B
4
6
8
10
12
14
16
18
20
22
24
26
28
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 2 Raman spectra of two samples produced at different manufacturing sites and possessing the same dosage (product 19 Xmg) Due todifferences in the chosen formulation the product presents heterogeneous spectra
Product 23 XmgProduct 18
468
1012141618202224262830323436384042
500 1000 1500 2000 2500Raman shift (cm‒1)
Ram
an in
tens
ity
Figure 3 Raman spectra of two different products (product 18 and product 23 Xmg) The products present similar excipients profiles andthus close Raman spectra
4 Journal of Spectroscopy
32 Identification Results
321 Calibration and Validation Results There are 33product families and among them 62 formulations couldbe successfully measured and integrated in the database ofthe instrument In total 31 methods have been created fortablets 6 for capsules and one for powders Most methodsgather spectra of several dosages per product For the productfamilies that presented heterogeneous formulations severalmethods have been created
100 of the spectra from the validation set has beencorrectly identified according to the pVal limit for positive
identification (superior to 005) According to the manufac-turer a pVal lower than 01 is considered too low for a robustmethod For all the developed methods the minimum pValwas higher than 01 (see Table 1)
Additionally no confusion could be observed whentesting the products on all the other methods than theirdedicated method 100 of all other products were rejectedby the tested method Table 1 presents the minimum p valuesobtained for the validation samples of the correspondingmethod together with the maximum p values of the closestother products These values show that the methods weresuccessfully validated and are very robust
Figure 4 Raman spectra of a brand product and one of its generic (product 20 Zmg) Both samples present a similar formulation and thesame dosage and thus similar Raman spectra
Product 32
20
25
30
35
40
45
50
55
60
65
70
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 5 Raman spectrum of product 32 While a strong fluorescence phenomenon can be observed the characteristic bands of the productare still visible enabling a correct identification
5Journal of Spectroscopy
A selectivity test proposed by the device was evaluatedafter the validation This test consists in evaluating for eachmethod the risks of getting mismatches The option wastested for the products providing the lowest p values duringthe validation For product 26 for instance the selectivity testsuggested that the closest other products would deliver a pvalue of 00026 Tested physically during the validation theproduct actually delivered a p value of 00352 using the
method of product 26 While the selectivity test might bean option to see quickly if possible mismatches might occurthe results delivered are not real The validation performed inthis study is thus considered more completed than theselectivity test Raman can thus not be considered a universalmethod for counterfeit detection
For six other products not included in the 33 presentedproduct families no method could be created since the
Table 1 Validation results obtained with the genuine products The number of signatures measured for each product has been added Theminimum p values (pVal min) obtained for the validation samples of the corresponding method are presented in the table together with thecorresponding maximum p values (pVal max) of the closest other products The device provides a positive identification for p values superioror equal to 005
Sample type Product (API) Number of signaturesValidation
pVal min ofvalidation samples
pVal max of closestother products
Tablets
Product 1 (71) 9 01404 00000
Product 2 (36) 9 05973 00105
Product 2 (72) 9 03576 00013
Product 3 (80 95) 18 05455 00000
Product 4 (67 75) 12 04137 00000
Product 5 (25 33) 18 03439 00171
Product 6 (61) 9 04847 00000
Product 7 (57) 9 04490 00007
Product 7 (56) 9 04106 00042
Product 8 (17) 9 04031 00083
Product 9 (3 6 12 30) 36 01292 00000
Product 10 (8) 9 05329 00044
Product 10 (4) 9 04481 00010
Product 11 (3) 9 06458 00129
Product 12 (75) 9 05785 00000
Product 13 (04 08 1 2 3) 54 04089 00145
Product 14 (61) 9 06009 00000
Product 15 (12) 9 04619 00167
Product 16 (51) 9 04299 00000
Product 17 (25 45) 27 04416 00000
Product 18 (2) 9 06275 00015
Product 19 (03 06 1) 24 01763 00009
Product 20 (24 32) 27 03453 00000
Product 21 (75) 6 05781 00000
Product 22 (70) 9 01562 00000
Product 23 (1 3 6) 36 04444 00348
Product 24 (41) 9 05872 00000
Product 25 (77 78) 18 05822 00000
Product 26 (08) 24 01807 00352
Capsules
Product 27 (83) 9 06223 00000
Product 28 (49) 9 05439 00000
Product 29 (76) 6 05920 00000
Product 30 (49) 9 01669 00000
Product 31 (83) 18 01218 00000
Product 32 (50) 9 01165 00000
Powders Product 33 (81) 30 06139 00000
6 Journal of Spectroscopy
fluorescence prevented the measurement of the signaturesAmong these six products two were tablets two werecapsules and two were lyophilized products Since nosignatures could be taken with these samples the resultsare not displayed in this paper
322 Second Validation with Counterfeits Placebos andGenerics All the tested counterfeits consisting in 44 samplesfailed for identification The obtained p values were verylowmdashinferior to 00003mdashwhich comforted the robustnessof the methods for the detection of counterfeits The detailsof the results of the second validation are presented inTable 2
Nine of the tested generics have the same formulationas the genuine brand products and their spectra are identicalas previously presented in Figure 4 These samples wereconsequently identified by the device as genuine Since theobjective of the methods is to reject counterfeits and genericswere only used to simulate them these results are notconsidered critical The other five generics were rejected bythe methods The p values obtained are highly dependenton the tested generic
Two of the three tested placebos were rejected by themethods The third one the placebo of product 26 wasdetectedby the instrumentas genuinewithamaximumpvalueof 05257 which is very high The method for product 26 alow-dosage product was then considered not specific enoughand was then deleted from the database
The results of the validation and the tests performed forthe second validation are summarized in Table 3
33 Robustness Results The results of the four robustnesstests are presented in Table 4
According to the first test a capsule of product 31 and atablet of product 20 were measured 20 times at the sameposition to check if the samples were damaged by the laserThe capsule and the tablet were correctly identified even after20 measurements Additionally no important variation ofthe p value range was observed which confirms that the laserdid not damage the samples At this point it should be notedthat no soft-gel capsules should be measured with the Ramaninstrument since their shell is very sensitive to the heat ofthe laser
A capsule of product 31 and a tablet of product 20 wereeach measured 10 times for the robustness test R2 They weremoved between each measurement in order to check if theposition of the sample had an influence on the results Thesamples were correctly identified during each measurementwith little difference in the p values This means that theposition of the samples has low influence on their identifica-tion However it is recommended to measure the capsules onthe side where there is only one layer of gelatin Also careshould be taken in order not to break the samples with thesample holder For instance if a tablet is broken and its coatingremoveddue to thebreaking thena slightly different spectrumcan be obtained which might alter the results
The robustness test R3 deals with the sampling that isavailable for the studied medicines Three products weremeasured either directly with the sample holder throughtransparent glass vials or through blisters depending onthe available packaging Two types of blister were evaluatedtransparent (plastic) and white (plastic) The samples wereall correctly authenticated when measured directly throughtransparent glass vials and through transparent blisters Theproducts could not be correctly identified through whiteblister It is consequently recommended to use the tablet
Table 2 Second validation results obtained with the challenging samples The number of samples measured for each product has been addedThe maximum p values (pVal) obtained for each counterfeit are presented in the table together with the corresponding passfail result
Sample set Counterfeited product Number of samples pVal max Identification against the corresponding method
Counterfeits
Product 27 2 00000 Fail
Product 6 6 00000 Fail
Product 28 3 00000 Fail
Product 26 1 00000 Fail
Product 19 5 00001 Fail
Product 23 10 00000 Fail
Product 25 7 00000 Fail
Product 32 10 00003 Fail
Placebos
Product 4 1 00000 Fail
Product 32 1 00040 Fail
Product 26 1 05257 Pass
Generics
Product 20 1 04937 Pass
Product 25 batch 1 2 00224 Fail
Product 25 batch 2 3 06326 Pass
Product 32 batch 1 1 00042 Fail
Product 32 batch 2 4 01548 Pass
Product 5 2 00110 Fail
Product 33 1 01295 Pass
7Journal of Spectroscopy
holder to measure the samples However it is interesting tonote that according to this test a product measured on thefield through transparent blister or in transparent glass vialsmight be correctly authenticated
In order to evaluate the human influence (test R4) a tabletof product 20 and a capsule of product 31 have beenmeasured three times by two different operators The sampleswere all correctly identified and the obtained p values are veryclose between the operators It seems that the operators haveno influence on the identification which is an importantfactor for the use on the field It should be noted that bothoperators were properly trained before the robustness test
34 Composition of the Counterfeits As soon as a counterfeithas been confirmed it is necessary to determine its chemicalcomposition in order to estimate the impact on the patientsrsquosafety In this frame the discovery mode of the instrumenthas been evaluated with the studied counterfeits This modeproposes in case of a ldquofailrdquo result the chemical compositionof the tested sample This composition is suggested according
to the database furnished by the supplier and integrated inthe device The database contains more than 8000 referencesof excipients and active substances and was completed by thestudied products during the calibration
Several samples per counterfeit seizures have been ana-lysed with the device and interpreted with the discoverymode Their chemical composition was previously deter-mined by the lab instruments a Raman a near-infraredand a nuclear magnetic resonance spectrometers
For 14 samples no match could be found with theinstrument database This can be explained by the factthat for these samples the chemical composition was amixture of at least three components part of the databasebut difficult to detect all with an automatic Raman data-base search
The second point is that for 10 counterfeits a differentcompositionwas proposed by the device between the differentsamples of the same seizure Once again it can be explainedby the complex mixture of the counterfeits Some compo-nents could be identified while others not For instance for
Table 4 Results of the four robustness tests The conditions of the test like the sampling have been added together with the number ofspectra measured The results consist in the p value range and the passfail answers
Robustness test Product ConditionsNumber ofspectra
pVal resultsIdentification against
the corresponding method
R1 degradationwith the laser
Product 20 No change in sample position 20 [05745ndash06669] Pass
Product 31 No change in sample position 20 [05616ndash07335] Pass
R2 influence ofthe sample position
Product 20 Sample position changed 10 [05523ndash06746] Pass
Product 31 Sample position changed 10 [03316ndash06481] Pass
R3 influence ofthe sampling
Product 20
Glass vial 3 [05386ndash05950] Pass
Transparent blister 3 [05891ndash06534] Pass
Direct 3 [05344ndash05746] Pass
Product 32
Glass vial 3 [01131ndash02471] Pass
Transparent blister 3 [00814ndash03385] Pass
Direct 3 [01628ndash04482] Pass
Product 27
White blister 3 [00175ndash00259] Fail
White blister 3 [00272ndash01164] Fail
White blister 3 [00017ndash00731] Fail
R4 influence ofthe operator
Product 20Operator A 3 [06815ndash06871] Pass
Operator B 3 [06143ndash06526] Pass
Product 31Operator A 3 [06159ndash06779] Pass
Operator B 3 [06599ndash07381] Pass
Table 3 Overview of the results obtained for the validation and the tests of the second validation (lowast) In these tests ldquofailrdquo is actually theexpected answer since these products should not be identified by the method
Step of the method development Sample set Results of the identification
Validation
Validation with positive samples(eg product 1 tested on the method of product 1)
100 pass
Validation against all other products(eg product 1 tested on the method of product 2)
100 faillowast
Second validation
Counterfeits 100 faillowast
Placebos 67 faillowast
Generics 35 faillowast
8 Journal of Spectroscopy
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
321 Calibration and Validation Results There are 33product families and among them 62 formulations couldbe successfully measured and integrated in the database ofthe instrument In total 31 methods have been created fortablets 6 for capsules and one for powders Most methodsgather spectra of several dosages per product For the productfamilies that presented heterogeneous formulations severalmethods have been created
100 of the spectra from the validation set has beencorrectly identified according to the pVal limit for positive
identification (superior to 005) According to the manufac-turer a pVal lower than 01 is considered too low for a robustmethod For all the developed methods the minimum pValwas higher than 01 (see Table 1)
Additionally no confusion could be observed whentesting the products on all the other methods than theirdedicated method 100 of all other products were rejectedby the tested method Table 1 presents the minimum p valuesobtained for the validation samples of the correspondingmethod together with the maximum p values of the closestother products These values show that the methods weresuccessfully validated and are very robust
Figure 4 Raman spectra of a brand product and one of its generic (product 20 Zmg) Both samples present a similar formulation and thesame dosage and thus similar Raman spectra
Product 32
20
25
30
35
40
45
50
55
60
65
70
500 1000 1500 2000 2500Raman shi (cm‒1)
Ram
an in
tens
ity
Figure 5 Raman spectrum of product 32 While a strong fluorescence phenomenon can be observed the characteristic bands of the productare still visible enabling a correct identification
5Journal of Spectroscopy
A selectivity test proposed by the device was evaluatedafter the validation This test consists in evaluating for eachmethod the risks of getting mismatches The option wastested for the products providing the lowest p values duringthe validation For product 26 for instance the selectivity testsuggested that the closest other products would deliver a pvalue of 00026 Tested physically during the validation theproduct actually delivered a p value of 00352 using the
method of product 26 While the selectivity test might bean option to see quickly if possible mismatches might occurthe results delivered are not real The validation performed inthis study is thus considered more completed than theselectivity test Raman can thus not be considered a universalmethod for counterfeit detection
For six other products not included in the 33 presentedproduct families no method could be created since the
Table 1 Validation results obtained with the genuine products The number of signatures measured for each product has been added Theminimum p values (pVal min) obtained for the validation samples of the corresponding method are presented in the table together with thecorresponding maximum p values (pVal max) of the closest other products The device provides a positive identification for p values superioror equal to 005
Sample type Product (API) Number of signaturesValidation
pVal min ofvalidation samples
pVal max of closestother products
Tablets
Product 1 (71) 9 01404 00000
Product 2 (36) 9 05973 00105
Product 2 (72) 9 03576 00013
Product 3 (80 95) 18 05455 00000
Product 4 (67 75) 12 04137 00000
Product 5 (25 33) 18 03439 00171
Product 6 (61) 9 04847 00000
Product 7 (57) 9 04490 00007
Product 7 (56) 9 04106 00042
Product 8 (17) 9 04031 00083
Product 9 (3 6 12 30) 36 01292 00000
Product 10 (8) 9 05329 00044
Product 10 (4) 9 04481 00010
Product 11 (3) 9 06458 00129
Product 12 (75) 9 05785 00000
Product 13 (04 08 1 2 3) 54 04089 00145
Product 14 (61) 9 06009 00000
Product 15 (12) 9 04619 00167
Product 16 (51) 9 04299 00000
Product 17 (25 45) 27 04416 00000
Product 18 (2) 9 06275 00015
Product 19 (03 06 1) 24 01763 00009
Product 20 (24 32) 27 03453 00000
Product 21 (75) 6 05781 00000
Product 22 (70) 9 01562 00000
Product 23 (1 3 6) 36 04444 00348
Product 24 (41) 9 05872 00000
Product 25 (77 78) 18 05822 00000
Product 26 (08) 24 01807 00352
Capsules
Product 27 (83) 9 06223 00000
Product 28 (49) 9 05439 00000
Product 29 (76) 6 05920 00000
Product 30 (49) 9 01669 00000
Product 31 (83) 18 01218 00000
Product 32 (50) 9 01165 00000
Powders Product 33 (81) 30 06139 00000
6 Journal of Spectroscopy
fluorescence prevented the measurement of the signaturesAmong these six products two were tablets two werecapsules and two were lyophilized products Since nosignatures could be taken with these samples the resultsare not displayed in this paper
322 Second Validation with Counterfeits Placebos andGenerics All the tested counterfeits consisting in 44 samplesfailed for identification The obtained p values were verylowmdashinferior to 00003mdashwhich comforted the robustnessof the methods for the detection of counterfeits The detailsof the results of the second validation are presented inTable 2
Nine of the tested generics have the same formulationas the genuine brand products and their spectra are identicalas previously presented in Figure 4 These samples wereconsequently identified by the device as genuine Since theobjective of the methods is to reject counterfeits and genericswere only used to simulate them these results are notconsidered critical The other five generics were rejected bythe methods The p values obtained are highly dependenton the tested generic
Two of the three tested placebos were rejected by themethods The third one the placebo of product 26 wasdetectedby the instrumentas genuinewithamaximumpvalueof 05257 which is very high The method for product 26 alow-dosage product was then considered not specific enoughand was then deleted from the database
The results of the validation and the tests performed forthe second validation are summarized in Table 3
33 Robustness Results The results of the four robustnesstests are presented in Table 4
According to the first test a capsule of product 31 and atablet of product 20 were measured 20 times at the sameposition to check if the samples were damaged by the laserThe capsule and the tablet were correctly identified even after20 measurements Additionally no important variation ofthe p value range was observed which confirms that the laserdid not damage the samples At this point it should be notedthat no soft-gel capsules should be measured with the Ramaninstrument since their shell is very sensitive to the heat ofthe laser
A capsule of product 31 and a tablet of product 20 wereeach measured 10 times for the robustness test R2 They weremoved between each measurement in order to check if theposition of the sample had an influence on the results Thesamples were correctly identified during each measurementwith little difference in the p values This means that theposition of the samples has low influence on their identifica-tion However it is recommended to measure the capsules onthe side where there is only one layer of gelatin Also careshould be taken in order not to break the samples with thesample holder For instance if a tablet is broken and its coatingremoveddue to thebreaking thena slightly different spectrumcan be obtained which might alter the results
The robustness test R3 deals with the sampling that isavailable for the studied medicines Three products weremeasured either directly with the sample holder throughtransparent glass vials or through blisters depending onthe available packaging Two types of blister were evaluatedtransparent (plastic) and white (plastic) The samples wereall correctly authenticated when measured directly throughtransparent glass vials and through transparent blisters Theproducts could not be correctly identified through whiteblister It is consequently recommended to use the tablet
Table 2 Second validation results obtained with the challenging samples The number of samples measured for each product has been addedThe maximum p values (pVal) obtained for each counterfeit are presented in the table together with the corresponding passfail result
Sample set Counterfeited product Number of samples pVal max Identification against the corresponding method
Counterfeits
Product 27 2 00000 Fail
Product 6 6 00000 Fail
Product 28 3 00000 Fail
Product 26 1 00000 Fail
Product 19 5 00001 Fail
Product 23 10 00000 Fail
Product 25 7 00000 Fail
Product 32 10 00003 Fail
Placebos
Product 4 1 00000 Fail
Product 32 1 00040 Fail
Product 26 1 05257 Pass
Generics
Product 20 1 04937 Pass
Product 25 batch 1 2 00224 Fail
Product 25 batch 2 3 06326 Pass
Product 32 batch 1 1 00042 Fail
Product 32 batch 2 4 01548 Pass
Product 5 2 00110 Fail
Product 33 1 01295 Pass
7Journal of Spectroscopy
holder to measure the samples However it is interesting tonote that according to this test a product measured on thefield through transparent blister or in transparent glass vialsmight be correctly authenticated
In order to evaluate the human influence (test R4) a tabletof product 20 and a capsule of product 31 have beenmeasured three times by two different operators The sampleswere all correctly identified and the obtained p values are veryclose between the operators It seems that the operators haveno influence on the identification which is an importantfactor for the use on the field It should be noted that bothoperators were properly trained before the robustness test
34 Composition of the Counterfeits As soon as a counterfeithas been confirmed it is necessary to determine its chemicalcomposition in order to estimate the impact on the patientsrsquosafety In this frame the discovery mode of the instrumenthas been evaluated with the studied counterfeits This modeproposes in case of a ldquofailrdquo result the chemical compositionof the tested sample This composition is suggested according
to the database furnished by the supplier and integrated inthe device The database contains more than 8000 referencesof excipients and active substances and was completed by thestudied products during the calibration
Several samples per counterfeit seizures have been ana-lysed with the device and interpreted with the discoverymode Their chemical composition was previously deter-mined by the lab instruments a Raman a near-infraredand a nuclear magnetic resonance spectrometers
For 14 samples no match could be found with theinstrument database This can be explained by the factthat for these samples the chemical composition was amixture of at least three components part of the databasebut difficult to detect all with an automatic Raman data-base search
The second point is that for 10 counterfeits a differentcompositionwas proposed by the device between the differentsamples of the same seizure Once again it can be explainedby the complex mixture of the counterfeits Some compo-nents could be identified while others not For instance for
Table 4 Results of the four robustness tests The conditions of the test like the sampling have been added together with the number ofspectra measured The results consist in the p value range and the passfail answers
Robustness test Product ConditionsNumber ofspectra
pVal resultsIdentification against
the corresponding method
R1 degradationwith the laser
Product 20 No change in sample position 20 [05745ndash06669] Pass
Product 31 No change in sample position 20 [05616ndash07335] Pass
R2 influence ofthe sample position
Product 20 Sample position changed 10 [05523ndash06746] Pass
Product 31 Sample position changed 10 [03316ndash06481] Pass
R3 influence ofthe sampling
Product 20
Glass vial 3 [05386ndash05950] Pass
Transparent blister 3 [05891ndash06534] Pass
Direct 3 [05344ndash05746] Pass
Product 32
Glass vial 3 [01131ndash02471] Pass
Transparent blister 3 [00814ndash03385] Pass
Direct 3 [01628ndash04482] Pass
Product 27
White blister 3 [00175ndash00259] Fail
White blister 3 [00272ndash01164] Fail
White blister 3 [00017ndash00731] Fail
R4 influence ofthe operator
Product 20Operator A 3 [06815ndash06871] Pass
Operator B 3 [06143ndash06526] Pass
Product 31Operator A 3 [06159ndash06779] Pass
Operator B 3 [06599ndash07381] Pass
Table 3 Overview of the results obtained for the validation and the tests of the second validation (lowast) In these tests ldquofailrdquo is actually theexpected answer since these products should not be identified by the method
Step of the method development Sample set Results of the identification
Validation
Validation with positive samples(eg product 1 tested on the method of product 1)
100 pass
Validation against all other products(eg product 1 tested on the method of product 2)
100 faillowast
Second validation
Counterfeits 100 faillowast
Placebos 67 faillowast
Generics 35 faillowast
8 Journal of Spectroscopy
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
A selectivity test proposed by the device was evaluatedafter the validation This test consists in evaluating for eachmethod the risks of getting mismatches The option wastested for the products providing the lowest p values duringthe validation For product 26 for instance the selectivity testsuggested that the closest other products would deliver a pvalue of 00026 Tested physically during the validation theproduct actually delivered a p value of 00352 using the
method of product 26 While the selectivity test might bean option to see quickly if possible mismatches might occurthe results delivered are not real The validation performed inthis study is thus considered more completed than theselectivity test Raman can thus not be considered a universalmethod for counterfeit detection
For six other products not included in the 33 presentedproduct families no method could be created since the
Table 1 Validation results obtained with the genuine products The number of signatures measured for each product has been added Theminimum p values (pVal min) obtained for the validation samples of the corresponding method are presented in the table together with thecorresponding maximum p values (pVal max) of the closest other products The device provides a positive identification for p values superioror equal to 005
Sample type Product (API) Number of signaturesValidation
pVal min ofvalidation samples
pVal max of closestother products
Tablets
Product 1 (71) 9 01404 00000
Product 2 (36) 9 05973 00105
Product 2 (72) 9 03576 00013
Product 3 (80 95) 18 05455 00000
Product 4 (67 75) 12 04137 00000
Product 5 (25 33) 18 03439 00171
Product 6 (61) 9 04847 00000
Product 7 (57) 9 04490 00007
Product 7 (56) 9 04106 00042
Product 8 (17) 9 04031 00083
Product 9 (3 6 12 30) 36 01292 00000
Product 10 (8) 9 05329 00044
Product 10 (4) 9 04481 00010
Product 11 (3) 9 06458 00129
Product 12 (75) 9 05785 00000
Product 13 (04 08 1 2 3) 54 04089 00145
Product 14 (61) 9 06009 00000
Product 15 (12) 9 04619 00167
Product 16 (51) 9 04299 00000
Product 17 (25 45) 27 04416 00000
Product 18 (2) 9 06275 00015
Product 19 (03 06 1) 24 01763 00009
Product 20 (24 32) 27 03453 00000
Product 21 (75) 6 05781 00000
Product 22 (70) 9 01562 00000
Product 23 (1 3 6) 36 04444 00348
Product 24 (41) 9 05872 00000
Product 25 (77 78) 18 05822 00000
Product 26 (08) 24 01807 00352
Capsules
Product 27 (83) 9 06223 00000
Product 28 (49) 9 05439 00000
Product 29 (76) 6 05920 00000
Product 30 (49) 9 01669 00000
Product 31 (83) 18 01218 00000
Product 32 (50) 9 01165 00000
Powders Product 33 (81) 30 06139 00000
6 Journal of Spectroscopy
fluorescence prevented the measurement of the signaturesAmong these six products two were tablets two werecapsules and two were lyophilized products Since nosignatures could be taken with these samples the resultsare not displayed in this paper
322 Second Validation with Counterfeits Placebos andGenerics All the tested counterfeits consisting in 44 samplesfailed for identification The obtained p values were verylowmdashinferior to 00003mdashwhich comforted the robustnessof the methods for the detection of counterfeits The detailsof the results of the second validation are presented inTable 2
Nine of the tested generics have the same formulationas the genuine brand products and their spectra are identicalas previously presented in Figure 4 These samples wereconsequently identified by the device as genuine Since theobjective of the methods is to reject counterfeits and genericswere only used to simulate them these results are notconsidered critical The other five generics were rejected bythe methods The p values obtained are highly dependenton the tested generic
Two of the three tested placebos were rejected by themethods The third one the placebo of product 26 wasdetectedby the instrumentas genuinewithamaximumpvalueof 05257 which is very high The method for product 26 alow-dosage product was then considered not specific enoughand was then deleted from the database
The results of the validation and the tests performed forthe second validation are summarized in Table 3
33 Robustness Results The results of the four robustnesstests are presented in Table 4
According to the first test a capsule of product 31 and atablet of product 20 were measured 20 times at the sameposition to check if the samples were damaged by the laserThe capsule and the tablet were correctly identified even after20 measurements Additionally no important variation ofthe p value range was observed which confirms that the laserdid not damage the samples At this point it should be notedthat no soft-gel capsules should be measured with the Ramaninstrument since their shell is very sensitive to the heat ofthe laser
A capsule of product 31 and a tablet of product 20 wereeach measured 10 times for the robustness test R2 They weremoved between each measurement in order to check if theposition of the sample had an influence on the results Thesamples were correctly identified during each measurementwith little difference in the p values This means that theposition of the samples has low influence on their identifica-tion However it is recommended to measure the capsules onthe side where there is only one layer of gelatin Also careshould be taken in order not to break the samples with thesample holder For instance if a tablet is broken and its coatingremoveddue to thebreaking thena slightly different spectrumcan be obtained which might alter the results
The robustness test R3 deals with the sampling that isavailable for the studied medicines Three products weremeasured either directly with the sample holder throughtransparent glass vials or through blisters depending onthe available packaging Two types of blister were evaluatedtransparent (plastic) and white (plastic) The samples wereall correctly authenticated when measured directly throughtransparent glass vials and through transparent blisters Theproducts could not be correctly identified through whiteblister It is consequently recommended to use the tablet
Table 2 Second validation results obtained with the challenging samples The number of samples measured for each product has been addedThe maximum p values (pVal) obtained for each counterfeit are presented in the table together with the corresponding passfail result
Sample set Counterfeited product Number of samples pVal max Identification against the corresponding method
Counterfeits
Product 27 2 00000 Fail
Product 6 6 00000 Fail
Product 28 3 00000 Fail
Product 26 1 00000 Fail
Product 19 5 00001 Fail
Product 23 10 00000 Fail
Product 25 7 00000 Fail
Product 32 10 00003 Fail
Placebos
Product 4 1 00000 Fail
Product 32 1 00040 Fail
Product 26 1 05257 Pass
Generics
Product 20 1 04937 Pass
Product 25 batch 1 2 00224 Fail
Product 25 batch 2 3 06326 Pass
Product 32 batch 1 1 00042 Fail
Product 32 batch 2 4 01548 Pass
Product 5 2 00110 Fail
Product 33 1 01295 Pass
7Journal of Spectroscopy
holder to measure the samples However it is interesting tonote that according to this test a product measured on thefield through transparent blister or in transparent glass vialsmight be correctly authenticated
In order to evaluate the human influence (test R4) a tabletof product 20 and a capsule of product 31 have beenmeasured three times by two different operators The sampleswere all correctly identified and the obtained p values are veryclose between the operators It seems that the operators haveno influence on the identification which is an importantfactor for the use on the field It should be noted that bothoperators were properly trained before the robustness test
34 Composition of the Counterfeits As soon as a counterfeithas been confirmed it is necessary to determine its chemicalcomposition in order to estimate the impact on the patientsrsquosafety In this frame the discovery mode of the instrumenthas been evaluated with the studied counterfeits This modeproposes in case of a ldquofailrdquo result the chemical compositionof the tested sample This composition is suggested according
to the database furnished by the supplier and integrated inthe device The database contains more than 8000 referencesof excipients and active substances and was completed by thestudied products during the calibration
Several samples per counterfeit seizures have been ana-lysed with the device and interpreted with the discoverymode Their chemical composition was previously deter-mined by the lab instruments a Raman a near-infraredand a nuclear magnetic resonance spectrometers
For 14 samples no match could be found with theinstrument database This can be explained by the factthat for these samples the chemical composition was amixture of at least three components part of the databasebut difficult to detect all with an automatic Raman data-base search
The second point is that for 10 counterfeits a differentcompositionwas proposed by the device between the differentsamples of the same seizure Once again it can be explainedby the complex mixture of the counterfeits Some compo-nents could be identified while others not For instance for
Table 4 Results of the four robustness tests The conditions of the test like the sampling have been added together with the number ofspectra measured The results consist in the p value range and the passfail answers
Robustness test Product ConditionsNumber ofspectra
pVal resultsIdentification against
the corresponding method
R1 degradationwith the laser
Product 20 No change in sample position 20 [05745ndash06669] Pass
Product 31 No change in sample position 20 [05616ndash07335] Pass
R2 influence ofthe sample position
Product 20 Sample position changed 10 [05523ndash06746] Pass
Product 31 Sample position changed 10 [03316ndash06481] Pass
R3 influence ofthe sampling
Product 20
Glass vial 3 [05386ndash05950] Pass
Transparent blister 3 [05891ndash06534] Pass
Direct 3 [05344ndash05746] Pass
Product 32
Glass vial 3 [01131ndash02471] Pass
Transparent blister 3 [00814ndash03385] Pass
Direct 3 [01628ndash04482] Pass
Product 27
White blister 3 [00175ndash00259] Fail
White blister 3 [00272ndash01164] Fail
White blister 3 [00017ndash00731] Fail
R4 influence ofthe operator
Product 20Operator A 3 [06815ndash06871] Pass
Operator B 3 [06143ndash06526] Pass
Product 31Operator A 3 [06159ndash06779] Pass
Operator B 3 [06599ndash07381] Pass
Table 3 Overview of the results obtained for the validation and the tests of the second validation (lowast) In these tests ldquofailrdquo is actually theexpected answer since these products should not be identified by the method
Step of the method development Sample set Results of the identification
Validation
Validation with positive samples(eg product 1 tested on the method of product 1)
100 pass
Validation against all other products(eg product 1 tested on the method of product 2)
100 faillowast
Second validation
Counterfeits 100 faillowast
Placebos 67 faillowast
Generics 35 faillowast
8 Journal of Spectroscopy
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
fluorescence prevented the measurement of the signaturesAmong these six products two were tablets two werecapsules and two were lyophilized products Since nosignatures could be taken with these samples the resultsare not displayed in this paper
322 Second Validation with Counterfeits Placebos andGenerics All the tested counterfeits consisting in 44 samplesfailed for identification The obtained p values were verylowmdashinferior to 00003mdashwhich comforted the robustnessof the methods for the detection of counterfeits The detailsof the results of the second validation are presented inTable 2
Nine of the tested generics have the same formulationas the genuine brand products and their spectra are identicalas previously presented in Figure 4 These samples wereconsequently identified by the device as genuine Since theobjective of the methods is to reject counterfeits and genericswere only used to simulate them these results are notconsidered critical The other five generics were rejected bythe methods The p values obtained are highly dependenton the tested generic
Two of the three tested placebos were rejected by themethods The third one the placebo of product 26 wasdetectedby the instrumentas genuinewithamaximumpvalueof 05257 which is very high The method for product 26 alow-dosage product was then considered not specific enoughand was then deleted from the database
The results of the validation and the tests performed forthe second validation are summarized in Table 3
33 Robustness Results The results of the four robustnesstests are presented in Table 4
According to the first test a capsule of product 31 and atablet of product 20 were measured 20 times at the sameposition to check if the samples were damaged by the laserThe capsule and the tablet were correctly identified even after20 measurements Additionally no important variation ofthe p value range was observed which confirms that the laserdid not damage the samples At this point it should be notedthat no soft-gel capsules should be measured with the Ramaninstrument since their shell is very sensitive to the heat ofthe laser
A capsule of product 31 and a tablet of product 20 wereeach measured 10 times for the robustness test R2 They weremoved between each measurement in order to check if theposition of the sample had an influence on the results Thesamples were correctly identified during each measurementwith little difference in the p values This means that theposition of the samples has low influence on their identifica-tion However it is recommended to measure the capsules onthe side where there is only one layer of gelatin Also careshould be taken in order not to break the samples with thesample holder For instance if a tablet is broken and its coatingremoveddue to thebreaking thena slightly different spectrumcan be obtained which might alter the results
The robustness test R3 deals with the sampling that isavailable for the studied medicines Three products weremeasured either directly with the sample holder throughtransparent glass vials or through blisters depending onthe available packaging Two types of blister were evaluatedtransparent (plastic) and white (plastic) The samples wereall correctly authenticated when measured directly throughtransparent glass vials and through transparent blisters Theproducts could not be correctly identified through whiteblister It is consequently recommended to use the tablet
Table 2 Second validation results obtained with the challenging samples The number of samples measured for each product has been addedThe maximum p values (pVal) obtained for each counterfeit are presented in the table together with the corresponding passfail result
Sample set Counterfeited product Number of samples pVal max Identification against the corresponding method
Counterfeits
Product 27 2 00000 Fail
Product 6 6 00000 Fail
Product 28 3 00000 Fail
Product 26 1 00000 Fail
Product 19 5 00001 Fail
Product 23 10 00000 Fail
Product 25 7 00000 Fail
Product 32 10 00003 Fail
Placebos
Product 4 1 00000 Fail
Product 32 1 00040 Fail
Product 26 1 05257 Pass
Generics
Product 20 1 04937 Pass
Product 25 batch 1 2 00224 Fail
Product 25 batch 2 3 06326 Pass
Product 32 batch 1 1 00042 Fail
Product 32 batch 2 4 01548 Pass
Product 5 2 00110 Fail
Product 33 1 01295 Pass
7Journal of Spectroscopy
holder to measure the samples However it is interesting tonote that according to this test a product measured on thefield through transparent blister or in transparent glass vialsmight be correctly authenticated
In order to evaluate the human influence (test R4) a tabletof product 20 and a capsule of product 31 have beenmeasured three times by two different operators The sampleswere all correctly identified and the obtained p values are veryclose between the operators It seems that the operators haveno influence on the identification which is an importantfactor for the use on the field It should be noted that bothoperators were properly trained before the robustness test
34 Composition of the Counterfeits As soon as a counterfeithas been confirmed it is necessary to determine its chemicalcomposition in order to estimate the impact on the patientsrsquosafety In this frame the discovery mode of the instrumenthas been evaluated with the studied counterfeits This modeproposes in case of a ldquofailrdquo result the chemical compositionof the tested sample This composition is suggested according
to the database furnished by the supplier and integrated inthe device The database contains more than 8000 referencesof excipients and active substances and was completed by thestudied products during the calibration
Several samples per counterfeit seizures have been ana-lysed with the device and interpreted with the discoverymode Their chemical composition was previously deter-mined by the lab instruments a Raman a near-infraredand a nuclear magnetic resonance spectrometers
For 14 samples no match could be found with theinstrument database This can be explained by the factthat for these samples the chemical composition was amixture of at least three components part of the databasebut difficult to detect all with an automatic Raman data-base search
The second point is that for 10 counterfeits a differentcompositionwas proposed by the device between the differentsamples of the same seizure Once again it can be explainedby the complex mixture of the counterfeits Some compo-nents could be identified while others not For instance for
Table 4 Results of the four robustness tests The conditions of the test like the sampling have been added together with the number ofspectra measured The results consist in the p value range and the passfail answers
Robustness test Product ConditionsNumber ofspectra
pVal resultsIdentification against
the corresponding method
R1 degradationwith the laser
Product 20 No change in sample position 20 [05745ndash06669] Pass
Product 31 No change in sample position 20 [05616ndash07335] Pass
R2 influence ofthe sample position
Product 20 Sample position changed 10 [05523ndash06746] Pass
Product 31 Sample position changed 10 [03316ndash06481] Pass
R3 influence ofthe sampling
Product 20
Glass vial 3 [05386ndash05950] Pass
Transparent blister 3 [05891ndash06534] Pass
Direct 3 [05344ndash05746] Pass
Product 32
Glass vial 3 [01131ndash02471] Pass
Transparent blister 3 [00814ndash03385] Pass
Direct 3 [01628ndash04482] Pass
Product 27
White blister 3 [00175ndash00259] Fail
White blister 3 [00272ndash01164] Fail
White blister 3 [00017ndash00731] Fail
R4 influence ofthe operator
Product 20Operator A 3 [06815ndash06871] Pass
Operator B 3 [06143ndash06526] Pass
Product 31Operator A 3 [06159ndash06779] Pass
Operator B 3 [06599ndash07381] Pass
Table 3 Overview of the results obtained for the validation and the tests of the second validation (lowast) In these tests ldquofailrdquo is actually theexpected answer since these products should not be identified by the method
Step of the method development Sample set Results of the identification
Validation
Validation with positive samples(eg product 1 tested on the method of product 1)
100 pass
Validation against all other products(eg product 1 tested on the method of product 2)
100 faillowast
Second validation
Counterfeits 100 faillowast
Placebos 67 faillowast
Generics 35 faillowast
8 Journal of Spectroscopy
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
holder to measure the samples However it is interesting tonote that according to this test a product measured on thefield through transparent blister or in transparent glass vialsmight be correctly authenticated
In order to evaluate the human influence (test R4) a tabletof product 20 and a capsule of product 31 have beenmeasured three times by two different operators The sampleswere all correctly identified and the obtained p values are veryclose between the operators It seems that the operators haveno influence on the identification which is an importantfactor for the use on the field It should be noted that bothoperators were properly trained before the robustness test
34 Composition of the Counterfeits As soon as a counterfeithas been confirmed it is necessary to determine its chemicalcomposition in order to estimate the impact on the patientsrsquosafety In this frame the discovery mode of the instrumenthas been evaluated with the studied counterfeits This modeproposes in case of a ldquofailrdquo result the chemical compositionof the tested sample This composition is suggested according
to the database furnished by the supplier and integrated inthe device The database contains more than 8000 referencesof excipients and active substances and was completed by thestudied products during the calibration
Several samples per counterfeit seizures have been ana-lysed with the device and interpreted with the discoverymode Their chemical composition was previously deter-mined by the lab instruments a Raman a near-infraredand a nuclear magnetic resonance spectrometers
For 14 samples no match could be found with theinstrument database This can be explained by the factthat for these samples the chemical composition was amixture of at least three components part of the databasebut difficult to detect all with an automatic Raman data-base search
The second point is that for 10 counterfeits a differentcompositionwas proposed by the device between the differentsamples of the same seizure Once again it can be explainedby the complex mixture of the counterfeits Some compo-nents could be identified while others not For instance for
Table 4 Results of the four robustness tests The conditions of the test like the sampling have been added together with the number ofspectra measured The results consist in the p value range and the passfail answers
Robustness test Product ConditionsNumber ofspectra
pVal resultsIdentification against
the corresponding method
R1 degradationwith the laser
Product 20 No change in sample position 20 [05745ndash06669] Pass
Product 31 No change in sample position 20 [05616ndash07335] Pass
R2 influence ofthe sample position
Product 20 Sample position changed 10 [05523ndash06746] Pass
Product 31 Sample position changed 10 [03316ndash06481] Pass
R3 influence ofthe sampling
Product 20
Glass vial 3 [05386ndash05950] Pass
Transparent blister 3 [05891ndash06534] Pass
Direct 3 [05344ndash05746] Pass
Product 32
Glass vial 3 [01131ndash02471] Pass
Transparent blister 3 [00814ndash03385] Pass
Direct 3 [01628ndash04482] Pass
Product 27
White blister 3 [00175ndash00259] Fail
White blister 3 [00272ndash01164] Fail
White blister 3 [00017ndash00731] Fail
R4 influence ofthe operator
Product 20Operator A 3 [06815ndash06871] Pass
Operator B 3 [06143ndash06526] Pass
Product 31Operator A 3 [06159ndash06779] Pass
Operator B 3 [06599ndash07381] Pass
Table 3 Overview of the results obtained for the validation and the tests of the second validation (lowast) In these tests ldquofailrdquo is actually theexpected answer since these products should not be identified by the method
Step of the method development Sample set Results of the identification
Validation
Validation with positive samples(eg product 1 tested on the method of product 1)
100 pass
Validation against all other products(eg product 1 tested on the method of product 2)
100 faillowast
Second validation
Counterfeits 100 faillowast
Placebos 67 faillowast
Generics 35 faillowast
8 Journal of Spectroscopy
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
the counterfeits of product 19 made ofmelatonin starch andsaccharose melatonin is missing in the search of the deviceWhile it is observable in the Raman spectra the bands aretoo small to be detected by an automatic search
In three cases (product 29 seizures number 2 and number3 and product 33) the chemical composition proposed by thediscovery mode was correct The counterfeits are only madeof one component an API easy to identify by Raman
It can be concluded that some of the compounds of thecounterfeits are common compared to the ldquorightrdquo composi-tion determined by the lab instruments While the discoverymode can help the analyst determine the chemical composi-tion of the confirmed counterfeits it cannot be reported asthe right composition It should be noted that in the discoverymode the operator also has access to the overlay of the testedsample and the encountered matches of the database Thespectra can then be visually compared to evaluate if theproposedmatches are correct or not In this case the operatorshould however be a Raman specialist
35 Comparison with a Lab Instrument In order to evaluatethe quality of the data generated by the handheld Ramanspectrometer these were compared with the Raman spectraobtained with a benchtop instrument The chosen instrumentis a Raman spectrometer equipped with a probe generating a6mm diameter laser spot The device possesses a laser of785nm providing a power of 270mW on the samples anda spectra range going from 150 to 1890 cmminus1
Figures 6(a) and 6(b) present examples of overlaidspectra generated by the handheld and the benchtop Ramanspectrometers Two types of spectra are represented for thehandheld device the ones generated while taking a signatureand the others generated during simple measurements itmeans routine analysis or method validation
The spectra generated by both devices for the signaturesare both of good quality and are almost identical The onesgenerated by the handheld Raman spectrometer duringsimple measurements are much noisier However the mainbands are easily observed and the spectra used for validationall provided 100 of correct identification
It can also be noted that the range used by the handheldRaman spectrometer longer than the one of the benchtopdevice does not bring much more information and is usuallythe noisiest part of the spectrum However based on thevalidation results this additional range (1890ndash2900 cmminus1)does not seem to have a negative impact on the identification
The results of the calibration obtained by both devicescan be partly compared for the tablets while referring to aprevious study performed with this lab instrument [48] Inboth cases 100 of the samples used for the calibrationand for the validation have been correctly identified Con-sequently the results obtained by the handheld Ramanspectrometer are in the present case as good as the onesobtained with the lab instrument
4 Discussion
In this paper methods have been presented for the authenti-cation of almost all the solid products produced by the firm
The database consists indeed in 33 product families andamong them are 62 product formulations The calibrationand validation could be successfully performed for all of theseproducts Moreover the methods were validated against allthe other products of the database which were successfullyrejected by the studied methods Several challenges raisedduring the spectra analysis have been encountered duringthe development of the presented methods The heterogene-ity of the product families and the closeness of formulationbetween different families could be solved by the highnumber of signatures taken and their correct gathering inthe methods The importance of taking several spectraespecially of samples from different manufacturing sitesshould be underlined All the tested counterfeits wererejected by the methods which confirms the dedicated useof the handheld Raman spectrometer for counterfeit detec-tion Some generics were recognized as genuine brandproducts however only when their formulation was reallyclose These results were not considered critical since theobjective of the methods was to detect counterfeits and notgeneric products The tested placebos were rejected by themethods apart from one sample the placebo of product 26Consequently product 26 was deleted from the database afterthe second validation phase This test suggests that for low-dosage products an additional analytical technique shouldbe used that would confirm the presence of the API In thesecases a fingerprint of the whole tablet is nevertheless availableby Raman meaning that at least the right excipients arepresent in the tested product [48]
Six products not included in the 33 presented productscould not be measured at all by the instrument since thefluorescence prevented the acquisition of the signaturesFluorescence constitutes the main drawback of the Ramantechnology which can be on other instruments diminishedfor instance by the use of a fluorescence correction optionor the use of a different wavelength like 1084nm In thepresent case no solution can be found for the products pre-senting fluorescence However a large majority of the testedproducts could be successfully measured with the handheldRaman spectrometer Furthermore the results obtainedwith the handheld Raman spectrometer were comparedto the ones generated by a lab instrument It resulted thatthe handheld instrument delivered good results as the labRaman spectrometer
The robustness tests performed during the validationdemonstrated that the laser did not damage the samplesMoreover no influence could be observed of the sampleposition or the operator However it was pointed out thatthe operators had to be trained so that they would positionthe samples carefully Also the samples should not be dam-aged when placed on the device for instance by removinginvoluntarily the coating or measured through a thick layerof capsule shell A trained operator could consequentlyperform the measurement on the field and be able toauthenticate the samples without needing further knowl-edge in Raman spectroscopy The fourth robustness testshowed that a measurement through a white blister did notallow the identification of the capsule Since the measuredproduct (product 27) contains 83 of API this confirms that
9Journal of Spectroscopy
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
this can be attributed to the type of packaging Furthermoretransparent packaging like glass vial or blister delivered thesame results as in direct contact with the instrument in thesample holder This fact could be of advantage during ameasurement on the field since a nondestructive measure-ment should always be privileged While Raman spectros-copy is not destructive in itself it can be considereddestructive if tablets or capsules have to be removed fromtheir packaging for the analysis
Handheld Raman spectrometers are also interesting forcounterfeit detection for the reason that a method can bedeveloped on a master instrument and then easily transferredto several other ones for a local measurement on the field forinstance at airports at the customs or in warehouses In this
frame the transfer of the presented methods to anotherinstrument is quite easy The transfer basically only consistedin the copying of files from the master device on the newdevice Then each method can be tested with an independentset of samples in order to check that the spectrometer has noinfluence on the measurement At this point the intro-duction of all possible manufacturing sites in the databaseis particularly important if the new instrument is going tobe used for the testing in another region of the world Thatway much fewer false negatives should be expected Theuse of the handheld devices on the field would help speedup the investigation by reducing the time spent by sendingthe samples It is however recommended to send the samplesthat failed the identification with the handheld Raman
Product 32 measured with the TruScan as signatureProduct 32 measured with the KaiserProduct 32 measured with the TruScan as routine measure
10
09
10
11
12
13
14
15
16
17
18
19
20
21
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(a)
Product 20 measured with the TruScan as signatureProduct 20 measured with the KaiserProduct 20 measured with the TruScan as routine measure
100607080910111213141516171819
Extin
ctio
n
500 1000 1500 2000 2500Raman shi (cm‒1)
(b)
Figure 6 Overlay of Raman spectra obtained with the TruScan Thermoreg (signature and routine measure) and the RXN1 Kaiser OpticalSystemsreg devices for product 32 capsule (a) and product 20 tablet (b)
10 Journal of Spectroscopy
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
spectrometer to labs in order to confirm the counterfeits andperform the analysis of the whole chemical compositionIndeed the test conducted in this study concerning thediscovery mode of the instrument revealed that in most casesand based on the previous analyses performed with diversetechnologies the proposed chemical composition was notcorrect This mode is able to determine the compositionwhen the counterfeits are only made of one component Forthe analysis of mixtures a manual spectrum interpretationand further analyses are required
The main advantage of the studied instrument residesin how easy the method calibration is Indeed no furtherknowledge in Raman spectroscopy or chemometrics isrequired although knowledge in method validation shouldbe privileged The proposed algorithm automatically calcu-lates the p values while taking signatures of the productsNevertheless this system might be considered a ldquoblack boxrdquowith no possible way of modifying the calibration of themethod The opportunity of choosing further chemometrictools would be welcome which seems to be possible in thenewest versions of the device Indeed the more products areintroduced in the database the more flexibility is needed todevelop a robust calibration Also it would be of advantageto be able to change the sample exposure during the routinemeasurements In fact the quality of the spectra is worseduring this phase than during the signature mode whichmight generate less reliable results The end of the range(1890ndash2900 cmminus1) is additionally not of great interest for thestudied products and represents the noisiest part of thespectrum A compromise might be found between the timeof measurement partly dependent on the sample exposureand the quality of the spectra Finally another possibleimprovement of the device would be the possibility ofimporting spectra from another device Even if the calibrationstep is as fast as possible concerning the computation takingsignatures of a large database of products and validating all ofthem require many hours of measurements The spectrashould in this case be transformed with calibration transfertechniques and the methods carefully validated since theyare spectrometer dependent but a lot of time would be sparedin the measurement phase
5 Conclusion
Handheld spectrometers are more and more used for the on-site analysis of pharmaceuticals In this paper a handheldRaman spectrometer has been evaluated for the analysis ofcounterfeits of 33 product families of medicines under theform of tablets capsules and powders Several challengeshad to be dealt with during the calibration of the methodslike the heterogeneity of several product families or thecloseness of formulation between different products Never-theless the methods could be successfully created and fullyvalidated The tested counterfeits were rejected by themethods thus confirming the specificity of the calibrationfor counterfeit detection The methods were also proved tobe robust in terms of degradation by the laser samplepositioning and influence of the operators The samples
could be partly measured through the packaging dependingon their opacity
Some limits could nevertheless be found concerningthe instrumentation like the positive identification of somegenerics and one placebo The device is not adapted to themeasurement of low-dosage medicines for which a comple-mentary method like infrared spectroscopy should be usedMoreover the analysis of the chemical composition sug-gested by the discovery mode of the device does not providecorrect results as soon as more than one component isincluded in the counterfeit While the calibration is per-formed quite quickly the chemometrics cannot be chosenand the method developer has to rely on the proposedalgorithm Finally the spectra obtained during the routinemeasurements are quite noisy and might generate falsenegative results Apart from these encountered limits thehandheld Raman spectrometer performed very well for theidentification of the 33 product families of medicines andis uncontestably a reliable tool for the analysis of counterfeitmedicines on the field More and more handheld devices arebeing used in this context which should be encouraged inorder to speed up the investigations and provide betterchances to pursue the criminals While the handheld Ramanspectrometer might be used as a first screening tool on thefield the counterfeits detected can be sent to a lab forconfirmation and determination of the chemical composi-tion for the evaluation of the patientrsquos health risk
Conflicts of Interest
The authors declare that there is no conflict of interestregarding the publication of this paper
References
[1] CdelCastilloRodriguez andM J LozanoEstevan ldquoCounterfeitmedicine A threat to health Legal situationrdquo Anales de laReal Academia Nacional de Farmacia vol 81 no 4pp 329ndash333 2015
[2] K Deacutegardin Y Roggo and P Margot ldquoUnderstanding andfighting the medicine counterfeit marketrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 87 pp 167ndash175 2014
[3] J Harris P Stevens and J Morris Keeping It Real Combatingthe Spread of Fake Drugs in Poor Countries InternationalPolicy Network 2009 April 2014 httpwwwafricanlibertyorgpdfKeepingitrealpdf
[4] E Medina E Bel and J M Sune ldquoCounterfeit medicines inPeru a retrospective review (1997-2014)rdquo BMJ Open vol 6no 4 pp 1ndash11 2016
[5] S Garrigues and M de la Guardia ldquoNon-invasive analysis ofsolid samplesrdquo Trends in Analytical Chemistry vol 43pp 161ndash173 2013
[6] F Adar E Lee A Whitley and M Witkowski Single-PointAnalysis and Raman Mapping of Tablet Dosage Formulationas a Means for Detecting and Sourcing Counterfeit Pharma-ceuticals Pharmaceutical Online 2007 March 2011 httpwwwpharmaceuticalonlinecomdownloadmvcSingle-Point-Analysis-And-Raman-Mapping-Of-Do-0001user=20
[7] M Boiret D Rutledge N Gorretta Y M Ginot and J MRoger ldquoUtilisation de la Microscopie Raman et des Meacutethodes
11Journal of Spectroscopy
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
Chimiomeacutetriques pour la Deacutetection de Comprimeacutes Contre-faitsrdquo Spectra Analyse vol 298 pp 74ndash80 2014
[8] K Deacutegardin Y Roggo F Been and P Margot ldquoDetection andchemical profiling of medicine counterfeits by Ramanspectroscopy and chemometricsrdquo Analytica Chimica Actavol 705 no 1-2 pp 334ndash341 2011
[9] K Kwok and L S Taylor ldquoAnalysis of the packaging enclosinga counterfeit pharmaceutical tablet using Raman microscopyand two-dimensional correlation spectroscopyrdquo VibrationalSpectroscopy vol 61 pp 176ndash182 2012
[10] J K Mbinze P-Y Sacreacute A Yemoa et al ldquoDevelopmentvalidation and comparison of NIR and Raman methods forthe identification and assay of poor-quality oral quininedropsrdquo Journal of Pharmaceutical and Biomedical Analysisvol 111 pp 21ndash27 2015
[11] S Neuberger and C Neusuumlss ldquoDetermination of counterfeitmedicines by Raman spectroscopy systematic study based ona large set of model tabletsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 112 pp 70ndash78 2015
[12] J Peters A Luczak V Ganesh E Park and R KalyanaramanldquoRaman spectral fingerprinting for biologics counterfeitdrug detectionrdquo American Pharmaceutical Review vol 19no 2 2016
[13] P De Peinder M J Vredenbregt T Visser and D De KasteldquoDetection of Lipitorreg counterfeits a comparison of NIR andRaman spectroscopy in combination with chemometricsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 47no 4-5 pp 688ndash694 2008
[14] F E Dowell E B Maghirang F M Fernandez P N NewtonandM D Green ldquoDetecting counterfeit antimalarial tablets bynear-infrared spectroscopyrdquo Journal of Pharmaceutical andBiomedical Analysis vol 48 no 3 pp 1011ndash1014 2008
[15] J Dubois J C Wolff J K Warrack J Schoppelrei and E NLewis ldquoNIR chemical imaging for counterfeit pharmaceuticalproducts analysisrdquo Spectroscopy vol 22 pp 36ndash41 2007httpwwwiesmatcomiesmatuploadfileMalvernProductos-MALNIR-NIR20Chemical20Imaging20for20Counterfeit20Pharmaceutical20Products20Analysispdf
[16] R da Silva Fernandes F S L da Costa P Valderrama P HMarccedilo and K M G de Lima ldquoNon-destructive detection ofadulterated tablets of glibenclamide using NIR and solid-phase fluorescence spectroscopy and chemometric methodsrdquoJournal of Pharmaceutical and Biomedical Analysis vol 66pp 85ndash90 2012
[17] M B Lopes and J C Wolff ldquoInvestigation into classificationsourcing of suspect counterfeit Heptodintrade tablets by nearinfrared chemical imagingrdquo Analytica Chimica Acta vol 633no 1 pp 149ndash155 2009
[18] S H F Scafi and C Pasquini ldquoIdentification of counterfeitdrugs using near infrared spectroscopyrdquo The Analyst vol 126no 12 pp 2218ndash2224 2001
[19] KDeacutegardin AGuillemainNViegasGuerreiro andY RoggoldquoNear infrared spectroscopy for counterfeit detection using alarge database of pharmaceutical tabletsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 128 pp 89ndash97 2016
[20] J R Lucio-Gutierrez J Coello and S Maspoch ldquoExpeditiousidentification and semi-quantification of Panax ginseng usingnear infrared spectral fingerprints and multivariate analysisrdquoAnalytical Methods vol 5 no 4 pp 857ndash865 2013
[21] E Deconinck P Y Sacreacute D Coomans and J De BeerldquoClassification trees based on infrared spectroscopic data to
discriminate between genuine and counterfeit medicinesrdquoJournal of Pharmaceutical and Biomedical Analysis vol 57pp 68ndash75 2011
[22] C Ricci C Eliasson N Macleod P Newton P Matousek andS Kazarian ldquoCharacterization of genuine and fake artesunateanti-malarial tablets using Fourier transform infrared imagingand spatially offset Raman spectroscopy through blister packsrdquoAnalytical and Bioanalytical Chemistry vol 389 no 5pp 1525ndash1532 2007
[23] K Deacutegardin and Y Roggo ldquoCounterfeit analysis strategyillustrated by a case studyrdquo Drug Testing and Analysis vol 8no 3-4 pp 388ndash397 2015
[24] H Ayvaz and L E Rodriguez-Saona ldquoApplication of hand-held and portable spectrometers for screening acrylamidecontent in commercial potato chipsrdquo Food Chemistry vol 174pp 154ndash162 2014
[25] A Keil N Talaty C Janflet et al ldquoAmbient mass spectrome-try with a handheld mass spectrometer at high pressurerdquoAnalytical Chemistry vol 79 no 20 pp 7734ndash7739 2007
[26] P Leary G S Dobson and J A Reffner ldquoDevelopmentand applications of portable gas chromatography-massspectrometry for emergency responders the military andlaw-enforcement organizationsrdquo Applied Spectroscopy vol 70no 5 pp 888ndash896 2016
[27] J D Dunn C M Gryniewicz-Ruzicka J F Kauffman B JWestenberger and L F Buhse ldquoUsing a portable ion mobilityspectrometer to screen dietary supplements for sibutraminerdquoJournal of Pharmaceutical and Biomedical Analysis vol 54no 3 pp 469ndash474 2010
[28] C R Appoloni and F L Melquiades ldquoPortable XRF andprincipal component analysis for bill characterisation inforensic sciencerdquo Applied Radiation and Isotopes vol 85pp 92ndash95 2014
[29] D Lauwers A Candeias A Coccato J Mirao L Moens andP Vandenabeele ldquoEvaluation of portable Raman spectroscopyand handheld X-ray fluorescence analysis (hXRF) for thedirect analysis of glypticsrdquo Spectrochimica Acta Part A Molec-ular and Biomolecular Spectroscopy vol 157 pp 146ndash1522015
[30] M D Hargreaves K Page T Munshi R Tomsett G Lynchand H G M Edwards ldquoAnalysis of seized drugs using porta-ble Raman spectroscopy in an airport environment - a proof ofprinciple studyrdquo Journal of Raman Spectroscopy vol 39 no 7pp 873ndash880 2008
[31] E L Izake ldquoForensic and homeland security applications ofmodern portable Raman spectroscopyrdquo Forensic ScienceInternational vol 202 no 1ndash3 pp 1ndash8 2010
[32] J Zheng S Pang T P Labuza and L He ldquoEvaluation ofsurface-enhanced Raman scattering detection using a hand-held and a bench-top Raman spectrometer a comparativestudyrdquo Talanta vol 129 pp 79ndash85 2014
[33] J Jehlicka P Viacutetek H G M Edwards M Heagraves andT Capoun ldquoApplication of portable Raman instruments forfast and non-destructive detection of minerals on outcropsrdquoSpectrochimica Acta Part A Molecular and BiomolecularSpectroscopy vol 73 no 3 pp 410ndash419 2009
[34] S Assi ldquoInvestigating the quality of medicines using handheldRaman spectroscopyrdquo European Pharmaceutical Reviewvol 19 no 5 pp 56ndash60 2014
[35] A J ONeil R D Jee G Lee A Charvill and A C MoffatldquoUse of a portable near infrared spectrometer for the
12 Journal of Spectroscopy
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
authentication of tablets and the detection of counterfeit ver-sionsrdquo Journal of Near Infrared Spectroscopy vol 16 no 3pp 327ndash333 2008
[36] J E Polli S W Hoag and S Flank ldquoComparison of authenticand suspect pharmaceuticalsrdquo Pharmaceutical Technologyvol 33 no 8 pp 46ndash52 2009
[37] M Alcalagrave M Blanco D Moyano et al ldquoQualitative andquantitative pharmaceutical analysis with a novel hand-heldminiature near infrared spectrometerrdquo Journal of Near InfraredSpectroscopy vol 21 no 6 pp 445ndash457 2013
[38] K Deacutegardin Y Roggo and P Margot ldquoEvaluation de Spec-tromegravetres Portables Raman Infrarouge et Proche Infrarougepour la Deacutetection de Contrefaccedilons de Meacutedicamentsrdquo SpectraAnalyse vol 276 pp 46ndash52 2010
[39] S Assi R A Watt and A C Moffat ldquoIdentification ofcounterfeit medicines from the internet and the world marketusing near-infrared spectroscopyrdquo Analytical Methods vol 3no 10 pp 2231ndash2236 2011
[40] A Luczak and R Kalyanaraman ldquoPortable and benchtopRaman technologies for product authentication and counterfeitdetectionrdquo American Pharmaceutical Review pp 1ndash9 2014
[41] R Bate and K Hess ldquoAnti-malarial drug quality in Lagos andAccra - a comparison of various qualityrdquo Malarial Journalvol 9 no 1 p 157 2010
[42] R Bate R Tren K Hess L Mooney and K Porter ldquoPilotstudy comparing technologies to test for substandard drugsin field settingsrdquo African Journal of Pharmacy and Pharmacol-ogy vol 3 no 4 pp 165ndash170 2009
[43] D E Bugay and R C Brush ldquoChemical identity testingby remote-based dispersive raman spectroscopyrdquo AppliedSpectroscopy vol 64 no 5 pp 467ndash475 2010
[44] M Hajjou Y Qin S Bradby D Bempong and P LukulayldquoAssessment of the performance of a handheld Raman devicefor potential use as a screening tool in evaluating medicinesqualityrdquo Journal of Pharmaceutical and Biomedical Analysisvol 74 pp 47ndash55 2012
[45] R Kalyanaraman M Ribick and G Dobler ldquoPortableRaman spectroscopy for pharmaceutical counterfeit detectionrdquoEuropean Pharmaceutical Review vol 17 no 5 pp 35ndash392012
[46] C Ricci L Nyadong F Yang et al ldquoAssessment of hand-heldRaman instrumentation for in situ screening for potentiallycounterfeit artesunate antimalarial tablets by FT-Raman spec-troscopy and direct ionization mass spectrometryrdquo AnalyticaChimica Acta vol 623 no 2 pp 178ndash186 2008
[47] J Spink D C Moyer and M R Rip ldquoAddressing the risk ofproduct fraud a case study of the Nigerian combating counter-feiting and sub-standard medicines initiativesrdquo Journal ofForensic Science and Criminology vol 4 no 2 pp 1ndash13 2016
[48] Y Roggo K Deacutegardin and P Margot ldquoIdentification ofpharmaceutical tablets by Raman spectroscopy and chemo-metricsrdquo Talanta vol 81 no 3 pp 988ndash995 2010
