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BETTER SOLUTIONSFOR FOOD AND
BEVERAGE ANALYSISTHIRD EDITION
Including ASE® Accelerated Solvent Extraction
LPN 0666-05
CarboPac and MicroBead are trademarks, and ASE, AutoSuppression, IonPac,and OmniPac are registered trademarks of Dionex Corporation.
Hydromatrix is a trademark of Varian Corporation.
All rights reserved. No part of this publication may be reproduced, stored in a retrievalsystem, or transmitted, in any form or by any means, electronic, mechanical, photocopying,recording, or otherwise, without the prior written permission of the publisher.
This book is published by:
Dionex CorporationP.O. Box 3603Sunnyvale, CA 94088-3603
Copyright © 2003 by Dionex Corporation.Printed in the United States of America.
LPN 0666-05 PDF 8/03
Second Edition printed January 1997.Revised and reprinted May 2000.Third Edition, August 2003.
INTRODUCTION ........................................................................................................... 1
OFFICIALLY APPROVED IC METHODS ....................................................................... 2
CHAPTER ONE: INORGANIC ANIONS AND CATIONS .............................................. 5Municipal Drinking Water ....................................................................................................................... 6Bromate in Drinking Water and Baked Goods ...................................................................................... 7Cations in Mineral Water and Drinking Water ...................................................................................... 7Cations in Soft Drinks and Wine ............................................................................................................. 7Transition Metals in Food ......................................................................................................................... 8Sulfite in Dried Apricot ............................................................................................................................ 8Iodide in Whole Milk ................................................................................................................................ 9Nitrate/Nitrite in Ham ............................................................................................................................. 9Polyphosphates ....................................................................................................................................... 10
CHAPTER TWO: ORGANIC ACIDS ............................................................................11Organic Acids in Fruit Juice ................................................................................................................... 12
Organic Acids in Cranberry Juice and Tomato Juice ..................................................................... 12Simultaneous High-Resolution Profiling of Organic Acids and Inorganic Ions in Fruit Juice .... 13
Anions and Organic Acids in an Irish Stout ......................................................................................... 14Food Dyes ................................................................................................................................................ 14
CHAPTER THREE: AMINES AND OTHER ORGANIC BASES .................................... 15Cations and Methylamines .................................................................................................................... 16Inorganic Cations, Choline, and Acetylcholine ................................................................................... 16Flavor Constituents and Additives ....................................................................................................... 17Amines as Indicators of Seafood Spoilage ........................................................................................... 17Triazine Herbicides in Raw Fruits and Vegetables .............................................................................. 18Water-Soluble Vitamins .......................................................................................................................... 18
CHAPTER FOUR: CARBOHYDRATES ........................................................................ 19Oligo- and Polysaccharides Derived from Hydrolyzed Glucose Syrup ........................................... 21Sugar Alcohols ......................................................................................................................................... 21
Sugar Alcohols in Dietetic Hard Candy and Chewing Gum ....................................................... 22Simultaneous Determination of Sugars and Sugar Alcohols ....................................................... 22
Nutritive Sweeteners .............................................................................................................................. 23Impurities in Sweeteners .................................................................................................................. 23Sugars in Molasses ............................................................................................................................ 23Sugars in Foods ................................................................................................................................. 24Sugars in High-Fat Foods ................................................................................................................. 24
Determining Authenticity or Adulteration with Sugar and Oligosaccharide Profiles .................... 25Coffee Adulteration ........................................................................................................................... 25Oligosaccharide Profiling of Beverages and Sweeteners .............................................................. 26Establishing Geographic Origin ...................................................................................................... 26
TABLE OF CONTENTS
Fermentation Monitoring ....................................................................................................................... 27Sugar and Oligosaccharide Profiles During Beer Production ...................................................... 27
Alternative Sweeteners, Bulking Agents and Fat Substitutes ............................................................ 28Sucralose ............................................................................................................................................. 28Inulin Products .................................................................................................................................. 28Artificial Sweetener from Japan ....................................................................................................... 29Maltodextrins ..................................................................................................................................... 29Amylopectins ..................................................................................................................................... 30
Fruit and Fruit Juice ................................................................................................................................ 31Sugars in Orange Juice ...................................................................................................................... 31Oligogalacturonic Acids from Citrus Pectin ................................................................................... 31
CHAPTER FIVE: DIONEX LC TECHNOLOGIES ......................................................... 33Column Technologies ............................................................................................................................. 35Detector Technologies ............................................................................................................................. 36
Suppressed Conductivity Detection ............................................................................................... 36Pulsed Amperometric Detection ..................................................................................................... 36
Pump Technology ................................................................................................................................... 37
CHAPTER 6: ACCELERATED SOLVENT EXTRACTION (ASE®) ................................. 39Overview of ASE Technology ................................................................................................................ 41
ASE System Features ........................................................................................................................ 42ASE 100 Accelerated Solvent Extractor ........................................................................................... 42ASE 200 Accelerated Solvent Extractor ........................................................................................... 42ASE 300 Accelerated Solvent Extractor ........................................................................................... 42
Extraction of Pesticides from Grains ..................................................................................................... 43Study No. 1: Pesticides ..................................................................................................................... 43Study No. 2: Pesticides, Herbicides, and Fungicides .................................................................... 44
Extraction of Organochlorine Pesticides From Fruits and Vegetables .............................................. 45Extraction of Organophosphorus Pesticides from Baby Food ........................................................... 45Selective Extraction of PCBs from Fish Tissue ..................................................................................... 44
A Comparison of “Nonselective” and “Selective” ASE Extractions ............................................ 47PCBs in Large-Volume Fish Tissue Samples ........................................................................................ 48Extraction of PCBs From Oyster Tissue ................................................................................................ 49
Sample Preparation and Analysis ................................................................................................... 49Analysis and Quantification ............................................................................................................ 49
TABLE OF CONTENTS
TABLE OF CONTENTS
Determination of Fat in Various Food Matrices ................................................................................... 50Comparison of ASE to Soxhlet Method .......................................................................................... 50Comparison of ASE to Mojonnier Method ..................................................................................... 51Extraction of Fat from Chocolate ..................................................................................................... 52Determination of Fat in Dried Milk Products ................................................................................ 52Extraction of Fat from Liquid Dairy Products ............................................................................... 53
Extraction of Oils From Oilseeds ........................................................................................................... 54Comparison of ASE to the Current Official Method ..................................................................... 54
APPENDIX ONE: AOAC INTERNATIONAL, OFFICIALLY APPROVEDHPLC METHODS .................................................................................................. 55
APPENDIX TWO: RECOMMENDED READING ......................................................... 59Journal Articles ........................................................................................................................................ 60Dionex Presentations, Application Notes, and Technical Notes ........................................................ 64
INDEX ...................................................................................................................... 67
1
The development of rapid, automat-
able methods for analyses of foods and
beverages is one of the most challenging
areas of analytical chemistry. Food and
beverage matrices can be very complex,
and analytes of interest may only be
present at trace levels. Methods are often
prone to interferences due to lack of
detector specificity and sensitivity, and
complex sample cleanup procedures may
be required.
The examples presented in Chapters
1–4 show analytical solutions for the
determination of carbohydrates, organic
acids, amines, and inorganic ions in
foods and beverages. The methods
employed are simple, direct, and inter-
ference-free, and require only minimal
sample cleanup. These solutons
achieved by combining specific, high-
sensitivity detection with column
selectivities tailored to the analytes.
Chapter 5 provides a brief overview of
these technologies.
INTRODUCTION
SIMPLER, INTERFERENCE-FREEANALYSIS
is the result of
High-Sensitivity DetectionSamples can be diluted 100–10,000-fold
for analysis, which greatly reduces theconcentration of matrix interferencesand the potential for column fouling.
plus
Specific DetectionAnalytes of interest can be detected at lowlevels even in the presence of much higher
levels of matrix components. Extensivesample cleanup is reduced or eliminated.
plus
Analyte-Specific SeparationsHigh-resolution ion-exchange and mixed-mode columns with selectivities tailoredto specific classes of analytes providein-situ cleanup by eliminating other
compound classes.
Chapter 6 focuses on Accelerated
Solvent Extraction (ASE®), a technique
introduced by Dionex in 1995 that
provides fast, automated extraction of
food matrices and requires only small
amounts of solvent.
2
OFFICIALLY APPROVED IC METHODS
The methods listed below employ either suppressed conductivity or pulsed
amperometric detection. See Appendix I for "AOAC International: Approved
HPLC Methods". All methods listed are suitable for Dionex systems, columns,
and reagents.
AOAC INTERNATIONAL OFFICIAL METHODS BOARD, 1ST ACTION APPROVALAOAC Method 993.30: Determination of Inorganic Anions in Water by Ion ChromatographyAOAC Method 996.04: Determination of Sugar in MolassesAOAC Method 995.13: Determination of Carbohydrates in Soluble (Instant) Coffee:
Anion-Exchange Chromatographic Method with Pulsed Amperometric DetectionAOAC Method 997.08: Determination of Fructans in Food Products
ANALYTICA-EBC INTERNATIONAL METHODDetermination of Anions in Beer by Ion Chromatography
Collaboratively Tested and Approved by the American Society for Brewing Chemists,the European Brewing Convention, and the Brewery Convention of Japan
INTERNATIONAL ORGANIZATION FOR STANDARDIZATIONISO 11292: Instant Coffee: Determination of Free and Total Carbohydrates—Method by
High Performance Anion-Exchange ChromatographyISO 10304-1: Anions in Natural and Contaminated Waters
INTERNATIONAL COMMISSION FOR UNIFORM METHODS OF SUGAR ANALYSIS (ICUMSA)Determination of Sugar in Molasses
AMERICAN SOCIETY FOR TESTING MATERIALS (ASTM)ASTM D4327-91: Anions in Water by Chemically Suppressed Ion Chromatography
U. S. NATIONAL INSTITUTE FOR OCCUPATIONAL SAFETY AND HEALTH (NIOSH)NIOSH 4110: Determination of Anions by Ion Chromatography
3
UNITED STATES ENVIRONMENTAL PROTECTION AGENCY (U.S. EPA)Method 218.6: Determination of Dissolved Hexavalent Chromium in Drinking Water,
Groundwater, and Industrial Wastewater Effluents by Ion ChromatographyMethod 300.0: Determination of Inorganic Anions in Water by Ion ChromatographyU.S. EPA Method 300.1: The Determination of Inorganic Anions in Drinking Water
by Ion Chromatography ........................................................................................ 0983U.S. EPA Method 314.0: Determination of Perchlorate in Drinking Water Using
Ion Chromatography ............................................................................................. 1195U.S. EPA Method 317.0: Determination of Inorganic Oxyhalide Disinfection By-Products
in Drinking Water Using Ion Chromatography with the Addition of Postcolumnreagent for Trace Bromate Analysis ...................................................................... 1259
5
CHAPTER ONE:INORGANIC ANIONS
AND CATIONS
6
The determination of inorganic and organic anions and cations in food and
beverages is important for meeting nutritional labeling requirements and for
processing and quality control. In many cases, the concentration levels of certain
ions have a direct bearing on food quality and flavor. Health concerns associated
with ions such as nitrite, bromide, bromate, iodide, cyanide, and chromium (VI)
necessitate their determination at trace levels in many foods and beverages. Ion
chromatography (IC) offers the food chemist multiple capabilities for automated
analysis of ionic and ionizable compounds, while eliminating time-consuming
sample preparation steps.
MUNICIPAL DRINKING WATERWater is an important raw material for
many beverages and food products. Theanalysis of incoming water is importantboth for quality control and flavor consid-erations, and for nutritional and healthconcerns. Nitrate and nitrite—present inmany water sources—are particularlyproblematic since nitrate can be reduced tonitrite in the human body and ultimatelyform carcinogenic nitrosamines by reactionwith amines. Nitrate and nitrite can bedetermined at low-µg/L (ppb) levels indrinking water either by ion chromatogra-phy with UV detection or by IC with sup-pressed conductivity detection accordingto United States Environmental ProtectionAgency (U.S. EPA) Method 300.0. IC analy-sis of municipal drinking water from twodifferent locations illustrates the significantvariations in ion concentrations thatcan occur.
7
75
0
µS3
5
0 5 10
0.0
2.5
µS 1
2 5
Minutes0 5 10
12 4
5
6
2.5
0.0
µS
Minutes
75
0
µS
Column: IonPac® AS12A and guardEluent: 2.7 mM sodium carbonate/
0.3 mM sodium bicarbonateFlow Rate: 1.5 mL/minInj. Volume: 10 µLDetection: Suppressed conductivity
10046
Peaks: 1. Fluoride2. Carbonate3. Chloride4. Bromide5. Nitrate6. Phosphate7. Sulfate
1
3
75
Union City Water Sunnyvale Water
30:1 Expanded Scale 30:1 Expanded Scale
Municipal Drinking Water
0 5 10Minutes
0 5 10Minutes
INORGANIC ANIONS & CATIONS
7
BROMATE IN DRINKING WATERAND BAKED GOODS
Bromate, a by-product of the ozonationdisinfection process for drinking water, hasbeen cited by the U.S. EPA and the WorldHealth Organization as a potential carcino-gen, even at low-µg/L concentrations.Bromate is also commonly used as a doughstabilizer for bread and other baked goods,and trace amounts may remain in the finalproduct. In both of these cases, trace levelsof bromate must be determined in the pres-ence of much higher concentrations ofchloride and other common inorganic ions.The IonPac AS9-HC column was developedspecifically to provide the high capacity andselectivity needed to quantitatively deter-mine bromate at low-µg/L concentrationlevels using a simple isocratic procedure.
CATIONS IN MINERAL WATERAND DRINKING WATER
Group I and Group II cations are easilyand rapidly determined by IC as shown forvarious water samples, even when thesodium concentration is very high relativeto ammonium and other cations present.
CATIONS IN SOFT DRINKS AND WINEGroup I and II cations are easily deter-
mined in soft drinks and other beveragessuch as wine. The high specificity of sup-pressed conductivity detection providesa very simple chromatogram, free frominterferences. Sample preparation is straight-forward—simply vacuum degas if thebeverage is carbonated, and then dilute.
INORGANIC ANIONS & CATIONS
0 5 10 15
3
2
4
5 6
Minutes
1
3
2 4
5 6
Mineral Water (310:1)* Drinking Water (680:1)*
8108-02
Column: IonPac CS12Eluent: Methanesulfonic acid,
step change at 5.1 minFlow Rate: 2 mL/minInj. Volume: 25 µLTrap Column: CTC-1Detection: Suppressed conductivity
Peaks: 1. Lithium2. Sodium3. Ammonium4. Potassium5. Magnesium6. Calcium
* Sodium:Ammonium Ratio
Cations in Mineral Water and Drinking Water
µS µS
0 5 10 15Minutes
10751/10747
Column: IonPac CS12 and guard,Cation Trap
Eluent: Water/methanesulfonic acid gradient
Flow Rate: 1 mL/minInj. Volume: 25 µL
Cations in Soft Drinks and Wine
1
2
3
4
5
10
µS
0
A Diet Cola (1:10)
0 2 4 6 8 10 12
4 5
1
23
0
µS
10
Minutes
B White Bordeaux Wine (1:160
0 2 4 6 8 10 12Minutes
Detection: Suppressed conductivity
Peaks: 1. Sodium2. Ammonium3. Potassium4. Magnesium5. Calcium
Peaks:1. Fluoride 1.0 mg/L2. Chlorite 0.013. Bromate 0.0054. Chloride 50.05. Nitrite 0.16. Bromide 0.017. Chlorate 0.018. Nitrate 10.09. o-Phosphate 0.1
10. Sulfate 50.0
Sample also contained 150-mg/L bicarbonate
0.4
µS
0.0
0
1
23
4
5
67
8
9
10
10Minutes 12952
20 30
Common Anions and Oxyhalides
Columns: IonPac AG9-HC and AS9-HCEluent: 9 mM sodium carbonateInj. Volume: 200 µLDetection: Suppressed conductivity, ASRS®,
AutoSuppression® ext. water mode
8
TRANSITION METALS IN FOODTransition metals in food products are
often determined by AA or ICP spectroscopy.However, IC with postcolumn derivatiza-tion and visible absorbance detection is anattractive alternative that also provides in-formation on speciation. An extension ofthis technique incorporates Chelation IC,a matrix elimination, and preconcentrationprocedure, which eliminates interferencefrom calcium and magnesium, elementsthat are often problematic with AA and ICPspectroscopic techniques. Chelation IC al-lows determination of transition metalsat the part-per-billion level in matricescontaining high levels of other ions.
SULFITE IN DRIED APRICOTSulfite is commonly used as a preserva-
tive in many foods and beverages. Levelsare closely regulated in many countriesbecause of reported allergic reactions exper-ienced by some individuals. U.S. Food andDrug Administration regulations requirethat any product containing 10 mg/kg ormore sulfite must be labeled as such. Thedetermination shown here is based on theprocedure of Kim and Kim1; however, pulsedrather than dc amperometric detection wasused because it provides greatly improvedreproducibility of detector response. Sampleswere diluted and homogenized with theeluent and then filtered prior to injection.
INORGANIC ANIONS & CATIONS
1Kim, H. J.; Kim, Y. K. J. Food Sci. 1986, 51, 1380.
11853
Sulfite in Dried Apricot
Flow Rate: 1 mL/minInj. Volume: 50 µLDetection: Pulsed amperometry,
Pt electrodePeaks:1. Mannitol —2. Sulfite 10.4 mg/L
Sample Preparation:20 g dried apricot blended in 100 mL mannitol buffer.Sulfite conc. 0.8 mg/g of dried apricot sample.
Column: IonPac ICE-AS1Eluent: 20 mN Sulfuric acid
200
nC
Minutes0 1263 9
300
1
2
Column: IonPac CS5A, CG5AEluent: MetPac™ PDCAFlow Rate: 1.2 mL/minInj. Vol.: 50 µLDetection: Absorbance, 530 nm,
with PAR in MetPacPostcolumn ReagentDiluent0.2
AU
0
0 2 4 6 8 10 12 14
1
2
3
45
6
7
Minutes
8
Peaks:1. Iron (III) 1.3 mg/L 2. Copper 1.3 3. Nickel 2.64. Zinc 1.35. Cobalt 1.36. Cadmium 6.07. Manganese 2.68. Iron (II) 1.3
Transition Metals in Food
11873
9
INORGANIC ANIONS & CATIONS
IODIDE IN WHOLE MILKThe determination of inorganic ions
such as nitrite, nitrate, and iodide in dairyproducts is particularly important becauseof potential health implications. Simpleprotein precipitation procedures followedby ion chromatography usually provide agood analytical solution. The determinationof iodide in whole milk in the low-µg/Lrange is shown here.
NITRATES AND NITRITES IN HAMCommonly used methods for deter-
mining nitrate and nirite in food are timeconsuming and involve a series of samplepretreatment steps using protein precipitat-ing reagents or solid-phase extraction(SPE) cartridges.
The method used here greatly simpli-fies sample preparation. Homogenizedmeat samples are extracted with water at70–80 °C for 15 min, and then centrifugedand filtered. An aliquot of the filtrate isinjected directly without further cleanup.At the end of each run, a 5-min wash with100 mM sodium hydroxide prevents col-umn fouling.
The IonPac AS11 column is ideal forthis application because it provides selec-tivity, not only for the separation of nitrateand nitrite, but also for the separation ofthe analytes from potentially interferingUV-absorbing matrix components, thatelute close to the column void.
10926
0 2 4 6 8 10
12
0
30
nA
Minutes
Iodide in Whole Milk
Column: IonPac AS7 and AG7 guard
Eluent: 200 mM Nitric acidFlow Rate: 1.5 mL/minInj. Volume: 100 µLDetection: DC Amperometry,
Pt electrode, 0.8VPeaks: 1. Iodide 38.5 µg/L*
2. Unidentified*Milk diluted 1:4. Actual conc. in milk sample is 154 µg/L.
0 5 10
0.02
0
AU1
2
Minutes 12622
Peaks:1. Nitrite 1.16 mg/L2. Nitrate 0.54
Nitrate and Nitrite in Ham
Column: IonPac AS11Eluent: 5 mM Sodium
hydroxideFlow Rate: 1 mL/minInj. Volume: 25 µLDetection: UV, 225 nm
Sample Preparation:Homogenize 10 g of sample with 100 mL of water. Heat to 75 °C for 15 min. Centrifuge. Filter through1.2 µm filter, then 0.2 µm filter.
10
POLYPHOSPHATESPolyphosphates are widely used addi-
tives in products such as fruit juices andcanned goods to prevent discoloration andoff-flavors. They are also used for curingham, tenderizing vegetables, as emulsionstabilizers for cheese, and to retain mois-ture in frozen entreés. The functionality ofpolyphosphates in these applications isstrongly dependent on their sequesteringpower and buffering capacity, which isrelated to the polyphosphate chain length.
Commercial polyphosphates are mix-tures of polyphosphates with differentchain lengths. The most widely acceptedmethod for characterizing these productshas been to determine average chain lengthby end-group titration.
Microbore ion chromatography is in-creasingly being adopted for lot-to-lotquality control and for identification ofpolyphosphate products in unknownsamples because it provides a “fingerprint”of the actual chain-length distribution.The chromatographic profiles shown are oftwo 50% sodium hexametaphosphatesolutions that were prepared from the samelot of dry powder but with producedcheese products having significantly differ-ent characteristics.2 These chromatogramsindicated a different degree of hydrolysis ata critical point in the processing and pin-pointed a problem with solution handling.
INORGANIC ANIONS & CATIONS
B “GOOD” BATCH
2 Reproduced from Baluyot, E.; Hartford, C.G.J. Chromatogr., A. 1996, 739, 217–222.
A “BAD” BATCH
Phosphates in Cheese Products
Column: IonPac AS11 (2 mm), AG11 guard (2 mm), and ATC trap (2 mm)
Eluent: Sodium hydroxidegradient
Flow Rate: 0.3 mL/minInj. Volume: 10 µLDetection: Suppressed con-
ductivity, ASRS,AutoSuppression
recycle modePeaks: 1. PO4
2. P2O73. P3O94. P3O105. P4O126. P4O13
11
CHAPTER TWO:ORGANIC
ACIDS
12
ORGANIC ACIDS
Organic acids are important flavor components in foods and also can be
indicators of product quality or deterioration due to storage. Trace levels of organic
acids in foods are best determined by ion chromatography because suppressed
conductivity detection is approximately 10 times more sensitive than low UV
detection. Inorganic ions and organic acids in food and beverage products can
usually be determined in the same analysis.
ORGANIC ACIDS IN FRUIT JUICEProfiling of the organic acids in fruit
juices is important both for establishingfreshness and detecting adulteration. Ra-tios of certain organic acids are often deter-mined because they are characteristic of aparticular juice.
Organic Acids in Cranberry Juice andTomato Juice
Quinic acid is a specific marker forcranberry juice and is used as a measure ofpurity and authenticity. A simple isocraticseparation by ion exclusion chromatographyand detection by suppressed conductivityprovides a rapid method to determinequinic acid.
Other fruit juices also show character-istic organic acid profiles that can be usedto monitor product purity.
Analysis of food products such as tomatojuice are greatly simplified since the highconcentration of salt does not interfere.Chloride elutes in the void along with otherinorganic ions. Only dilution and filtrationof the sample is required.
9990/11397
A StandardsPeaks: 1. Oxalate 2. Tartrate 3. Citrate 4. Malate 5. Glycolate 6. Formate 7. Lactate 8. HIBA 9. Acetate 10. Succinate 11. Fumarate 12. Propionate 13. Glutarate
Organic Acids in Cranberry Juice and Tomato Juice
Column: IonPac ICE-AS6Eluent: 0.4 mM hepta-
fluorobutyric acidFlow Rate: 1.0 mL/minInj. Volume: 50 µLDetection: Suppressed conductivity
0 5 10 15 20 25 30 35
30
µS
0
3
12
4
Minutes0 5 10
0 5 10 15 20 25 30 35 40
15 20
5
µS
0
12
µS
0
2 56
4
3
75
4
8
6
11 1213
109
12
3
B Cranberry JuicePeaks: 1. Oxalate 2. Tartrate 3. Citrate 4. Quinate
C Tomato JuicePeaks: 1. Inorganics
2. Malonate3. Citrate4. Malate5. Formate6. Unknown
Minutes
Minutes
13
ORGANIC ACIDS
Simultaneous High-Resolution Profilingof Organic Acids and Inorganic Anionsin Fruit Juice
As shown here for orange juice, grapejuice, and apple juice, high-resolutionprofiles of major and minor organic acids,as well as inorganic anionic components,can be determined simultaneously usinggradient high-performance anion-ex-change chromatography. The IonPac AS11column functionality is tailored for gradi-ent elution with a sodium hydroxide elu-ent and can be reequilibrated to initialconditions in approximately 5 min. Metha-nol is incorporated in the eluent to opti-mize selectivity for separation of certainorganic acid pairs.
8508A/8589/11398
0 2 4 6 8 10 12 14 16
4
7
15
17
13
2 56 8
1916910
14
11
18
20
115
µS
02
34
5
6
7 8
9
12
13
10
15
14
1617
2018
5
µS
0
5
µS
0
1214
1615
17 20
11
10
987
645
21 3
Minutes
C Apple Juice
B Grape Juice
A Orange Juice
Peaks: 9. Glutarate10. Succinate11. Malate12 Malonate
13 Tartrate14. Sulfate15. Oxalate16. Phosphate17. Citrate18. Isocitrate19. cis-Aconitate20. trans-Aconitate
Organic Acids in Orange Juice, Grape Juice,and Apple Juice
Column: IonPac AS11Eluents: Sodium hydroxide/
Methanol gradientFlow Rate: 2.0 mL/minDetection: Suppressed conductivityPeaks: 1. Quinate
2. Lactate3. Acetate4. Glycolate5. Formate6. d-Galacturonate7. Chloride8. Nitrate
0 2 4 6 8 10 12 14 16Minutes
0 2 4 6 8 10 12 14 16Minutes
14
ANIONS AND ORGANIC ACIDS IN ANIRISH STOUT
Organic acids and inorganic anions areimportant flavor constituents in brewingliquors. Inorganic anions also affect physi-cal appearance. A complete high-resolutionprofile of both organic acids and inorganicanions in beer can be obtained in less than18 min.
ORGANIC ACIDS
FOOD DYESSynthetic food dyes are widely used in
foods and beverages. They usually fall intoone of four classes: azo (mono-, di-, and tri-),indol, triphenylmethane, and methin dyes.For the most part, these dyes are acidic oranionic, and contain sulfonate, carboxyl,or phenolic groups. These strongly ioniccompounds are not easily separated byconventional reversed-phase HPLC andrequire the use of ion-pairing reagents. Byusing a multiphase column (as shown),excellent separations are achieved withoutinvoking the use of ion-pairing reagents.
Column: OmniPac® PCX-500Eluent: Perchloric acid/
Sodium perchlorate/Acetonitrile gradient
Detection: UV, 254 nmPeaks: 1. Indigo carmine
2. Orange G3. Tropaeolin O4. Orange I5. Alizarian red S6. Orange II
5399-020 5 10 15 20
Minutes
1
2
3
4
5
6
7 8
9
10
11
12
1314
15
16
17
Peaks: 7. Chrome azurol S8. Acid blue 409. Thymol blue
10. Acid blue 11311. Fluorescein 12. Methyl green13. Acid red 11414. Acridine orange15. Nile blue16. Rhodamine B17. Malachite green
Food Dyes
107450 2 4 6 8 10 12 14 16 18
16
171 236
1454 8
910
1112
13 15
714
µS
0
Minutes
Peaks: 1. Fluoride2. Lactate3. Acetate4. Formate5. Unknown6. Pyruvate7. Chloride8. Nitrate9. Unknown
10. Unknown11. Succinate12. Malate13. Maleate14. Sulfate15. Oxalate16. Phosphate17. Citrate
Column: IonPac AS11 & AG11 guardEluent: Sodium hydroxide/ methanol gradientFlow Rate: 2 mL/minInj. Volume: 25 µLDetection: Suppressed conductivity
141312111098
4
µS
08
9
1112
13
14
15
10
Anions and Organic Acids in an Irish Stout
7
15
CHAPTER THREE:AMINES AND OTHER
ORGANIC BASES
16
AMINES & OTHER ORGANIC BASES
The determination of amines in foods and beverages is important because
evidence shows that amines can act as precursors in the formation of carcinogenic
nitrosamines, and may be indicators of food spoilage.
CATIONS AND METHYLAMINESLow-molecular-weight amines such as
trimethylamine and dimethylamine areindicators of quality in fish and other foodproducts. By tailoring the column selectivity,low-molecular-weight amines and inor-ganic cations can be determined simulta-neously. Methyl-, dimethyl-, and trimethyl-amines are all resolved from the commoncations with a run time of approximately12 min.
INORGANIC CATIONS, CHOLINE,AND ACETYLCHOLINE
Choline is essential to proper metabo-lism and is often added to infant formulaand vitamin formulations. Separation ofcholine using silica reversed-phase HPLCwith ion-pairing and detection by low UVis shown in Panel A. Ion chromatographyprovides an alternative column selectivityand has the advantage of highly sensitiveand specific detection. Nonionic UV absorb-ing matrix components that are oftenpresent in food samples do not interfere.Panel B shows a separation of choline andacetylcholine by cation exchange usingsuppressed conductivity detection. Theelution order of the choline and acetyl-choline has been reversed, and sodium andpotassium are determined simultaneously.
Peaks:1. Lithium 0.5 mg/L2. Sodium 23. Ammonium 2.54. Methylamine 105. Dimethylamine 106. Potassium 57. Trimethylamine 308. Magnesium 2.59. Calcium 5
Column : IonPac CS14Eluent: 10 mM methanesulfonic
acid/0.3% acetonitrileFlow Rate: 1 mL/minInj. Volume: 18 µLDetection: Suppressed conductivity
0 5 10 15Minutes
98
7
65
4
3
2
1
5
µS
0
8919-01
Cations and Methylamines
0 5 10 15 20
1
2
3
4
430.2
AU
0.0
2
µS
0
Minutes 7130
A Reversed Phase HPLCColumn: C-18Eluent: 5 mM heptanesulfonic
acid, pH 4.0/1% acetonitrile
Detection: UV, 190 nmInjection: Choline, acetylcholine
(10 µg each)
B Cation Exchange ICColumn: OmniPac PCX-100Eluent: 75 mM hydrochloric
acid/1% methanolDetection: Suppressed
conductivityInjection: Choline,
Acetylcholine (100 ng each)Sodium, potassium (10 ng each)
Peaks(A & B): 1. Sodium
2. Potassium3. Choline4. Acetylcholine
A Reversed- Phase HPLC
B Cation- ExchangeIC
Inorganic Cations, Choline, and Acetylcholine
0 5 10Minutes
17
FLAVOR CONSTITUENTS AND ADDITIVESNaturally occuring alkaloids such as
caffeine, theophylline, and theobromine areimportant bitter flavor constituents in coffee,tea, cocoa, and cola-type beverages. Thisseparation of 10 alkaloids was performed ona multiphase polymeric pellicular columnthat gives different retention characteristicsto a typical C-18 reversed-phase separation.The difference in selectivity may provide abetter separation from potentially interferingmatrix components.
AMINES AS INDICATORS OF SEAFOOD SPOILAGE
Biogenic amines in fish are used as in-dicators of quality and spoilage. VariousHPLC methods have been developed, butderivatization techniques are requiredbecause of the lack of a suitable chromophore.An improved method developed by agroup at the Laboratorio Alimenti, InstitutoSuperiore di Sanità in Rome allows biogenicamines to be determined directly at µg/Llevels without derivatization by usingintegrated pulsed amperometry. Panel Ashows the chromatogram of aminesextracted from spoiled canned herrings;Panel B shows the same extract spikedwith 300 µg/g of each amine.
AMINES & OTHER ORGANIC BASES
7023-020 5 10
98
21
4
35
6
7
Minutes
Peaks: 1. Theobromine2. Theophylline3. Caffeine4. Morphine5. Colchicine6. Strychnine7. Papaverine8. Nicotine9. Cinchonine
10. Quinine10
Column: OmniPac PCX-500Eluent: Hydrochloric acid/
Potassium chloride/Acetonitrile gradient
Flow Rate: 1.0 mL/minDetection: UV, 254 nm
Flavor Constituents and Additives
10758Minutes0 5 10 15 20
1
23
45
4321
3
µC
0
3
µC
0
Column: IonPac CS10 and guard
Eluent: Acetonitrile/Perchloric acid/Sodium perchlorategradient
Flow Rate: 1.0 mL/min
Reproduced with permission from Draisci, R., et al., Chromatographia 1993, 35, No. 9–12, 584–590.
Amines as Indicators of Seafood Spoilage
A Canned Herring Sample
B Spiked Sample (300 µg/g of each amine)
Detection: Integrated amperometryPeaks: 1. Putrescine 16 µg/g 2. Histidine 103 3. Cadaverine 187 4. Histamine 172 5. Spermidine 294
5
Minutes0 5 10 15 20
18
AMINES & OTHER ORGANIC BASES
Column: OmniPac PCX-500Eluent: Hydrochloric acid/
Ammonium acetate/Acetonitrile gradient
Flow Rate: 1.0 mL/minUV, 254 nm
0 5 10 15
0.2
0.0
AU1
2
3 45
8
6
7
Minutes 7214
Peaks: 1. Picloram2. Aminen3. Simazine4. Atrazine5. Propanine6. Prometryn7. Trifluralin,
Planavin Component8. Planavin
Triazine HerbicidesTRIAZINE HERBICIDES IN RAW FRUITSAND VEGETABLES
Due to their widespread use, triazineherbicides are typically monitored in foodssuch as raw fruits and vegetables by usingmultiresidue methods. Both gas chroma-tography (GC) and C-18 silica reversed-phase HPLC methods have been devel-oped. Potential interferences vary for eachfood product, and a different column selec-tivity may be advantageous for a particularmatrix. The common triazine herbicidesshown here were separated on a multi-phase cation-exchange reversed-phasepolymeric column that provides alternativeselectivity to silica reversed-phase. Theeluent used in this separation is compatiblewith mass spectrometry if positive peakidentification is required.
WATER-SOLUBLE VITAMINSVitamin assays in foods are needed for
nutritional labeling, quality assurance, andmonitoring changes due to processing,storage, and so on. HPLC using silicareversed-phase with ion pairing is mostcommonly used for the determination ofwater-soluble vitamins. Separations can beachieved on a multiphase column withoutrequiring ion-pairing reagents—this alsopermits two analytical determinations tobe combined into a single run, as shown inPanels A and B, by connecting UV andconductivity detectors in series.
11.5
µS
0.0
0.65
AU
0.00
12
3
7
56
8
9114
10
Na+
K+ Mg2+Ca2+
7027-02
Column: OmniPac PCX-500Eluent: Hydrochloric acid, DAP/
Acetonitrile gradientDetection: A: UV, 215 nm
B: Conductivity
Peaks: 1. Ascorbic acid2. Pantothenic acid3. Riboflavin4. Biotin5. Niacin6. Niacinamide7. PABA8. Adenine9. Folic acid
10. Pyridoxamine11. Thiamine
Inorganic Cations and Water-Soluble Vitamins
A UV Detection of Water-Soluble Vitamins
B Conductivity Detection of Inorganic Cations
0 5 10 15Minutes
0 5 10 15Minutes
19
CHAPTER FOUR:CARBOHYDRATES
20
Carbohydrates are important constituents in many food and beverage products
and are determined for a variety of reasons, including quality control; monitoring
of food-labeling claims; establishing authenticity; analysis of sweeteners, bulking
agents, and fat substitutes; and fermentation monitoring in the production of
alcoholic beverages.
HPLC on aminopropyl-bonded silica and polymeric phases, or metal-loaded
cation-exchange resins in conjunction with refractive index (RI) or low-wave-
length UV detection, often provide simple isocratic methods for determining
common sugars. The need for improved methods has been recognized3 because
these approaches are unsatisfactory for some applications due to inadequate
resolution of sugars from sugar alcohols and organic acids, lack of detector
specificity, and insufficient sensitivity. Improved methods are particularly
important for nutritional labeling since total sugar content must be stated.
Sodium chloride interference and the use of acetonitrile have also been cited as
additional problems.3
High-performance anion-exchange chromatography at high pH coupled with
pulsed amperometric detection (HPAE-PAD) solves these problems. Sugars,
sugar alcohols, oligo-, and polysaccharides can be separated with very high
resolution in a single run without derivatization, and quantified down to picomole
levels. The technique is being applied to a wide variety of routine monitoring and
research applications and official methods have been approved by the International
Standards Organization and other official regulatory agencies. Alcohols, glycols,
and aldehydes can also be determined with this technique.
CARBOHYDRATES
3 De Vries, J. W.; Nelson, A. L. Food Technology 1994, July, pp 76–77.
21
CARBOHYDRATES
107440 10 20 30 40 50
12
10
11
678 9
5
4
30.14
µC
0.00
Minutes
Column: CarboPac MA1Eluent: 500 mM sodium hydroxideFlow Rate: 0.4 mL/minInj. Volume: 10 µL
Detection: Pulsed amperometry
Peaks: 1. myo-Inositol2. Glycerol3. i-Erythritol4. Xylitol5. Arabitol6. Sorbitol7. Dulcitol8. Mannitol9. Glucose
10. Fructose11. Sucrose
Sugar Alcohols
Column: Amino-PropylEluent: 75% acetonitrile/
WaterFlow Rate: 2.0 mL/minDetector: Refractive indexTemp.: 25 °CPeaks: 1. DP-1
2. DP-23. DP-34. DP-45. DP-5
0 5
1
2
34 5
0 5 10 15Minutes
123
4
5
8576-8579
0 10
1
2
34 5
Column: Reversed-phaseEluent: WaterFlow Rate: 0.5 mL/minDetection: Refractive indexTemp.: Ambient
Column: CarboPac PA1Eluent: Sodium hydroxide/
Sodium acetate gradient
Flow Rate: 1.0 mL/minDetector: Pulsed amperometryTemp.: Ambient
0 8 12
54
32
1
Column: Cation-exchange, Calcium form
Eluent: WaterFlow Rate: 0.6 mL/minDetector: Refractive indexTemp.: 80 °C
Oligo- and Polysaccharides Derived fromHydrolyzed Glucose Syrup
OLIGO- AND POLYSACCHARIDES DERIVEDFROM HYDROLYZED GLUCOSE SYRUP
This comparison of chromatographictechniques used for the analysis of ahydrolyzed glucose syrup illustrates theremarkable resolving power of theHPAE-PAD technique. Elution order isreversed with the CarboPac® PA1 columnas compared with conventional metal-loaded cation-exchange columns. Thatis the higher homologues elute later asresolved peaks rather than in a poorlyresolved group at the beginning of thechromatogram.
SUGAR ALCOHOLSNutritional labeling requirements for
sugar alcohols are presently optional in theU.S., but as with sugars, the total sugaralcohol content is listed. Minor sugaralcohols must therefore be determined.Gas chromatogarphy (GC) methods forsugar alcohols have been developed, butare complicated by the requirement forderivatization. The only official method inexistence is AOAC 973.28, a GC method forsorbitol. A simpler, more direct approachthat does not require derivatization isshown here. The selectivity of the columnwas designed to allow sugar alcohols as agroup to elute ahead of sugars with highresolution.
22
CARBOHYDRATES
12236
Column: CarboPac PA10, PA10 Guard,Borate Trap™
Eluent: 52 mM sodiumhydroxide
Flow Rate: 1.5 mL/min
Inj. Volume: 5 µLDetection: Pulsed amperometry,
gold electrodePeaks: 1. Glycerol 4.0 µg/mL
2. Xylitol 2.03. Sorbitol 1.54. Mannitol 3.05. Glucose 5.06. Fructose 8.07. Sucrose 2.08. Lactose 8.0
Isocratic Separation of Food Sugarsand Sugar Alcohols
0
123
4
5
6 78
42 6 12108 14
40
nC
Minutes
0
8587-02/8850-01
A Dietetic Hard Candy
0 10 20 30
1
3
2
4
Minutes
Column: CarboPac MA1Eluent: 500 mM sodium
hydroxideFlow Rate: 0.4 mL/minDetection: Pulsed amperometry,
Au electrode
Peaks A:1. Sorbitol 106 µg2. Mannitol 17
Peaks B:1. Glycerol2. Sorbitol3. Mannitol4. Glucose
B Chewing Gum
Sugar Alcohols in Dietetic Hard Candyand Chewing Gum
µC
2
15
00 10 20 30
Minutes
µC
Sugar Alcohols in Dietetic Hard Candyand Chewing Gum
Use of sugar alcohols as alternativesweeteners is increasing rapidly particularlyfor dietetic foods and in products such aschewing gum because of their noncariogenicproperties. In these cases, sugar alcoholsmust be routinely determined in foods tomeet regulatory requirements. Simple pro-cedures for sugar alcohols in hard candy(boiled sweets) and chewing gum areshown here. Sorbitol and mannitol can beeasily determined, interference-free, in hardcandy; sample preparation consists of dis-solution and dilution in water (Panel A).Panel B shows the determination of glycerol,sorbitol, mannitol, and glucose in a chewinggum sample; sample preparation consistsof sonication with deionized water,OnGuard® A cartridge pretreatment, follow-ed by filtration through a 0.45-µm filter.
Simultaneous Determination of Sugarsand Sugar Alcohols
Sugars and sugar alcohols commonlyfound in food and beverage samples can beroutinely determined in the same run on theCarboPac PA10 column. Under isocraticconditions, glycerol, xylitol, sorbitol, andmannitol elute rapidly followed by themono- and disaccharides.
23
CARBOHYDRATES
0 2 4 6 8 10 12
12
3
Minutes 7660
Column: CarboPac PA1 and guard
Eluent: 160 mM sodium hydroxide
Flow Rate: 1.0 mL/minDetection: Pulsed
amperometry, Au electrode
Peaks: 1. Glucose2. Fructose3. Sucrose
µA
Impurities in Sweeteners
0 2 4 6 8
1
23
4
8
µA
0
Minutes 10930
Column: CarboPac PA1 and guard
Eluent: 150 mM sodium hydroxide
Flow Rate: 1.0 mL/minInj. Volume.: 50 µLDetection: Pulsed amperometry,
Au electrodePeaks: 1. Glucose 4.39%
2. Fructose 6.673. Lactose (int. std.)4. Sucrose 30.8
Sugars in Molasses
10
NUTRITIVE SWEETENERSHPLC is in daily use in the sugar
industry to determine organic acids andcarbohydrates. Strong cation-exchangecolumns in various metal forms have beenmost commonly used; however, HPAE-PAD now offers a powerful alternativewith significant advantages.
Impurities in SweetenersThe common crystalline sugar products
—sucrose, maltose, lactose, dextrose andfructose—have highly predictable func-tionality when used in food products be-cause of their high purity. Trace impuritiesare easily and rapidly determined, asshown in this typical sucrose analysis.
Sugars in MolassesThe determination of sugars in molas-
ses using the official method of the Interna-tional Commission for Uniform Methodsof Sugar Analysis (ICUMSA) approved in1994 is shown here. Approval of themethod was based on an internationalcollaborative study involving 11 laborato-ries. Excellent reproducibility was obtainedand the results were in close agreementwith a parallel GC collaborative study.
Among the advantages cited for thenew method were: (1) lack of coelutionwith nonsugar impurities; (2) greatlyreduced possibility of overestimation ofsugars due to coeluting impurities; and(3) no column heater is required.
24
Sugars in FoodsSugars can easily be determined in foods
such as tomato ketchup by a simple extrac-tion followed by dilution and filtration, asillustrated in Panel A. In Panel B, glucose,fructose, maltose, and maltotriose contentwere determined in a butterscotch candysample. Samples were diluted 1:2000 andpassed through a 0.2-µm filter prior to injec-tion. In Panel C, sugars present in a flavoredpotato chip extract were determined directlyfollowing a simple extraction. The unidenti-fied peaks may be other sugars or easilyoxidizable ingredients such as aldehydes.
0 5 10 15 20 25
3
1
2
200
0
nC
Minutes 10753
0 10 20Minutes
1
2
3
5
1
4µC
0
10755
0 10 20Minutes
1
2
3
4
5
4133
B Butterscotch CandyColumn: CarboPac PA1Eluent : Sodium hydroxide/
sodium acetate gradient
C Flavored Potato ChipColumn: CarboPac PA1Eluent: Sodium hydroxide
Flow Rate: 1.0 mL/minDetection: Pulsed amperometry,
Au electrodePeaks: 1. Arabinose
2. Glucose3. Fructose4. Lactose5. Sucrose
Flow Rate: 1.0 mL/minInj. Volume: 25 µLDetection: Pulsed amperometry,
Au electrodePeaks: 1. Glucose 6.8%
2. Fructose 3.0% 3. Unidentified 4. Maltose 4.5%
5. Maltotriose 3.9%
Column: CarboPac PA1Eluent: 150 mM sodium hydroxideFlow Rate: 1 mL/minInj. Volume: 50 µLDetection: Pulsed amperometry, Au electrodePeaks: 1. Glucose 950 µg/mL
2. Fructose 5903. Sucrose 4.1Sample diluted 1:100 with DI water.
Sugars in Food
A Ketchup
µC
0 2 4 6 8 10
12
1.5 3
0
µC
Minutes 10172-01
Flow Rate: 1.0 mL/minInj. Volume: 50 µL (~1 g in 16 L)Detection: Pulsed amperometry,
Au electrodePeaks:1. Glycerol not quantified2. Lactose 1.3 µg3. Sucrose 2.9
Sugars in Milk Chocolate
Column: CarboPac PA1 and guard
Eluent: 180 mM sodium hydroxide
Sugars in High-Fat Foods A challenge in determining sugars in
high-fat food samples is that the fat mayinterfere with the chromatography. Thisproblem can be overcome by extracting thefat prior to analysis. Supercritical fluidextraction was used in this example.
CARBOHYDRATES
25
0 10 20 30 40Minutes
0
12
34
5 6 7
6007-02
Carbohydrates in Instant Coffee
Inj. Volume: 25 µL of 10 g/L solutionDetector: Pulsed amperometry,
Au electrode; postcolumnaddition of 0.3 M NaOH
Peaks: 1. Mannitol 21 mg/L2. Arabinose1403. Galactose 764. Glucose 445. Xylose 266. Mannose 517. Fructose 93Sample Preparation: Phenolics removed with On-Guard P cartridge.
Column: CarboPac PA1Eluent: 150 mM sodium
hydroxide/deionizedwater gradient
1000
0
nA
CARBOHYDRATES
DETERMINING AUTHENTICITY ORADULTERATION WITH SUGAR ANDOLIGOSACCHARIDE PROFILES
Coffee AdulterationThe fraudulent addition of cheaper
coffee substitutes to commercial productscan be detected by determination of freeand total carbohydrate profiles. For ex-ample, high levels of free mannitol andtotal xylose are indicative of adulterationwith coffee husks and parchments,whereas so-called “pure” soluble coffeescontaining cereals or caramelized sugarcontain high levels of glucose.
Previous methods for determination ofcarbohydrates in coffee have been limitedeither by complex sample preparation,enzyme availability, or lack of specificity.Complete carbohydrate profiles could onlybe obtained by combining results fromdifferent techniques. Using HPAE-PAD, allthe major carbohydrates present in solublecoffee can be determined in a single run.
The separation shown here employs a“reverse” gradient. A similar method hasbeen validated by a collaborative studyinvolving 11 laboratories from differentcountries and has been approved as anofficial ISO method (ISO 11292). Severalpapers have been published relating to ISO11292:
• Prodolliet, J; Bruelhart, M; Lador, F; Mar-tinez, C; Obert, L; Blanc, M. B; Parchet, J-M. “Determination of Free and Total Carbohydrate Profile in Soluble Coffee”J. Assoc. Off. Anal. Chem. Int. 1995, 78,749–761.
• Prodolliet, J; Blanc, M. B; Leloup, V;Cherix, G; Donnelly, C. M; Viani, R.“Adulteration of Soluble Coffee withCoffee Husks and Parchments”J. Assoc. Off. Anal. Chem. Int. 1995, 78,761–767.
• Prodolliet, J; Bugner, E;Feinberg, M.“Determination of Carbohy-drates in Soluble Coffee by AnionExchange Chromatography with PulsedAmperometric Detection:Interlaboratory Study” J. Assoc. Off. Anal. Chem. Int. 1995, 78,768–782.
For additional literature of interest,see Appendix Two: Recommended Reading.
26
CARBOHYDRATES
0 10 20 30
0
µA
5
11399
B Orange Juice Adulterated with Beet Medium Invert Sugar
Oligosaccharide Profiles of Pure and Adulterated Orange Juice
Minutes
Column: CarboPac PA-100Eluent: Sodium hydroxide/
Sodium acetate gradient
Flow Rate: 1 mL/minInj. Volume: 25 µLDetection: Pulsed
amperometry,Au electrode
0
µA
5A Pure Orange Juice
0 10 20 30Minutes
1
4938-01/4937-01
Sucrose
0
µA
Minutes
0
µA
1
Oligosaccharide Profile of Beet Sugar Molasses
C British Column: CarboPac PA1Eluent: 100 mM sodium
hydroxide/20 mM Sodium acetate
Flow Rate: 1 mL/minDetection: Pulsed
amperometry, Au electrode
0 5 15
D U.S.
Sucrose
Minutes0 5 15
Oligosaccharide Profiling of Beveragesand Sweeteners
The determination of oligosaccharidecomposition profiles is a powerful tech-nique for detection of adulteration of natu-ral fruit juices by inexpensive sweetenerssuch as beet sugar (Panels A and B).
Profiles can be rapidly determined, asshown in this comparison of pure orangejuice (A) and orange juice adulterated with20% beet medium invert sugar (B).
Establishing Geographic OriginOligosaccharide profiling is also a useful
tool for establishing the geographical origin ofcarbohydrate food ingredients such as molas-ses. Panels C and D show the difference inoligosaccharide profiles for beet sugar fromBritain (C) and the United States (D).
27
CARBOHYDRATES
0 5 10 15 2520Minutes
0
nC
70 2
34
5 6
78
910
11
2000
nC
0
2
34
56 7 8 9 1011
10927/11358
800
nC
0
1
23
45
6 7 8 9 10 11
Column: CarboPac PA-100 and guardEluent: Sodium hydroxide/
Sodium acetate gradientFlow Rate: 1.0 mL/minInject Vol.: 10 µLDetection: Pulsed amperometry,
Au electrode
Sugars and Oligosaccharide ProfilesDuring the Beer Brewing Process
Peaks: 1. Ethanol2. Glucose
3. Maltose 4. Maltotriose 5. Maltotetraose
6. Maltopentaose7. Maltohexaose8. Maltoheptaose9. Maltooctaose
10. Maltononaose11. Maltodecaose
C Wort
A Maltose Oligomers
D Finish
0 5 10 15 2520Minutes
1600
nC
0
2
3
4
5 6
B First Mash
0 5 10 15 2520Minutes
0 5 10 15 2520Minutes
FERMENTATION MONITORING
Sugar and Oligosaccharide ProfilesDuring Beer Production
Determining the levels of fermentableand nonfermentable sugars at every stage ofbeer production is important because fer-mentable sugars determine the final alcoholcontent, and nonfermentable sugars contrib-ute to the flavor and “body” of the finalproduct. Sugars, sugar alcohols, alcohols,and glycols can be rapidly determined withhigh resolution at all phases of beer, wine,or cider production, as shown here.
A separation of maltose oligomers upto DP10 (“DP” refers to their degree ofpolymerization) with baseline resolution isshown in Panel A, and sugar and oligosac-charide profiles at various stages of thebrewing process in Panels B, C, and D.All samples were diluted 1:10.
Similarly, oligosaccharide profiles canbe determined for comparing regular andlow calorie beers.
28
CARBOHYDRATES
0 10 20 30 40 50
1
Minutes
2
3780
nA
0
10898
Column: CarboPac MA1Eluent: 150 mM sodium
acetate/0.2% (v/v) Acetic acid, pH 5.5
Flow Rate: 0.4 mL/minInj. Volume: 25 µLDetection: Pulsed amperometryPeaks:1. 4-Cl-Galactose 7.8 mg/L2. 1,6-di-Cl-Fructose 2.43. Sucralose
Sucralose
0 10 20 30 40Minutes
10
20
50
3550/3073
3
0
µA
Column: CarboPac PA1Eluent: Sodium hydroxide/
Sodium acetate gradientFlow Rate: 1 mL/minDetection: Pulsed amperometry,
Au electrodeSample*: 0.3% Water washed
Inulin (polyfructose) in 0.1 M sodium hydroxide
*Sample courtesy of Dr. C. Mitchell. California Natural Products, Manteca, CA
Purified Inulin
β
CH2
O
H HO
OH HCH2OH
O
OO
O OO
HHOCH2
HOH2Cαβ
CH2
β
H HH
OH
HOCH2
H
HOCH2
HH HO
OH H
H HO
OH H H HO
OH H
n
ALTERNATIVE SWEETENERS, BULKINGAGENTS, AND FAT SUBSTITUTES
Increasing public concern over healthand nutrition has led to the developmentof new low-calorie sweeteners and fat sub-stitutes. Products have been developed notonly to emulate the sweetness of sucrose,or act as a fat substitute, but also to provideother important properties such as textureand bulk.
SucraloseSucralose is a new high-intensity sweet-
ener with 400–800 times the sweetness ofsucrose. Sucralose is manufactured by selec-tive chlorination of sucrose and is currentlythe only nonnutritive sweetener based onsucrose. The product was approved inCanada in September 1991 for use in avariety of food and beverage categories. Itis presently under review in other countries,including the United States and the UnitedKingdom. A sucralose sample shows traceimpurities when analyzed by HPAE-PAD.
Inulin ProductsInulin-based products derived from
chicory root and Jerusalem artichokes aremarketed as fat replacers and dietary fiberadditives to food formulations. These inulinproducts are mixtures of linear polyfructosechains incorporating a few glucose units.Their degree of polymerization (DP) istailored for a particular end use, so it iscritical to determine the chain-length distri-bution to maintain quality control of theend product. Determination of the chain-length distribution is shown here for DPvalues up to 50 and more.
29
CARBOHYDRATES
0 5 10 15
123
4
5
Minutes 4940-01/6055
0
µA
1
Column: CarboPac PA1Eluent: Sodium hydroxide/sodium acetateFlow Rate: 1 mL/minDetector: Pulsed amperometry,
Au electrodePeaks: 1. Glucose
2. Fructose3. Sucrose4. 1-Kestose5. Nystose
Kestose — Artificial Sweetener from Japan
OH
HO
HO
HOCH2
HOCH2
HOCH2 O
OHOH
OH
OH
O
O
CH2
O
CH2
O
Sucr
ose
HigherHomologs
1α
2β
2β
1-Kestose
OH
HO
HO
HOCH2
HOCH2
HOCH2 O
OHOH
OH
OH
O
O
CH2
O
CH2
O
Nystose
HO
HOCH2
OH
O
CH2OH
O
0 5 10 15 20 25 30 35 0
175
nC
Minutes
0 5 10 15 20 25 30 35 0
175
nC
Minutes
Maltodextrins — Maltrin® MO40 & M700
Inj. Volume: 25 µLFlow Rate: 1 mL/minDetection: Pulsed amperometry,
Au electrode
Maltrin is a trademark of Grain Processing Corporation.
Column: CarboPac PA10Eluent: Sodium hydroxide/
Sodium acetate gradient
A Maltrin MO40
B Maltrin M700
Artificial Sweetener from JapanKestoses improve the flavor of products
such as yogurt, and are also used as alterna-tive sweeteners with a sweetness 0.4–0.6times that of sucrose. Commercial productsare obtained from sucrose by an enzymaticprocess that results in a mixture of 1-kestose,nystose, glucose, sucrose, and fructose.HPAE-PAD provides an excellent method formonitoring the enzymatic process and forquality control of the final product.
MaltodextrinsMany commercial low-calorie bulk
sweeteners, bulking agents, and fat substi-tutes are polysaccharide or polyol materi-als derived from various types of starch.Determination of the distribution of poly-saccharide chain lengths in these productshas a direct bearing on the product func-tionality. HPAE-PAD is the method ofchoice because other approaches cannotseparate the higher DP polysaccharidechains. Chain-length distribution profilesfor two maltodextrins derived from corn-starch are shown here. Maltrin® MO40 isused in film-forming applications and toprovide a smooth texture. Maltrin M700 isprocessed to produce very low density par-ticles with excellent dissolution character-istics. The differences in physical proper-ties of the two maltodextrins are clearly re-flected in the different chain-length distri-bution profiles.
30
CARBOHYDRATES
D Edible Canna
Column: CarboPac PA1Eluent: Sodium hydroxide/sodium acetate gradientFlow Rate: 1.0 mL/minDetection: Pulsed amperometry, Au electrodeFrom Koizumi, K.; Fukuda, M.; Hizukuri, S. J. Chromatogr. 1991, 585, 233–238.
Amylopectin Chain-Length Distribution
504030
20
12
A Rice
B Corn
C Sweet Potato
6
6
12
20
3040 50 60
6
12
20
3040 50 60
0 10 20 30 40Minutes 9661
6
12
20
3040 50 60
0 10 20 30 40Minutes
0 10 20 30 40Minutes
0 10 20 30 40Minutes
AmylopectinsStarch from various sources varies in its
functional properties as a consequence ofdifferences in chemical structure. Anunderstanding of the relationship betweenmolecular structure and functional proper-ties is important for fundamental researchand in selecting and developing starch-derived additives for food formulations.Chain-length distribution is an importantparameter in characterizing both the amy-lose and amylopectin portion of starch.Shown here are complete chain-lengthdistributions up to DP60 for debranchedamylopectin from different sources.
31
FRUIT AND FRUIT JUICEHPLC is a powerful technique for the
analysis of carbohydrates, organic acids,and preservatives in fruit juice to ensureproduct quality, support nutritional label-ing claims, and detect adulteration.
Sugars in Orange JuiceAs shown, sugars can be determined
directly in juices, free of interference fromorganic acids. The only sample preparationrequired in this case is dilution and filtra-tion.
Oligogalacturonic Acidsfrom Citrus Pectin
Pectin has been used for years to thickenor gel products such as jams and jellies, butnew applications are being discovered.One specialty pectin product used as a fatsubstitute consists of partially methylatedpolygalacturonic acid extracted from citruspeel. Pectin from different sources has aunique oligogalacturonic acid “fingerprint”that can be used for identification andquality control purposes. A typical profileof oligogalacturonic acids from citrus pec-tin can be obtained in less than 40 min.Sample preparation consists of direct injec-tion of the diluted pectin hydrolysate.
CARBOHYDRATES
0 5 10 15 20 25 30 35 40 45
6
5
78 9
1210
1311 14
1516
1720
18
19
4939-01Minutes
10
µA
0
Oligogalacturonic Acids from Citrus Pectin
Column: CarboPac PA1Eluent: Sodium acetate/
Sodium hydroxide gradient
DP
Flow Rate: 1 mL/minDetection: Pulsed amperometry,
Au electrode
Sugars in Orange Juice
0 2 4 6 8 10 12
3
2
1
Minutes
0
40
nC
11400
Column: CarboPac PA1Eluent: Sodium hydroxideFlow Rate: 1 mL/minDetection: Pulsed amperometry,
Au electrodePeaks: 1. Glucose
2. Fructose3. Sucrose
33
CHAPTER FIVE:DIONEX LIQUID
CHROMATOGRAPHY
TECHNOLOGIES
34
Dionex LC instrumentation accommodates all of the conventional HPLC
techniques. New detector, column, and pump technologies
have been developed—often as a result of working with our customers to
solve specific analytical problems. Some of our most unique and effective
innovations include:
• Ion-exchange stationary phases with unique selectivities tailored to provide
fast, high-resolution separations of ionic and polar compounds.
• Metal-free HPLC system flow paths that guard against corrosion and
protein denaturation.
• Advanced suppressed conductivity detection technology for quantification
of organic and inorganic anions and cations with high specificity and
sensitivity.
• Pulsed amperometric detection (PAD) technology that greatly simplifies
the determination of carbohydrates.
• A revolutionary HPLC pumping system that integrates unique feedback
control with digital signal processing and artificial intelligence to eliminate
pulsation and provide exceptional flow accuracy—even under the changing
backpressure of gradient elution.
DIONEX LC TECHNOLOGIES
35
DIONEX LC TECHNOLOGIES
COLUMN TECHNOLOGIESResolution in HPLC is highly depen-
dent on column selectivity, which, in turn,depends on the stationary phase.
Dionex MicroBead™ high-efficiency,high-polymeric stationary phases weredeveloped specifically to optimize theseparation of specific classes of analytes.These stationary phases offer:• High-speed equilibration for gradient
elution.• Excellent mass transport characteristics
that result in higher efficiency.• Compatibility with high flow rates for
rapid separations.• Complete pH compatibility (pH 0 to 14).• High mechanical (4000 psi) and chemical
stability for exceptionally long column life.• Noncompressible resins that simplify
linear scale-up for preparative-scaleapplications.
The MicroBead stationary phase is madeby agglomerating a nonporous, noncom-pressible, polymeric substrate with beads ofquaternary-functionalized latex. The resultis a highly stable particle with a thin surfacelayer rich in ion-exchange sites that can betailored to separate specific analytes.
A variation on the MicroBead station-ary phases, also developed at Dionex, arepackings in which ion-exchange resins aregrafted in a thin layer on the surface of thecore polymer particle. Packings of this typehave been developed for applications suchas ion analysis of drinking water and wastewater.
Dionex has also developed multiphasepackings that incorporate both reversed-phase and ion-exchange functionality,which often eliminate the need for the ion-pairing reagents used in conventionalHPLC reversed-phase separations.
Figure 1Construction of IonPac’spellicular anion-exchangeresin bead:• Highly cross-linked inert,
nonporous core providesHPLC solvent compatibilityfor cleanup and formodifying eluent selectivity.
• Surface-sulfonated regioncompletely covers the core.
• Submicron pellicularMicroBead layerconcentrates a vast numberof ion-exchange sites into avery thin layer.
0.1-µmDiameter
3183-02
Core Ion-ExchangeSurface
SO3–
SO3–
SO3–
SO3–
SO3–
SO3–
SO3–
R3N+
R3N+
N R3+
N R3+
N R3+
R3N+
N R3+
N R3+
N R3+
N R3+
N R3+
N R3+
R3N+
N R3+
N R3+
R3N+
IonPac's Pellicular Anion-Exchange Resin Bead
5 µm
36
DIONEX LC TECHNOLOGIES
DETECTOR TECHNOLOGIESThe simplicity of many of the applica-
tion solutions shown in this book can, inmany cases, be attributed to the combina-tion of unique column selectivities witheither suppressed conductivity or pulsedamperometric detection.
Figure 3 Electrolysis of water within the SRS produces thehydronium and hydroxide ions required for eluent neutralization.Effluent from the detector cell is recycled to provide a continuouswater source.
8416-03
Eluent
Waste
Detector CellSRS
Recycled to SRS
ASRS Eluents: Hydroxide, carbonate/bicarbonate, boric acid/tetraborateCSRS Eluents: Methanesulfonic acid (MSA). (Eluents containingCl– or N0
3– cannot be used in this mode).
AutoSuppression Recycle Mode
Time
E1
Delay Integration
Poten
tial
E2
E3
t2
t3
5031-01
Pulsed Amperometry Triple-Pulse Sequence
t1
Figure 4 The triple-pulse potential sequence employed inpulsed amperometric detection ensures a clean electrode foraccurate, consistent detection results.
Figure 2 The principle of AutoSuppression® suppressedconductivity with the SRS® Self-Regenerating Suppressor.
–2
11309
AnalyticalColumn
Time
µS
Cl–F– SO4–2
Anion Suppressor
Waste
µS
SO4Cl–F–
Eluent (NaOH)Sample F–, Cl–, SO4
–2
H2O
NaF, NaClNa2SO4 in NaOH
Without Suppression
With AutoSuppression
The Role of AutoSuppression
Time
Suppressed Conductivity DetectionThe principle of suppressed conductivity
detection is illustrated in Figure 2. Thesuppressor reduces the eluent conductivityto a very low level while enhancing theanalyte conductivity. The result is excep-tional sensitivity and high specificity forionic analytes.
The SRS Self-Regenerating Suppressoris a self-contained unit that operates unat-tended and requires no maintenance(Figure. 3).
Pulsed Amperometric DetectionAnalytes such as carbohydrates cannot
be determined by DC amperometry due toelectrode fouling. Pulsed amperometryovercomes this problem by using a repeat-ing triple-pulse voltage sequence thatmaintains a clean electrode surface, thusensuring a consistent detector response(Figure. 4). This technique is now wellestablished for the determination ofcarbohydrates, and provides high sensitiv-ity and specificity without the need forderivatization.
37
PUMP TECHNOLOGYAccuracy and precision of retention
times and peak areas can be adverselyaffected with conventional HPLC pumpdesigns due to flow rate variations. Thesechanges result from the effect of changingsystem backpressure on the compressibilityand viscosity of the mobile phase, systemcompliance, and other factors such as mi-nor seal leakage. Flow rate variations areparticularly problematic during gradient
Primary Information Pathways of the PumpDigital Control System
DIONEX LC TECHNOLOGIES
elution where large changes in systembackpressure often occur. The Dionexquaternary gradient pumping system wasdesigned to directly address these prob-lems by using a flow control system engi-neered around state-of-the-art digital sig-nal processing (DSP) and fuzzy-logic (arti-ficial intelligence) algorithms. This engi-neering results in accurate, precise andpulse-free flow over a broad range of mo-bile phase compositions and systembackpressures.
Artificial Intelligence
Memory
MotorControlCircuit
TachCircuit
Motor
Sensor
Mechanism
EncoderDisk
Piston A&
Piston B
ProportioningValves
PressureTransducer
DSP
Eluents
To Column
39
CHAPTER SIX:ASE® ACCELERATED
SOLVENT
EXTRACTION
40
ACCELERATED SOLVENT EXTRACTION
Extraction is often a necessary first step in the analysis of food samples.
Traditional extraction methods such as Soxhlet use large volumes of solvent and
are slow, often requiring hours to obtain a satisfactory recovery. Sonication is a
faster technique, but it too requires large amounts of solvents and is not easily
automated. The high initial cost of solvents and their subsequent disposal makes
these methods less than desirable for routine use.
Accelerated Solvent Extraction (ASE) is an automated extraction method
developed by Dionex that takes advantage of the temperature dependence of
extraction kinetics and requires only small amounts of solvent. Extraction times
are reduced from hours to just minutes by operating at higher temperatures and
pressures than are typically used for traditional solvent-based extraction tech-
niques. ASE significantly streamlines sample preparation and is being rapidly
adopted for extraction of environmental, food, polymer, and other types of solid
and semisolid samples. For example, ASE meets all of the requirements of U.S.
EPA SW-846 Method 3545A for Pressurized Fluid Extraction of bases, neutrals,
and acids (BNAs), chlorinated pesticides and herbicides, polychlorinated
biphenyls (PCBs), and organophosphorus pesticides.
41
ACCELERATED SOLVENT EXTRACTION
OVERVIEW OF ASE TECHNOLOGYAccelerated Solvent Extraction is
achieved by using organic or aqueous sol-vents at elevated temperature and pressure.The solvent is pumped into an extractioncell containing the sample, which is thenbrought to a specified temperature (fromambient to 200 °C) and pressure. Followingextraction, the extract is transferred from theheated cell, through a filter, to a standardcollection vial for cleanup and/or analysis.
Oven
Collection Vial
Extraction Cell
Solvent
Pump
Vent
Load sample into cell.
Fill cell with solvent.
Heat and pressurize cell.
Hold sample at pres- sure and temperature.
Pump clean solvent into sample cell.
Purge solvent from cell with N2 gas.
Extract ready for analysis.
Schematic of the Accelerated Solvent Extraction Process
Increased temperature accelerates theextraction kinetics, while elevated pressureprevents boiling at temperatures above thenormal boiling point of the solvent. Onlysmall amounts of solvent are used andextraction times can be as short as 15 min.Time and solvent consumption are thussignificantly reduced compared to othersolvent extraction techniques. Methoddevelopment is generally straightforwardbecause the same solvent used for a Soxhletor other current methods is generally usedwith ASE.
42
ACCELERATED SOLVENT EXTRACTION
ASE System FeaturesAll ASE Systems Feature:• Automated sample extraction• Automatic extract filtration• Easy-to-fill sample cells with hand-tight
fittings• Easy-to-use collection vials and bottles• Convenient front-panel operation with
multiple method storage• Sensors for temperature, pressure, and
solvent vapors ensure safe operation atall times
• Easy method transfer between systems• Patented technology (patent numbers
5,843,311; 5,647,976; 5,660,727; and5,785,856)
• Established methodologies andEPA approval
• Temperature range from ambientto 200 °C
ASE 100 Accelerated Solvent ExtractorThe ASE 100 is the entry-level ASE
system designed for use in lower-through-put labs. This system is priced economi-cally and offers fast and efficient extractionfor a large range of sample sizes.• Automated extraction of a single sample• Sample cell sizes: 10, 34, 66, and 100 mL• Collection bottle: 250 mL• Operating pressure: 1500 psi (100 bar)• Small footprint requires less than 36 cm
(14 in.) of bench space ASE 200 Accelerated Solvent Extractor
The ASE 200 is designed for high-throughput labs with sample sizes of 1 to30 grams. With automation capabilities forup to 24 samples, the ASE 200 maximizes
sample throughput and extraction effi-ciency. The ASE 200 is ideal for standardenvironmental analysis, pharmaceuticalprocesses, and routine food and polymerapplications.• Unattended extraction for up to
24 samples• Samples cell sizes: 1, 5, 11, 22, and 33 mL• Scheduling programming for automated
method optimization• Collection vial sizes: 40 and 60 mL• Automatic rinsing of system between
sample extractions• Compatible with AutoASE® computer
control software and ASE SolventController
• Operating pressure: 500–3000 psi(30–200 bar)
ASE 300 Accelerated Solvent ExtractorThe ASE 300 is designed for high-
throughput labs with large sample volumerequirements. With automation capabilitiesfor up to 12 cells and sample cell volumesup to 100 mL, the ASE 300 is ideal for thebusy environmental and food analysis lab.• Unattended extraction for up to
12 samples• Samples cell sizes: 34, 66, and 100 mL• Collection bottles: 250 mL• Automatic rinsing of system between
sample extractions• Scheduling programming for automated
method optimization• Compatible with AutoASE computer
control software and ASE SolventController
• Operating pressure: 1500 psi (100 bar)
43
EXTRACTION OF PESTICIDES FROM GRAINSThe determination of pesticides, herbi-
cides, and other related compounds infoods and agricultural commodity productsis important to ensure the safety of the foodsupply. Increasingly, multiresidue analysistechniques are being used that require aninitial extraction of groups of pesticideswith high efficiency.
The data shown here are from collabora-tive studies conducted with two independentresearchers on the extraction of various classesof pesticides from wheat. Ground wheatsamples were sent to Dionex for extraction byASE. The extracts were then returned to theresearchers for cleanup and/or analysis.
Study No. 1: PesticidesTables 1 and 2 show the data from the
first study. Table 1 compares the currentand ASE extraction conditions used toobtain the recovery data shown in Table 2.ASE provides a more efficient extraction ina much shorter time with greatly reducedsolvent usage, and does not require a post-extraction SPE cleanup step.
ACCELERATED SOLVENT EXTRACTION
TABLE 1. PESTICIDES IN WHEATStudy No. 1: Comparison of Current
and ASE Extraction Conditions
Current Method ASESample Size 3–20 g 3–20 gSolvent Volume 130 mL 15 mLPost Extraction SPE cleanup NoneTotal Time 60 min 15 minSample Analysis GC-FPD GC-FPD
TABLE 2. PESTICIDES IN WHEATStudy No. 1: Comparison of Recoveriesfor Current and ASE Extraction Methods
Malathion Methylchlorpyrifos(µg/L) (µg/L)
Sample Current ASE Current ASEMethod Method Method Method
1 40 50 70 90
2 40 50 80 100
3 60 70 50 60
5 40 100 30 70
10 60 80 60 80
11 60 70 70 90
44
ACCELERATED SOLVENT EXTRACTION
TABLE 4. ASE EXTRACTION OF WHEATStudy No. 2: Recoveries of Spiked
Pesticides, Herbicides, and Fungicides
Spike Recovery(µg/L) (%)
OrganophosphorusPesticidesAzinphos-methyl 56 94.2Chlorpyrifos 20 60.1Chlorpyrifos-methyl 8 115.7Demeton-S 38 96.7Diazinon 26 96.9Dichlovos 18 60.5Dimethoate 58 87.8Disulfoton 22 87.9Disulfoton-sulfone 98 77.7Omethoate 74 85.4Parathion 84 101.2Parathion-methyl 40 115.7Phorate 18 92.8Phorate-sulfone 32 105.7
Chlorinated PesticidesEndosulfan-alpha 56 94.1Endosulfan-beta 68 93.3Endosulfan-sulfate 20 77.0Methoxychlor-o,p 48 89.9Methoxychlor-p,p’ 50 114.9
Carbamate PesticidesCarbaryl 92 54.1Carbofuran 22 96.6
HerbicidesAtrazine 14 92.8Diclofop-methyl 36 81.8Linuron 102 83.6Triallate 68 87.8Trifluralin 44 99.6
FungicidesImazalil 40 108.8Thiabendazole 44 158.8
TABLE 3. PESTICIDES IN WHEATStudy No. 2: ASE Extraction Conditions
Sample Size 3.0 gStatic Time 5 minSolvent AcetonitrileTotal Time 12 minPressure 2000 psiSolvent Volume 14 mLTemperature 100 °CSample Analysis GC-MSHeatup Time 5 min
Study No. 2: Pesticides, Herbicides,and Fungicides
Table 3 summarizes the ASE conditionsused in the second study and Table 4shows the recovery data. In this study,samples were spiked at 2 times the limit ofquantification, as determined by theresearcher using GC-mass spectrometry inthe single-ion monitoring mode. In thiscase, extracts were cleaned up prior toanalysis. The data indicates that ASE givesexcellent recoveries for a wide variety ofpesticide classes.
45
EXTRACTION OF ORGANOCHLORINEPESTICIDES FROM FRUITS & VEGETABLES
Diatomaceous earth is typically addedto fruit and vegetable samples because oftheir high water content. In the examplesshown, all the pesticides were spiked at100 µg/kg. The same extraction conditionswere used for both banana and potato, andall extracts were analyzed by GC. Goodrecoveries and RSDs were obtained for allthe organochlorine pesticides, as shown inTables 5 and 6.
TABLE 5 . ASE EXTRACTION OF ORGANOCHLORINEPESTICIDES FROM BANANASa,b
Compound Avg. Rec. Std. RSD(n = 3) Dev. (%)
alpha-BHC 100.3 2.3 2.3beta-BHC 102.2 2.3 2.3gamma-BHC 98.9 3.2 3.2Heptachlor 89.2 7.6 8.5Aldrin 89.4 2.2 2.5Heptachlor Epoxide 93.5 2.1 2.2Dieldrin 93.7 1.6 1.74,4'-DDE 92.1 1.8 1.92,4'-DDD 95.4 2.5 2.6Endrin 94.4 2.7 3.04,4'-DDD 88.0 2.7 3.04,4'-DDT 89.6 5.8 6.4a100 µg/kg per compoundbConditions: 10-g samples mixed with 6-g diatomaceous earth, 100 °C,10 MPa (1500 psi); 5-min heatup, 5-min static, 60% flush; 60-s purge,hexane/10% acetone.
ACCELERATED SOLVENT EXTRACTION
Compound Avg. Rec. Std. RSD(n = 3) Dev. (%)
alpha-BHC 96.3 6.3 6.6beta-BHC 108.6 2.3 2.1gamma-BHC 97.4 6.6 6.8Heptachlor 93.9 3.5 3.7Aldrin 95.9 3.3 3.4Heptachlor Epoxide 95.2 2.4 2.6Dieldrin 97.1 0.55 0.574,4'-DDE 95.4 0.67 0.702,4'-DDD 95.7 0.85 0.89Endrin 97.8 1.8 1.94,4'-DDD 93.7 1.8 1.94,4'-DDT 93.0 4.5 4.8a100 µg/kg per compoundbConditions: 10-g samples mixed with 6-g diatomaceous earth, 100 °C,10 MPa (1500 psi); 5-min heatup, 5-min static, 60% flush; 60-s purge,hexane/10% acetone.
TABLE 6. ASE EXTRACTION OF ORGANOCHLORINEPESTICIDES FROM POTATOESa,b
EXTRACTION OF ORGANOPHOSPHORUSPESTICIDES FROM BABY FOOD
Sample of 30 g of baby food carrots andapples were weighed out. For this study,7.5 mL of a pesticide mixture at 0.2 mg/mLwas added to the baby food for a finalconcentration of 50 mg/kg on a samplemass basis. The samples were mixed withenough Hydromatrix™ to make them easyto work with and load into the extractioncells, usually around 1:1 (w/w). Table 7shows the results.
These results confirm that pesticideresidues can be easily extracted from large-volume food samples using the ASE 300.
46
ACCELERATED SOLVENT EXTRACTION
Traditional extraction methods would takefrom one to several hours for each sampleand several hundred milliliters of solventwould be used for each sample. With theASE 300, these samples can be extracted in
about 15 min each, with about 160 mL ofsolvent for each sample. In addition, theASE 300 can extract up to 12 samples se-quentially without user intervention.
50 µg/kg per compound. Conditions: 30-g samples mixed with 30-g Hydromatrix, 100 °C, 10 MPa (1500 psi); 5-min
heatup, 5-min static, 2 cycles, 60% flush; 180-s purge, methylene chloride/acetone (1:1, v/v).
TABLE 7. PERCENT RECOVERY OF ORGANOPHOSPHORUS PESTICIDES FROMSPIKED APPLE OR CARROT PUREE
Carrot Puree Apple PureeAvg. Rec. % RSD Avg. Rec. % RSD
(n = 12) (n = 12)
Dichlorvos/Naled 82 8 87 12Mevinphos 94 8 100 12TEPP 90 14 121 16Demeton-O 97 11 65 19Ethoprophos (Ethoprop) 89 8 95 12Sulfotep 85 6 95 10Phorate 88 7 86 9Demeton-S 93 18 59 18Dimethoate 125 9 128 15Diazinon 86 7 93 10Disulfoton 97 11 63 18Parathion-methyl 87 8 95 10Fenchlorphos 84 7 91 10Malathion 86 8 94 15Fenthion 88 7 86 8Chlorpyrifos 83 8 91 10Parathion-ethyl 84 12 99 11Trichloronat 86 7 89 10Tetrachlorvinphos 85 7 91 9Prothiofos 84 7 85 11Merphos 87 9 82 10Fensulfothion 91 8 98 11Sulprofos 86 8 80 10EPN 89 7 97 11Azinphos-methyl 95 9 98 11Coumaphos 90 7 98 8
47
ACCELERATED SOLVENT EXTRACTION
SELECTIVE EXTRACTION OF PCBSFROM FISH TISSUE
The analysis of extracts containing PCBcontaminants from fish tissue and fishhomogenates can be hindered by the pres-ence of coextracted fatty materials thatinterfere with the chromatographic analy-sis. Cleanup procedures, including size-exclusion chromatography (SEC), columnchromatography, and acid treatment, areusually required to remove the coextractedlipids from such samples prior to analysis.These procedures are time consuming andincrease the potential for analyte losses. Analternative selective extraction procedureusing accelerated solvent extraction hastherefore been developed.
Selective ASE extractions can be per-formed with the proper choice of solventand sorbent in the extraction cell. An im-portant benefit of using a sorbent, such asalumina in this case, is that the extracts, ascollected, may not require further cleanupprior to analysis by gas chromatography.
The fish tissue sample was obtainedfrom the National Research Council ofCanada (NRC-CNRC). It is characterizedas a ground whole Carp reference materialfor organochlorine compounds (CARP-1).
A Comparison of “Nonselective” and“Selective” ASE Extractions
A chromatogram of a "nonselective" hex-ane ASE extract of the fish tissue is showncompared to that of a "selective" ASE extractof the same sample. The use of alumina in
the outlet end of the extraction cell adsorbslipids and other coextractable materials, thusyielding a much cleaner extract and greatlysimplifying the quantification of PCBs.
In general, the selective extraction usingASE gives acceptable results (see Table 7),and eliminates the need for additionalcleanup, such as sulfuric acid treatment orsize-exclusion chromatography.
Using this method, both sample prepa-ration time and the potential for analytelosses are decreased significantly.
0
1000
mV
0 10 20 30Minutes
40 50
0
1000
mV
12018
ASE Extraction of Fish Tissue
A Nonselective Hexane Extract
B Selective Extract with Alumina in Cell
Columns: DB-608 & DB-1701 (J&W)30-m × 0.53-mm i.d.
Carrier: Helium, 30 cm/sInjector: Splitless at 220 ˚C, 5 µLDetector: Electron capture at 320 ˚COven: 60–200 ˚C at 28 ˚C/min after
1-min hold, then to 265 ˚C at 10 ˚C/min and 20.5-min holdAnalyses performed byMountain States Analytical, Inc.
0 10 20 30Minutes
40 50
48
ACCELERATED SOLVENT EXTRACTION
Congener Cert.a Value Extract 1 Extract 2 Extract 3 Average Std. Dev. RSD (%)
52 124 ± 32 100 107 99 102 4.4 4.3101/90 124 ± 37 101 103 100 101 1.5 1.5
105 54 ± 24 124 128 125 126b 2.1 1.7118 132 ± 60 107 109 107 108 1.2 1.1
138/163/164 102 ± 23 48 48 48 48b 0.0 N/A153 83 ± 39 48 48 48 48 0.0 N/A
170/190 22 ± 8 30 31 31 31 0.58 1.9180 46 ± 14 65 62 64 64b 1.5 2.4
187/182 36 ± 16 30 30 30 30 0.0 N/A
a 95% confidence limits are given. b Values fall outside the 95% confidence limitsConditions: 3-g samples mixed with 15-g sodium sulfate, dried, then combined with 5-g alumina; hexane; 100 °C,10 MPa (1500 psi), 5-min heatup, 5-min static, 60% flush, 90-s purge, 2 static cycles, 17-min per sample total time.
TABLE 8. RECOVERY OF PCBS FROM FISH TISSUE USING SELECTIVE ASE(concentration expressed as µg/kg)
PCBS IN LARGE-VOLUME FISH TISSUESAMPLES
The fish tissue used in this study wascod fillet obtained from a local source. Thesample had a fat content of 0.25% and amoisture level of 81%. The samples werepremixed with 20 g of pelleted diatoma-
ceous earth (Hydromatrix) prior to cellloading. Extraction results are shown inTable 9. Average recovery for the nine PCBcongeners was 96.9% with an averagepercent RSD of 6.1 (n = 5).
TABLE 9. RECOVERY OF SPIKED PCB CONGENERS FROM30-g FISH TISSUE SAMPLES
Congener BZ # Spike (µg) % Recovery % RSD2-Chlorobiphenyl 1 2.5 99.8 3.02,3-Dichlorobiphenyl 5 2.5 103.8 8.82,4,5-Trichlorobiphenyl 29 2.5 107.1 3.12,2’,4,6-Tetrachlorobiphenyl 50 5.0 98.4 2.42,2’,3,4,5’-Pentachlorobiphenyl 87 5.0 92.3 7.92,2’,4,4’,5,6’-Hexachlorobiphenyl 154 5.0 89.0 5.92,2’,3,4’,5,6,6’-Heptachlorobiphenyl 186 7.5 91.1 8.52,2’,3,3’,4,5’,6,6’-Octachlorobiphenyl 200 7.5 96.0 6.5Decachlorobiphenyl 209 12.5 94.2 8.7
Conditions: 125 °C, 1500 psi, 5-min heatup, 3-min static, 60% flush, 120-s purge, 3 static cycles, methylene chloride solvent.
100-mL cells containing 10 g alumina.
49
ACCELERATED SOLVENT EXTRACTION
TABLE 10. PCB RECOVERYFROM OYSTER TISSUEa
PCB Congener Avg. Rec., n = 6 RSD(as % of Soxhlet) (%)
PCB 28 90.0 7.8PCB 52 86.9 4.0PCB 101 83.3 1.5PCB 153 84.5 3.5PCB 138 76.9 3.0PCB 180 87.0 4.3
a Analyte concentration range: 50–150 µg/kg per component.Conditions: 5–10 g, 100 °C, 14 MPa (2000 psi); 5-min heatup,5-min static, 60% flush; 60-s purge, hexane/acetone (1:1), (v/v)
EXTRACTION OF PCBS FROMOYSTER TISSUE
Table 10 shows the ASE extraction ofPCBs from oyster tissue. The table showsthe average recoveries and percent RSDsfor PCB congener content.
Sample Preparation and AnalysisOyster tissue samples were obtained
from the National Oceanographic andAtmospheric Administration (NOAA)Laboratory (Seattle, Washington, USA).Samples were mixed in equal proportionswith Hydromatrix to bind water.
Analysis and QuantificationASE extracts were passed first through
a silica gel loaded with silver nitrate/sulfuric acid; then through alumina col-umns; and concentrated to 1 mL for GCanalysis with ECD using a 30-m × 0.25-mmi.d. Rtx-5 (or equivalent) column.
50
ACCELERATED SOLVENT EXTRACTION
TABLE 14. EXTRACTION OF FATFROM SWEET CEREAL
Method Solvent % Fat Std. RSD(wt. %) Dev. (%)
Soxhlet Methanol/ 10.0-12.0 N/A N/ACHCl3 (2:1)
ASEa Hexane/ 11.6 0.09 0.73IPA (3:2)
a Conditions: same as Table 5
DETERMINATION OF FAT IN VARIOUSFOOD MATRICES
The fat content of foods has been ofgrowing concern worldwide. In the U.S.,the Nutrition Labeling and Education Actrequires the labeling of total saturated andunsaturated fats contained in foods. Foodmanufacturers also require a method forroutine determination of fat content forquality-control purposes.
In currently used methods, such asSoxhlet and automated Soxhlet, the fatcontent is determined gravimetrically afterextraction with organic solvents such aschloroform or petroleum ether. Thesemethods require large volumes of solventsand time periods ranging from 2 to 16 h.Faster techniques requiring less solvent aretherefore needed.
Comparison of ASE to Soxhlet MethodsASE was applied to the determination of
fat content in various solid or semisolidfoods. The fat content was determined bycollecting the extracts in preweighed vials,evaporating the solvent with a nitrogenstream, and reweighing the vials.
Samples were provided by a number ofdifferent food companies. In all cases, theSoxhlet results shown were determined bythe food company supplying the sample.
Table 11 shows the results of extractingvarious snack foods by ASE and determin-ing their fat content. Recoveries were equi-valent to Soxhlet. Precision, expressed asrelative standard deviation (RSD), is good.
Tables 12–16 compare results for otherfoods. In most cases, there is close agreementbetween Soxhlet and ASE using the sameextraction solvent. Good agreement may
TABLE 11. EXTRACTION OF FATFROM SNACK FOODS
Sample Avg. % Fat Std. RSD(n = 5) (wt. %) Dev. (%)
Potato Chips 34.0 0.11 0.33Corn Chips 32.8 0.08 0.25Cheese Snacks 33.3 0.17 0.51Tortilla Chips 21.5 0.07 0.34Snack Chip 19.2 0.10 0.53
*Conditions: 3-g samples, 125 °C, 6.7 MPa (1000 psi), 6-min heatup,25-min static, 60% flush, 60-s purge, chloroform, 3 static cycles.
Method Solvent Avg. % Fat Std. RSD(wt. %) Dev. (%)
Soxhlet Methanol/ 20.0–22.0 N/A N/ACHCl3 (2:1)
ASE, n=3a Hexane/ 20.8 0.18 0.85IPA (3:2)
a Conditions: 3-g samples, 125 °C, 6.7 MPa (1000 psi), 6-min heatup,25-min static, 60% flush, 60-s purge, 1 static cycle.
TABLE 12. EXTRACTION OF FATFROM COOKIES
Sample Method Avg. % Fat Std. RSD(wt. %) Dev. (%)
Cracker 1 Soxhleta 15.4 N/A N/ACracker 1 ASEb, n=3 14.6 0.09 0.65Cracker 2 Soxhleta 28-30 N/A N/ACracker 2 ASEb, n=3 28.1 0.20 0.70
TABLE 13. EXTRACTION OF FATFROM SNACK CRACKERS
a After acid hydrolysisb Conditions: 5-g samples, 125 °C, 6.7 MPa (1000 psi), 6-min heatup,25-min static, 60% flush, 60-s purge, 1 static cycle, hexane/isopropanol (3:2).
51
ACCELERATED SOLVENT EXTRACTION
also be obtained with a different solvent.For fat extraction of sweet cereal (Table 14),methanol/chloroform mixture was usedfor the Soxhlet extraction. However, thecompany providing the sample wanted toremove the methanol from the productionline and eliminate the use of chlorinatedsolvents (e.g., chloroform). Equivalent datawas generated for the ASE extraction bychanging the solvent to hexane/isopro-panol (3:2).
Comparison of ASE to the MojonnierMethod
The traditional Mojonnier method for thedetermination of fat in dairy products in-volves a base pretreatment (usually ammo-nium hydroxide) followed by extraction withdiethyl ether/ethanol, and petroleum ether/ethanol. The purpose of the base petreatmentis to dissolve casein and release interstitialfat. The method is both labor and time inten-sive, and requires multiple extraction steps.Comparison of ASE with Mojonnier forextraction of a variety of high-fat foods andcheeses indicates that a base pretreatment isnot required for ASE. The results obtainedwith a 30-min ASE extraction show goodagreement with the Mojonnier method.
Sample Method Solvent Avg. % Fat Std. RSD(wt. %) Dev. (%)
SRMa Soxhlet Petroleum 8.80 0.50 5.7ether
SRM ASEb Petroleum 9.12 0.15 1.6ether
Brand X Soxhlet Petroleum 10.3 N/A N/Aether
Brand X ASEb Hexane/ 10.4 N/A N/AIPA (3:2)
TABLE 15. EXTRACTION OF FATFROM DOG BISCUITS
a SRM = Standard Reference Materialb Conditions: 7-g samples, 125 ˚C, 6.7 MPa (1000 psi), 6-min heatup,25-min static, 60% flush, 60-s purge, 1 static cycle.
ASE vs Mojonnier Fat Extraction (Gravimetric)High-Fat Content Foods Common Cheeses
0
20
40
60
80
Salad Mayonnaise Cream Processed PeanutDressing Cheese Cheese Butter
Wei
ght %
Fat
ASE
Mojonnier
0
10
20
30
Wei
ght %
Fat
Mild Cheddar Monterey MozzarellaCheese Jack Cheese String Cheese
ASE/Acetone
Mojonnier
ASE/Hex:IPA
TABLE 16. EXTRACTION OF FATFROM LOW-FAT SNACK CRACKERS
Method % Fat Std. RSD(wt. %) Dev. (%)
Soxhlet 1.40 N/A N/AASEa 1.43 0.03 2.1
a Conditions: 5-g samples, 125 °C, 6.7 MPa (1000 psi), 6-min heatup,25-min static, 60% flush, 60-s purge, 1 static cycle, hexane/isopropanol(3:2).
52
ACCELERATED SOLVENT EXTRACTION
Extraction of Fat from ChocolateThe extraction of fat from chocolate by
ASE is compared with an acid hydrolysis/ether extraction Mojonnier method (AOAC922.06). The ASE extraction is completed inonly 18 min and uses 20 mL of solvent ascompared with the Mojonnier method thattakes over 2 h and uses more than 110 mLof solvent. See Table 17.
TABLE 17. EXTRACTION OF FAT FROMCHOCOLATE
Sample (n = 3) Avg. % Fat SD % RSDBaking Chocolate ASE 52.80 0.35 0.67Mojonnier 51.69 0.26 0.50Milk Chocolate ASE 31.80 0.32 1.02Mojonnier 32.34 0.33 1.02Cocoa Powder ASE 11.82 0.12 1.01Mojonnier 11.52 0.15 1.33
ASE Conditions: 1-g samples mixed with Hydromatrix, 125 °C, 10 MPa
(1500 psi), 6-min heatup, 3-min static, 60% flush, 60-s purge, 3 static
cycles, Petroleum ether 100%.
Mojonnier conditions: Followed AOAC Method 922.06.
Determination of Fat in Dried MilkProducts
The samples range from very low-fatproducts such as skim milk powder to veryhigh-fat products such as cream powder.The extraction of fat from these matrices israpid and the results are equivalent to thereferenced traditional methods. Samples of2 grams are placed directly into 11-mLextraction cells. See Tables 18 and 19. Datasupplied courtesy of New Zealand DairyResearch Institute.
Samples Extraction Solvent Ratio(hexane:dichloro-
methane:methanol)Whole Milk Powder 5:2:1Cream Powder 5:2:1Skim Milk Powder 3:2:1Whey Protein Concentrate 2:3:3Whey Protein Isolate 2:3:3Sodium Caseinate 2:3:3
TABLE 18. GRAVIMETRIC COMPARISON OF ASEAND ROESE-GOTTLIEB METHODS
Sample % Fat ASE % Fat RGCream Powder 54.88 54.96Whole Milk Powder 29.41 29.45Skim Milk Powder 0.96 0.95
Conditions: 80 °C, 1500 psi, 5-min heatup, 1-min static, 100% flush, 40-s
purge, 3 static cycles, solvent as noted above.
TABLE 19. GRAVIMETRIC COMPARISON OF ASEWITH SBR AND SOXHLET METHODS
Sample % Fat ASE % Fat SBR % Fat SoxhletLactic Acid WPC Powder 5.47 4.95 5.50Acid WPC Powder 5.71 5.66 6.38Cheese WPC Powder 6.93 6.75 7.32Whey Protein Isolate 0.45 0.58 0.50Sodium Caseinate 0.66 0.65 0.55
Conditions: 80 °C, 1500 psi, 5-min heatup, 1-min static, 100% flush, 40-s
purge, 3 static cycles, solvent as noted above. SBR stands for Schmidt-
Bondzynski-Ratzlaff Method.
53
ACCELERATED SOLVENT EXTRACTION
Extraction of Fat from Liquid DairyProducts
The current methods for determiningfat in dairy products, though acceptable,have several drawbacks. Many dairy-based products require a pretreatmentprior to extraction. The standard fat extrac-tion method, including the pretreatment, ismanual and thus very time consuming.Large amounts of solvent are required toremove the fat from each sample matrix.
ASE is an automated extraction tech-nique that uses the same solvents as cur-rent extraction techniques but in signifi-cantly smaller amounts and in a fraction ofthe time. ASE extraction of the followingliquid milk samples takes only 10 min anduses less than 30 mL of solvent. See Tables20 and 21. Data supplied courtesy of NewZealand Dairy Research Institute.
TABLE 20. SAMPLE PREPARATION FOR THE EXTRACTION OF FAT FROM LIQUIDMILK PRODUCTS
SAMPLE CELL SIZE, HYDROMATRIX EXTRACTION SOLVENT COMBINATION(HM) AMOUNT, SAMPLE AMOUNT
Cream 11-mL cell, 0.9 g HM, 1 g sample. Petroleum ether:acetone:isopropanol (3:2:1)Whole Milk 11-mL cell, 2 g HM, 1 g sample. Petroleum ether:isopropanol (2:1)Homogenized/UHT Milk 11-mL cell, 2 g HM, 1 g sample. Petroleum ether:isopropanol (3:2)Skim Milk 33-mL cell, 5.5 g HM, 3-5 g sample. Petroleum ether:isopropanol (3:2)
TABLE 21. MILK AND CREAM % FATRECOVERY: ASE VS ROESE-GOTTLIEB METHOD
Sample ASE Mean ± SD (n) RG MethodCream 40.62 ± 0.06 (3) 40.58Whole Milk 4.42 ± 0.02 (4) 4.50Homogenized Milk 3.39 ± 0.03 (6) 3.39Skim Milk 0.053 ± 0.010 (7) 0.053
ASE Conditions: 120 °C, 1500 psi, 6-min heatup, 1-min static, 100%
flush, 60-s purge, 3 static cycles.
54
ACCELERATED SOLVENT EXTRACTION
EXTRACTION OF OILS FROM OILSEEDSOils for foods and cooking are derived
from oilseeds such as canola, soybeans,corn, flax, cotton, etc. The accurate determi-nation of seed oil content is therefore essen-tial for optimizing oil output.
Existing methods for extraction of oilfrom oilseeds use large volumes of solvent(typically several hundred milliliters) andlong extraction times (8 to 16 h).
Comparison of ASE to the CurrentOfficial Method
Extraction of canola seeds, which con-tain approximately 45-weight percent oil,serves as an excellent example for compar-ing ASE to the AOCS (American Oil Chem-ist Society) Official Method AM 2-93,which is based on the FOSFA (Federationof Oil Seeds and Fat Association) OfficialMethod. Table 22 gives the specifics of thismethod. Conditions used for the ASEextraction are listed in Table 23.
Results showed close agreement be-tween ASE and the official method. ForASE, the weight percent of oil in the seedswas 44.9% with 0.31% RSD (n = 3) as com-pared to 45.2% with 0.24% RSD (n = 12) forthe AOCS method. The histograms to theright show the percent of the total oil ex-tracted as a function of time. ASE givescomparable results faster and with lesssolvent usage than the FOSFA procedure.
Peroxide value (PV) and free fatty acid(FFA) determinations show that no signifi-cant triglyceride degradation occurs dur-ing the ASE extraction. 0
20
40
60
80
100
(%) E
xtrac
ted
240 480 720Minutes
FOSFA
10258
Extraction of Oil from Oilseeds: Comparison of ASE and FOSFA
Solvent Extraction Methods
30 60 90
ASE 200
Minutes
TABLE 22. EXTRACTION OF OIL FROM OILSEEDSAOCS Method AM 2-93 Conditions
Sample Size: 4 g ground seedsOven: 130 °C, 2 hExtract: 4 h, drain solvent and coolRegrind: 7 minExtract: 2 h, drain solvent and coolSolvent: Petroleum etherTotal Vol. Solvent Used: 150–250 mLTotal Time: 10.5 h
TABLE 23. EXTRACTION OF OIL FROM OILSEEDSASE Extraction Conditions
System Pressure: 6.7 MPa (1000 psi)Oven Temperature: 105 °COven Heatup Time: 5 minStatic Time: 10 minFlush Volume: 100%Purge Time: 60 sSolvent: Petroleum etherStatic Cycles: 3
55
APPENDIX ONE:AOAC INTERNATIONAL
APPROVED
HPLC METHODS
56
Aflatoxins in Cottonseed Products 980.20 Silica, 25 cm × 4.6 mm, 5 µm
Aflatoxins M1 and M2 in Fluid Milk 986.16 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Antioxidants in Oils and Fats 983.15 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Aprinocid in Feeds 981.27 Silica, 25 cm × 4.6 mm, 5 µm
Bacitracin in Premix Feeds 982.44 C8 Reversed Phase, 15 cm × 4.6 mm, 5 µm
Benzoate, Caffeine, and Saccharin in Soda Beverages 979.08 C18 Reversed Phase, 30 cm × 4.6 mm, 5 µm
Benzoic Acid in Orange Juice 994.11 SupelcoGel™ TPR, 15 cm × 4.6 mm, 5 µm
Domoic Acid in Mussels 991.26 C18 Reversed Phase, 15 cm × 4.6 mm, 5 µm
Fenbendazole in Beef Liver 991.17 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Furazolidone in Feeds and Premixes 985.51 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Glucose, Fructose, Sucrose, and Maltose in 982.14 NH2-bonded Silica, 25 cm × 4.6 mm, 5 µmPresweetened Cereals
Glycerol in Wine and Grape Juice 991.46 SupelcoGel C-610H, 30 cm × 7.8 µm
Glycyrrhizic Acid or Acid Salts in Licorice Products 982.19 C18 Reversed Phase, 30 cm × 4.6 mm, 5 µm
Glyphosate, Technical and Formulations 983.10 Silica-SAX, 25 cm × 4.6 mm, 5 µm
Intermediates and Reaction By products in 982.28 Silica-SAX, 25 cm × 4.6 mm, 5 µmFD&C Yellow No. 5
Intermediates in FD&C Red No. 40 981.13 Silica-SAX, 25 cm × 4.6 mm, 5 µm
Intermediates in FD&C Yellow No. 8 977.23 Silica-SAX, 25 cm x 4.6 mm, 5 µm
Iodine in Pasteurized Liquid Milk and Skim Milk Powder 992.22 C18 Reversed Phase, 15 cm × 4.6 mm, 5 µm
N-Methylcarbamate Residues in Grapes and Potatoes 985.23 C8 Reversed Phase, 15 cm × 4.6 mm, 5 µm
N-Methylcarbamoyloximes and N-Methylcarbamates 991.06 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µmin Finished Drinking Water
Ochratoxin A in Corn and Barley 991.44 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Pesticides in Water 992.14 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
AOAC METHOD NUMBER COLUMN TYPE
* Dionex equipment can be used for all methods listed
AOAC INTERNATIONAL:APPROVED HPLC METHODS
57
AOAC INTERNATIONAL:APPROVED HPLC METHODS
AOAC METHOD NUMBER COLUMN TYPE
Phenolic Antioxidants in Oils, Fats, and Butter Oil 983.15 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Purity of Lactose 984.22 NH2-bonded Silica, 25 cm × 4.6 mm, 5 µm
Quinic, Malic, and Citric Acids in Cranberry Juice 986.13 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µmCocktail and Apple Juice
Saccharides (major) in Corn Syrups and Sugars 979.23 Cation exchange-Ca form resin, 30 × 7.8
Saccharides (minor) in Corn Syrups and Sugars 979.23 Cation exchange-Ca form resin, 30 × 7.8
Separation of Sugars in Honey 977.20 NH2-bonded Silica, 25 cm × 4.6 mm, 5 µm
Sugars in Licorice Extracts 984.17 NH2-bonded Silica, 25 cm × 4.6 mm, 5 µm
Sulfamethazine in Raw Bovine Milk 992.21 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Total Malic Acid in Apple Juice 993.05 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Trans-Vitamin K1 in Infant Formula 992.27 Silica, 25 cm × 4.6 mm, 5 µm
Triglycerides in Vegetable Oils 993.24 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Vanillin, Vanillic Acid, p-Hydroxybenzaldehyde, and 990.25 C8 Reversed Phase, 15 cm × 4.6 mm, 5 µmp-Hydroxybenzoic Acid in Vanilla Extract
Vitamin A in Milk and Milk-Based Infant Formula 992.04 C18 Reversed Phase, 25 cm × 4.6 mm, 5 µm
Vitamin D in Fortified Milk and Milk Powder 981.17 Silica, 25 cm × 4.6 mm, 5 µm
Vitamin D in Infant Formula 992.26 Silica, 15 cm × 4.6 mm, 5 µm
Vitamin D in Mixed Feeds, Premixes, and Pet Foods 982.29 Silica, 25 cm × 4.6 mm, 5 µm
Vitamin D in Multivitamin Preparations 980.26 Silica, 25 cm × 4.6 mm, 5 µm
Vitamin D in Vitamin A & D Concentrates 985.27 Silica, 25 cm × 4.6 mm, 5 µm
Vitamin D in Vitamin Preparations 979.24 Silica, 15 cm × 4.6 mm, 5 µm
Vitamin E Activity in Milk-Based Infant Formula 992.03 Silica, 25 cm × 4.6 mm, 5 µm
Zearalenone and α-Zearalenol in Corn 985.18 C18 Reversed Phase, 30 cm × 4 mm, 5 µm
59
APPENDIX TWO:RECOMMENDED
READING
60
RECOMMENDED READING
JOURNAL ARTICLES
Amines and Other BasesDraisci, R.; Cavalli, S.; Lucentini, L.; Stacchini,A. “Ion Exchange Separation and PulsedAmperometric Detection for Determinationof Biogenic Amines in Fish Products”Chromatographia 1993, 35 (9–12), 584–590.
Draisci, R.; Giannetti, L.; Boria, P.; Lucentini,L.; Palleschi, L.; Cavalli, S. “Improved IonChromatography—Integrated PulsedAmperometric Detection Method for theEvaluation of Biogenic Amines in Food ofVegetable or Animal Origin and inFermented Foods.” Chromatogr., A 1998, 798,109–116.
Hagar, A. F.; Madsen, L.; Wales, Jr., L.;Bradford, Jr., H. B. J. Assoc.“Reversed-PhaseLiquid Chromatographic Determination ofVitamin D in Milk” Off. Anal. Chem. 1994, 77,1047–1051.
Amino AcidsSingleton, J. A.; Grimm, D. T.; Sanders, T. H.“Interference of Amino Acids in PulsedAmperometric Detection of Peanut Sugars”Peanut Science 1996, 23, 61–65.
CarbohydratesAlonso, S.; Setser, “Functional Replacementsfor Sugars in Foods” C. Trends Food Sci.Tech. 1994, 5 (May), 139–146.
Akers, A. A.; Hoseney, R. C. “Water SolubleDextrins from α-Amylase-Treated Bread andTheir Relationship to Bread Firming” CerealChemistry 1994, 71, 224–226.
Corradini, C.; Canali, G.; Cogliandro, E.;Nicoletti, I. “Determination of Sugars andSugar Alcohols in Dietetic Sweeteners andFood by High-Performance Anion-ExchangeChromatography (HPAEC) Coupled withPulsed Amperometric Detection (PAD)”Proceedings of EURO FOOD CHEM VIII,Vienna, Austria, September 18–20, 1995, 2,307–310.
Corradini, C.; Canali, G.; Nicoletti, I. “Appli-cation of HPAEC to Carbohydrate Analysisin Food Products and Fruit Juices” Seminarsin Food Analysis 1997, 2, 99–111.
Corradini, C.; Cristalli, A.; Corradini, D.“HighPerformance Anion-Exchange Chroma-tography with Pulsed Amperometric Detectionof Nutritionally Significant Carbohydrates” J. Liq. Chromatogr. 1993, 16, 3471–3485.
Craig, S. A. S.; Holden, J. F.; Khaled, M. Y.“Determination of Polydextrose as DietaryFiber in Foods”J. AOAC Int. 2000, 83 (4),1006–1009.
Craig, S. A. S.; Holden, J. F.; Khaled, M. Y.“Determination of Polydextrose in Foods byIon Chromatography: Collaborative Study” J.AOAC Int. 2001, 84 (2), 472–478.
Day-Lewis, C. M. J.; Schäffler, K. J.“Analysis of Sugar in Final Molasses byIon Chromatography” Proc. S. A. SugarTechnol. Assoc., June 1992.
Déséveaux, S.; Daems, V.; Delvaux, F.;Derdelinckx, G. “Analysis of FermentableSugars and Dextrins in Beer by Anion-Exchange Chromatography withElectrochemical Detection” Seminars in FoodAnalysis 1997, 2, 113–117.
Eggleston, G.; Clark, M. A. “Applications ofHPAE-PAD in the Sugar Industry” Seminarsin Food Analysis 1997, 2, 119–127.
61
RECOMMENDED READING
Garleb, A. K.; Bourquin, L. D.; Fahey, Jr., G.C. “Neutral Monosaccharide Composition ofVarious Fibrous Substances: A Comparisonof Hydrolytic Procedures and Use of Anion-Exchange HPLC with PAD Detection ofMonosaccharides” J. Agric. Food Chem 1989,37, 1287–1293.
Giese, J. H. “Alternative Sweeteners andBulking Agents” Food Technol. 1993, (January),114–126.
Hotchkiss, Jr., A. T.; Hicks, K. B.“Analysis ofOligogalacturonic Acids with 50 or FewerResidues by High-Performance AnionExchange Chromatography and PulsedAmperometric Detection” Anal. Biochem.1990, 184, 200–206.
Hoebregs, H. “Fructans in Foods and FoodProducts, Ion Exchange ChromatographicMethod: Collaborative Study” PosterPresented at the 110th AOAC InternationalMeeting, Orlando, FL, Sept. 8–12, 1996.
Lamb, J. D.; Myers, G. S.; Edge, N. “IonChromatographic Analysis of Glucose,Fructose, and Sucrose Concentrations in Rawand Processed Vegetables” J. Chromatogr.Sci. 1993, 31, 353–357.
Madigan, D.; McMurrough, I.; Smyth, M. R.“Application of Gradient Ion Chromatographywith Pulsed Electrochemical Detection to theAnalysis of Carbohydrates in Brewing” J. Am. Soc. Brew. Chem. 1996, 54 (1), 45–49.
Quemencer, B.; Thibault, J. F.; Coussement,P.“Determination of Inulin and Oligofructosein Food Products, and Integration in theAOAC Method for Measurement of TotalDietary Fibre” Lebensm.-Wiss. Technol. 1994,27, 125–132
Schäffler, K.; Day-Lewis, C. M. J.; Clarke, M.;Jekot, J. “Determination of Sugars in Beet andCane Final Molasses by Ion Chromatography:Collaborative Study”J. of AOAC Int. 1997, 80(3), 603–610.
Stumm, I.; Baltes, W. Z. “Determination ofPolydextrose in Food by Means of IonChromatography and Pulsed AmperometricDetection” Unters. Forsch 1992, 195, 246.
Swallow, K. W.; Low, N. H. “Detection ofOrange Juice Adulteration with Beet MediumInvert Sugar Using Anion Exchange LiquidChromatography with Pulsed AmperometricDetection” J. Assoc. Off. Anal. Chem. 1991, 74,341–343.
Thayer, A. M. “Food Additives” Chem. Eng.News 1992, June 15, 26–43.
Thompson, J. C. “Ion ChromatographicAnalysis of Sugars in Foods and Molasses”Proceedings of Sugar Processing ResearchInstitute Workshop on Analysis of Sugars inFoods 1992, M. C. Clarke, Ed.
Tsang, W. S. C.; Cargel, G.-L. R.; Clarke, M. A.“Ion Chromatographic Analysis of Oligosac-charides in Beet Sugar” Zuckerind. 1991, 116,No. 12, 1058–1061.
White, D. R., Jr.; Widmer, W. W. “Applicationof High Performance Anion-ExchangeChromatography with Pulsed AmperometricDetection to Sugar Analysis of Citrus Juices”J. Agric. Food Chem. 1990, 38, 1918–1921.
Wong, K. S.; Jane, J. “Effect of PushingAgents on the Separation and Detection ofDebranched Amylopectin by High-Performance Anion-ExchangeChromatography with Pulsed Ampero-metric Detection” J. Liq. Chromatogr. 1995,18 (1), 63–80.
62
Inorganic Anions and CationsBaluyot, E.; Hartford, C. G. “Comparisonof Polyphosphate Analysis by IonChromatography and by Modified End-Group Titration” J. Chromatogr. 1996, 739,217–222.
Bettler, K. M.; Chin, H. B.“ImprovedDetermination of Chlorite and Chlorate inRinse Water from Carrots and Green Beansby Liquid Chromatography and Ampero-metric and Conductivity Detection” J. Assoc.Off. Anal. Chem. Int. 1995, 78 (3), 878–883.
Buckee, G. K. “Determination of Anions inBeer by Ion Chromatography” J. Inst. Brew.1995, 101 (6), 429–430
de Bruijn, J. M.; Heringa, R. “Determinationof Anions and Cations in Sugar FactorySamples by Ion Chromatography” Presentedat the 1992 Conference on Sugar ProcessingResearch, New Orleans, LA, September 1992.
Dolenc, J.; Gorenc, D. “Ion ChromatographicDetermination of Inorganic Anions in VinegarSamples” Die Nahrung 1994, 4, 434–438.
Gaucheron, F; Le Graet, Y; Piot, M; Boyaval,E. “Determination of Anions of Milk by IonChromatography” Lait 1996, 76, 433–443.
Madigan, D; McMurrough, I; Smyth, M. R.“Determination of Oxalate in Beer and BeerSediments Using Ion Chromatography”J. Am. Soc. Brew. Chem. 1994, 52 (3), 134–137.
Pereira, C. F. “Application of IonChromatography to the Determination ofInorganic Anions in Food-stuffs”J. Chromatogr. 1992, 624, 457–470.
Perez-Cerrada, M.; Casp, A.; Maquieira, A.“Chromatographic Determination of theAnion Content in Spanish Rectified Musts”Am. J. Enol. Vitic. 1993, 44 (3), 292–296.
Sekiguchi, Y.; Matsunaga, A.; Yamamoto, A.;Inoue, Y. “Analysis of Condensed Phosphatesin Food Products by Ion Chromatographywith an On-Line Hydroxide EluentGenerator” J. Chromatogr., A 2000, 881,639–644.
Wagner, H. L.; McGarrity, M. J. “The Use ofPulsed Amperometry Combined with Ion-Exclusion Chromatography for the Simultan-eous Analysis of Ascorbic Acid and Sulfite”J. Chromatogr. 1991, 546, 119–124.
Organic AcidsBarber, E. L.“The Analysis of Organic Acidsby Ion Chromatography in Beer and Wort”J. Am. Soc. Brew. Chem. 1990, 48, 44–46.
Boyles, S.“Method for the Analysis ofInorganic and Organic Acid Anions in AllPhases of Beer Production Using GradientIon Chromatography” J. Am. Soc. Brew.Chem. 1992, 50, 61–63.
Johnson, D. C.; Ngoviwatchai “PulsedAmperometric Detection of Sulfur-Containing Pesticides in Reversed-PhaseLiquid Chromatography” Anal. Chim. Acta1988, 215, 1–12.
Saccani, G.; Gheradi, S.; Trifiro, A.; SoresiBordini, C.; Calza, M.; Freddi, C. “Use of IonChromatography for the Measurement ofOrganic Acids in Fruit Juices” J. Chromatogr.1995, 706, 395–403.
Talmond, P.; Doulbeau, S.; Rochette, I.;Guyot, J. P.; Treche, S. “Anion-ExchangeHigh-Performance Liquid Chromatographywith Conductivity Detection for the Analysisof Phytic Acid in Food0” J. Chromatogr., A2000, 871, 7–12
RECOMMENDED READING
63
VitaminsAlbalá-Hurtado, S.; Novella-Rodrígues, S.;Veciana-Nogués, M. T.; Mariné-Font, Abel.“Determination of Vitamins A and E in InfantMilk Formulae by High-Performance LiquidChromatography” J. Chromatogr, A, 1997, 778,243–246.
Chen, B. H.; Peng, H. Y.; Chen, H. E. “Changesof Carotenoids, Color, and Vitamin A Contentsduring Processing of Carrot Juice” J. Agric.Food Chem. 1995, 43, 1912–1918.
Haroon, Y.; Shearer, M. J.; Rahim, S.; Gunn,W. G.; McEnery, G.; Barkhan, P. “The Contentof Phylloquinone (Vitamin K1) in HumanMilk, Cows’ Milk and Infant Formula FoodsDetermined by High-Performance LiquidChromatography” J. Nutr. 1985, 112, 1105–1117.
Other“Application of High Performance AnionExchange Chromatography with PulsedAmperometric Detection in Food andBeverage Analysis” Seminars in Food Analysis1997, 2, 3–4.
Epler, K. S.; Ziegler, R. G.; Craft, N. E.“Liquid Chromatographic Method for theDetermination of Carotenoids, Retinoids andTocopherols in Human Serum and in Food”J. Chromatogr 1993, 619, 37–48.
Henshall, A. “Liquid ChromatographicTechniques for Detecting EconomicAdulteration of Foods” Cereal Foods World1998, 43 (2), 98–103.
Henshall, A. “Use of Ion Chromatography inFood and Beverage Analysis” Cereal FoodsWorld 1997, 42 (5), 414–419.
Koswig, S.; Fuchs, G.; Hotsommer, H. J.“The Use of HPAE-PAD for the Analysis ofThickening Agents in Fruit Juice and FoodAnalysis” Seminars in Food Analysis 1997, 2,71–83.
LaCourse, W. R.; Dasenbrock, C. O.; Zook, C.M. “Fundamentals and Applications ofPulsed Electrochemical Detection in FoodAnalysis” Seminars in Food Analysis 1997, 2,5–41.
Low, N. H. “Food Authenticity Analysis byHigh Performance Anion ExchangeChromatography with Pulsed AmperometricDetection” Seminars in Food Analysis 1997, 2,55–70.
Obana, H.; Kikuchi, K.; Okihashi, M.; Hori, S.“Determination of OrganophosphorusPesticides in Foods Using an AcceleratedSolvent Extraction System” The Analyst 1997,122, 217–220.
Okihashi, M.; Obana, H.; Hori, S.“Determination of N-MethylcarbamatePesticides in Foods Using an AcceleratedSolvent Extraction with a Mini-ColumnCleanup” The Analyst 1998, 123, 711–714.
RECOMMENDED READING
64
Dionex Presentations, Application Notes,and Technical NotesA Comparison of Conductivity andPostcolumn Derivatization Methods for theDetermination of DBP Anions in DrinkingWater Using Ion Chromatography”; DionexPresentation 2338.
“Recent Advances in IC-PAD (HPAE-PAD)Detection for the Analysis of Carbohydratesin Grain Products” Dionex Presentation 2328.
“Single Injection Determination of Sugars,Organic Acids, and Alcohols in Beveragesand Food” Dionex Presentation 1826.
“Accelerated Extraction of Vitamins fromTablets and Food Samples” DionexPresentation 2349.
Accelerated Solvent Extraction (ASE) ofMycotoxin Contaminants in Food Matrices”Dionex Presentation 2354.
“Use of Accelerated Solvent Extraction (ASE)for Analysis of Trace Contaminants in Foods”Dionex Presentation 2228.
“Use of Accelerated Solvent Extraction (ASE)for Analysis of Trace Contaminants in Foods”Dionex Presentation 2337.
AN#21: Organic Acids in Wine
AN#25: Analysis of Inorganic Anions andOrganic Acids in Carbonated Beverages
AN#37: Determination of Iodide in DairyProducts
AN#46: Ion Chromatography: A VersatileTechnique for the Analysis of Beer
AN#54: Determination of Sulfite in Foodand Beverages with Pulsed AmperometricDetection
AN#67: Determination of Plant-DerivedNeutral Oligo- and Polysaccharides
AN#70: Choline and Acetylcholine
AN#71: Determination of PolyphosphatesUsing Ion Chromatography with SuppressedConductivity Detection
AN#82: Analysis of Fruit Juice Adulteratedwith Medium Invert Sugar from Beets
AN#83: Size-Exclusion Chromatography ofPolysaccharides with Pulsed AmperometricDetection
AN#87: Determination of Sugar Alcohols inConfections and Fruit Juices by High-Perfor-mance Anion Exchange Chromatographywith Pulsed Amperometric Detection
AN#92: Determination of Sugars inMolasses by High-Performance AnionExchange Chromatography with PulsedAmperometric Detection
AN#112: Determination of Nitrate andNitrite in Meats
AN#124: Determination of Choline in Milkand Infant Formula
AN#142 : Determination of TryptophanUsing AAA-Direct
AN#143: Determination of Organic Acids inFruit Juices
AN#147: Determination of Polydextrose inFoods by AOAC Method 2000.11
AN#149: Determination of Chlorite, Bromate,Bromide, and Chlorate in Drinking Water byIon Chromatography with an On-LineGenerated Postcolumn Reagent for Sub-µg/LBromate Analysis
AN#155: Determination of Trans-Galactooligosaccharides in Foods by AOACMethod 2001.02
RECOMMENDED READING
65
AN#314: Determination of Unbound Fat inVarious Food Matrices Using AcceleratedSolvent Extraction (ASE)
AN#316: Extraction of PCBs from Environ-mental Samples Using Accelerated SolventExtraction (ASE)
AN#321: Determination of Unbound Fat inVarious Food Matrices Using AcceleratedSolvent Extraction (ASE)
AN#322: Selective Extraction of PCBs fromFish Tissue Using Accelerated SolventExtraction (ASE)
AN#325: Extraction of Oils from Oilseeds byAccelerated Solvent Extraction (ASE)
AN#329: Determination of Total Fat in InfantFormula Using Accelerated SolventExtraction (ASE)
AN#340 : Determingtion of Fat in Dried MilkProducts Using Accelerated SolventExtraction (ASE)
AN#342: Determingtion of PCBs in Large-Volume Fish Tissue Samples UsingAccelerated Solvent Extraction (ASE)
AN#343: Determination of Pesticides inLarge-Volume Food Samples UsingAccelerated Solvent Extraction (ASE)
AN#344 : Extraction of Fat from ChocolateUsing Accelerated Solvent Extraction (ASE)
AN#345: Extraction of Fat from DairyProducts (Cheese, Butter, and Liquid Milks)Using Accelerated Solvent Extraction (ASE)
AN#409: Fast Determination of Acrylamidein Food Samples Using Accelerated SolventExtraction (ASE) Followed by IonChromatography with UV or MS Detection
TN#20: Analysis of Carbohydrates by High-Performance Anion Exchange Chromatogra-phy with Pulsed Amperometric Detection(HPAE-PAD)
TN#21: Optimal Settings for PulsedAmerometric Detection of CarbohydratesUsing Dionex Pulsed Electrochemical andAmperometric Detectors
TN#50: Determination of the Amino AcidContent of Peptides by AAA-Direct
TN#55 : Screening of Sample Matrices andIndividual Matrix Ingredients for Suitabilityin AAA-Direct
RECOMMENDED READING
66
RECOMMENDED READING
The following series of journal articles is published in Seminars in Food Analysis;Hurst, W. J., Ed.; Chapman & Hall, 1997; Vol. 2, No. 1/2.
Campbell, J. M.; Flickinger, E. A.;Fahey, G., Jr. “A Comparative Study ofDietary Fiber Methodologies Using PulsedElectrochemical Detection of MonosaccharideConstituents” Dept. of Animal Sciences,University of Illinois, Urbana, IL, USA.
Coraddini, C.; Canali, G.; and Nicoletti, I.“Application of HPAEC-PAD to CarbohydrateAnalysis in Food Products and Fruit Juices”C.N.R. Istituto di Cromatografia del C.N.R.,Rome, Italy.
Deseveaux, S.; Daems, V.; Delvaux, F.;Derdelincx, G. “Analysis of FermentableSugars and Dextrins in Beer by HPAEC-PAD”University of Louvain, Centre for Maltingand Brewing Science, Leuven, Belgium.
Durgnat, J. M.; Martinez, C. “Determinationof Fructooligosaccharides in Raw Materialsand Finished Products by HPAE-PAD”Nestec Ltd., Nestlé Research Centre,Lausanne, Switzerland.
Eggleston, G. “Applications of HPAE-PAD inthe Sugar Industry” U.S. Dept. of Agriculture,Agricultural Research Service, SRRC, NewOrleans, LA; and Clarke, M., Sugar ProcessingResearch Institute, New Orleans, LA.
Koswig, S.; Fuchs, G.; Hotsommer, H. J.;Graefe, U.“The Use of HPAE-PAD for theAnalysis of Thickening Agents in Fruit Juiceand Food Analysis” Gesellschaft fürLebensmittel-Forschung mbH, Berlin,Germany.
Lacourse, W. R.; Dasenbrock, C. O.; Zook,C. M. “Fundamentals and Applications ofPulsed Electrochemical Detection in FoodAnalysis” Dept. of Chemistry and Biochem-istry, University of Maryland, BaltimoreCounty, MD, USA.
Low, N. H. “Food Authenticity Analysis byHigh-Performance Anion-ExchangeChromatography with Pulsed AmperometricDetection” Dept. of Applied Microbiologyand Food Science, University ofSaskatchewan, Saskatchewan, Canada.
67
INDEX
69
INDEX
AAccelerated Solvent Extraction 39–54
vs Mojonnier Method 51vs Soxhlet 40–41, 50–51
Additives 17Adulteration of:
Coffee 25Natural Fruit Juices 26
Amines and Other Organic Bases 15–18Biogenic 17Choline 16Vitamins 18
Amylopectins 30Anions and Cations, Inorganic 5–10Anions and Organic Acids in an
Irish Stout 14AOAC International: Officially
Approved HPLC Methods 55–57AOAC Method 973.28 21AOCS (American Oil Chemist
Society) 54Apricots (Dried) and Sulfite 8Approved IC Methods 2Artificial Sweetener from Japan 29ASE (Accelerated Solvent Extraction):
ASE 100, 200, and 300 42Schematic of Operation 41Technology and Features 41–42
BBaby Food 45–46Baked Goods and Bromate 7Bananas and Pesticides 45Beer, Wine, or Cider Production 27Beet Sugar 26Beverages and Sweeteners 26
Bread and Bromate 7Bromate in:
Baked Goods 7Drinking Water 7
Bulking Agents 28
CCandy (Hard) 22Carbohydrates 19–31Cations in:
Mineral Water and DrinkingWater 7
Soft Drinks and Wine 7Cereal, Extraction of Fat 50–51Cheese:
Extraction of Fat 51Phosphates in Cheese Products 10
Chewing Gum 22Chocolate 24, 52Coffee Adulteration 25Column Technologies 35Cookies, Extraction of Fat 50Crackers, Fat Extraction 51
DDairy Products:
Iodide in 9Fat, Extraction of 52–53
Detector Technologies 36Dionex Application and Technical
Notes 64Dionex LC Technologies 33–37Dog Biscuits, Fat Extraction 51Drinking Water 6–7Dyes 14
70
EEstablishing Geographic Origin 26EPA, U.S. 3Extraction (ASE) of:
Fat from:Cheese 51Chocolate 52Cookies 50Dog Biscuits 51Dried Milk Products 52High-Fat Content Food
(Salad Dressing, Mayonnaise,Cheeses, Peanut Butter) 51
Liquid Dairy Products 53Low-Fat Snack Crackers 51Snack Foods 50Sweet Cereal 50
Fungicides from Grain 44Herbicides from Grain 44Oils From Oilseeds 54Organochlorine Pesticides from
Fruits and Vegetables 45PCBs from:
Fish Tissue 47–48Oyster Tissue 49
Pesticides from:Baby Food 45–46Grain 41–42
FFat, Extracted from:
Cheese 51Chocolate 52Cookies 50Dog Biscuits 51Dried Milk Products 52High-Fat Content Food
(Salad Dressing, Mayonnaise,Cheeses, Peanut Butter) 51
Liquid Dairy Products 53
Low-Fat Snack Crackers 51Snack Foods 50Sweet Cereal 50
Fat Substitutes 28Fermentable Sugars 27Fish:
Extraction of PCBs 47–48Seafood Spoilage 17
Flavor Constituents and Additives 17Flavored Potato Chip Extract 24FOSFA (Federation of Oilseeds and
Fat Association) 54Fruit and Fruit Juices:
Oligogalacturonic Acidsfrom Citrus Pectin 31
Organic Acids in:Cranberry Juice 12Orange Juice 13Grape Juice 13Apple Juice 13
Sugars in Orange Juice 31Sulfite in Dried Apricot 8
Fruits and Vegetables:Extraction of Organochlorine
Pesticides 45Triazine Herbicides in 18
GGlucose, Fructose, Maltose, and
Maltotriose 23–24Glucose Syrup 21Grape Juice, Organic Acids in 13
HHam, Nitrates/Nitrites in 9Herbicides in:
Fruits and Vegetables 18Grain 44
High-Fat Foods 24HPAE-PAD, description of 20
INDEX
71
IIC Methods for Food 2–4ICUMSA 23Inorganic Anions and Cations 5–10Inorganic Cations, Choline, and
Acetylcholine 16Inulins 28Iodide in Whole Milk 9Irish Stout 14ISO/DIS 11292, for coffee
adulteration 25
JJournal Articles 60–63Juices. See Fruit and Fruit Juices.
KKestoses 29
LLow-Fat Snack Crackers,
Extraction of Fat 51
MMaltodextrins 29Mayonnaise, Extraction of Fat 51Methylamines 16Milk and Iodide 9Milk Products, Fat Extraction 52–53Molasses:
Beet Sugar, Geographic Origin 26Sugars in 23
Monjonnier Method, vs ASE 51–52Municipal Drinking Water 6
NNatural Fruit Juices 26Nitrite/Nitrate in Ham 9Nonselective vs Selective ASE 47Nutritional Labeling Requirements 21Nutritive Sweeteners 23
OOfficially Approved IC Methods 2–3Oils From Oilseeds, Extraction of 54Oligo- and Polysaccharides 21Oligogalacturonic Acid:
Fingerprinting 31From Citrus Pectin 31
Oligosaccharide Profiling 26Oligosaccharides 21, 26Orange Juice. See Fruit and Fruit Juices.Organic Acids in:
Irish Stout 14Fruit Juice 12–13
Organochlorine Pesticides in Fruitsand Vegetables 45
Organophosphorus and Baby Food45–46
Oyster Tissue, Extraction of PCBs 49
PPCBs, Extraction from:
Fish Tissue 47–48Oyster Tissue 49
Peanut Butter, Extraction of Fat 51Pectin 31Pesticides, Extraction from:
Fruits and Vegetables 45Grain 43–44
Polyphosphates 10Polysaccharides 21Potato Chips:
Extraction of Fat 50Flavor Additives 22
Potatoes and Pesticides 45Pulsed Amperometric Detection 36Pump Technology 37Purity. See Adulteration.
RRecommended Reading 25, 59–65
INDEX
72
SSalad Dressing, Extraction of Fat 51Seafood Spoilage, Amines 17Selective vs Nonselective ASE 47Snack Foods, Extraction of Fat
from 50–51Soft Drinks 7Stout, Organic Acids in 14Sucralose 28Sucrose, Maltose, Lactose, Dextrose
and Fructose 23Sugar Alcohols 21–22Sugar Alcohols in Dietetic Hard
Candy and Chewing Gum 22Sugars in:
Beer 27Foods 24High-Fat Foods 24Molasses 23Orange Juice 31
Sulfite in Dried Apricot 8Suppressed Conductivity Detection 36Sweet Cereal, Extraction of Fat 50Sweeteners 23–24, 26, 28–29Syrup, Glucose 21
TTomato Ketchup 24Transition Metals 8Triazine Herbicides in Raw Fruits
and Vegetables 18
UUnited States Environmental
Protection Agency (U.S. EPA) 3
VVitamins, Water-Soluble 18
WWater:
Drinking 6–7Mineral 7
Wheat and Pesticides 43–44Wine 7
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Corporate HeadquartersDionex Corporation1228 Titan WayP. O. Box 3603Sunnyvale, CA 94088-3603TEL: (408) 737- 0700FAX: (408) 730-9403
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© 2003 Dionex CorporationLPN 0666-05 PDF 08/03Printed in U.S.A.
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