Preformulation study of ivermectin raw material

Larissa Araujo Rolim • Flavia Cassia Maria dos Santos • Luıse Lopes Chaves •

Maria Luıza Carneiro Moura Goncalves • Jose Lourenco Freitas-Neto •

Andre Luiz da Silva do Nascimento • Jose Lamartine Soares-Sobrinho •

Miracy Muniz de Albuquerque • Maria do Carmo Alves de Lima •

Pedro Jose Rolim-Neto

Received: 30 January 2013 / Accepted: 9 February 2014

� Akademiai Kiado, Budapest, Hungary 2014

Abstract The aim of this study was to characterize the

raw material ivermectin (IVC) using different analytical

techniques used for drugs with its pharmacological char-

acteristics. Mass spectroscopic and infrared absorption

analyses were carried out to identify the molecule, and

further analyses to confirm its crystalline structure (elec-

tron sweep microscopy and X-ray diffraction), as well as

granulometric analysis, apparent, and compacted density,

leading to the conclusion that, even with a crystalline

structure IVC has good flow and compressibility. Differ-

ential Scanning Calorimetry and thermogravimetric ana-

lysis were used for the infrared thermal characterization,

determination of the melting point (157 �C), initial degra-

dation temperature (305 �C), loss of mass with the increase

of the temperature (3 events, the first dissolution and

degradation in two consecutive stages). Using the afore-

mentioned techniques, it was possible to carry out a com-

patibility study of IVC with some excipients used in solid

pharmaceutical form, which demonstrated an incompati-

bility between IVC and lactose and amide. These results

can be used to develop new pharmaceutical forms and for

more rational quality control of forms already commer-

cialized, with better understanding of the characteristics of

IVC.

Keywords Physico-chemical characterization � Raw

material � Analytical techniques

Introduction

Ivermectin is a semi-synthetic product obtained from

avermectin, naturally synthesized by the microorganism

Streptomyces avermitilis. It consists of a mixture of two

homologs dihydroavermectin B1a (H2B1a) and dihydro-

avermectin B1b (H2B1b) [1].

It is used as an active ingredient with broad-ranging

medical applications for the treatment of rashes, worms, and

lice, acting on the nervous system and functioning of the

muscles, resulting in paralysis and death of the parasites [2].

Ivermectin should be considered as a critical drug,

whose physical and chemical characteristics need to be

controlled in the pharmaceutical industry. Chemically, IVC

is a mixture of structural isomers, as mentioned, which act

in different ways and have different levels of toxicity. It

being expected that the raw material does not have less

than 80 % of the B1a isomer [3], while physically it pre-

sents itself as a poorly soluble drug with crystalline

structure (class II in the biopharmaceutical classification).

All these critical points need to be controlled when

obtaining technological forms of the drug.

The concept of quality by design (QbD) is systematic

and scientific, based on integral risk, and a proactive

approach to the development of medicines. It starts out

with previously defined objectives and emphasizes the

product and understanding of the manufacture process and

the control of processes [4]. QbD means guaranteed qual-

ity, improving the scientific methods that should be used in

the research and development and design phases, so that

the processing of the product is as quick as possible [5, 6].

QbD identifies characteristics of drugs that are critical for

quality from the point of view of production, which

translates into attributes that the drug should have to insure

safety and efficiency for users.

L. A. Rolim � F. C. M. dos Santos � L. L. Chaves �M. L. C. M. Goncalves � J. L. Freitas-Neto �A. L. da Silva do Nascimento � J. L. Soares-Sobrinho (&) �M. M. de Albuquerque � M. do Carmo Alves de Lima �P. J. Rolim-Neto

Universidade Federal de Pernambuco, Av. Prof. Arthur de Sa

S/N, Recife, PE, Brazil

e-mail: joselamartine@hotmail.com

123

J Therm Anal Calorim

DOI 10.1007/s10973-014-3691-9

Thus, there is an imminent need to review and refine the

physico-chemical characterization of IVC, since the phar-

macopeia tests required at present are not sufficient to

identify subtle differences between raw materials. This

evaluation may be one of the tools for establishing the

quality of the raw material, which, even though it origi-

nates from various manufacturers and is used for the

manufacture of pills, should provide discrimination of

results and consistent interpretations.

Experimental

Materials

Two batches of ivermectin were used, one of raw material

provided by Laboratorio Veterinario Vallee� (Batch

07016/2010, made in China), and the standard ivermectin

acquired from Sigma Aldrich�, batch 70288-86-7 (purity:

95 % H1B1a ? 2 % H1B1b) to calc the purity of raw

material used.

The binary mixtures (BM) 1:1 (p/p) of IVC with excipients

were prepared using a mortar and a pestle, breaking down the

mixtures for 3 min each. The BM were prepared using the

excipients: microcrystalline cellulose (MCC), talc, polyvinyl-

pyrrholidone (PVP) K-30, sodium croscarmellose, Starch�

(pre-gelatinized starch), Starlac� (85 % monohydrated lac-

tose ? 15 % starch—spray-dried compound), Flowlac�

(spray-dried monohydrated lactose), Tabletose� (monohy-

drated lactose), and Aerosil� (colloidal silicon dioxide).

Methods

Chemical identification

The infrared with fourier transform (FT-IR) spectrum were

obtained using a Spectrum 400 PerkinElmer� with an

attenuated total reflectance device (ATR) with a selenium

crystal. The samples to be analyzed were transferred

directly to the compartment of the ATR device, and the

result obtained using the mean of 10 sweeps, from 650 to

4000 cm-1 at a resolution of 4 cm-1. In addition to IR

identification, mass spectrometry was also carried out using

a Shimadzu� IT-TOF machine, with ionization by thermal

nebulization and flight time mass analyzer, the drug being

diluted using an acetonitrile:water system (50:50), by way

of positive ionization, with a 80–950 m/z scan.

Content determination and solubility study

The samples and the working standard IVC were obtained

using an initial dilution of 10 mg of IVC in solution of

acetonitrile:methanol:water (26:64:10), with 5 min of stir-

ring by sonication, in a 100 mL volumetric flask, to obtain

a final concentration of 200 lg mL-1, with subsequent

dilutions to 50, 100, and 150 lg mL-1 to obtain the cali-

bration curve for the raw material and plot three authentic

curves for 50, 75, 100, 125 and 150 lg mL-1 to determine

the linearity of the IVC standard, making it possible to

determine the IVC content of the raw material used for this

study.

First, the solutions obtained were submitted separately to

HPLC–DAD analysis, initially following the parameters laid

out by Cione and Silva [7], which involve the use of a C-18,

250 9 4.6 mm (5 lm) column, 25 �C, as the stationary

phase, a 0.2 % methanol:acetonitrile:acetic acid solution

(260:640:100 mL v/v), as the mobile phase, with a flow of

1.5 mL min-1 at 254 nm. After a number of analyses had

been conducted, the method was optimized, modifying the

composition and the flow of the mobile phase to 0.2 %

methanol:acetonitrile:phosphoric acid (260:640:100 mL

v/v), with a flow of 1 mL min-1 at 254 nm with completed

validation of this new method according ICH Q2(R1).

The solubility of IVC was determined by two studies. An

initial semi-quantitative study [3, 8] is to test the solvents:

ethyl acetate, acetone, acetonitrile, 0.2 M hydrochloric

acid, water, absolute ethyl alcohol, dichloromethane methyl

alcohol, ethylic ether, 0.2 M sodium hydroxide, n-hexane,

and 3 % hydrogen peroxide [5, 8]. In parallel, A quantita-

tive study was also conducted using the method described

for determining the IVC content by way of HPLC–DAD, in

which IVC samples were added to solutions of: water, pH

6.8 (50 mL of H2KPO4 0.1 M solution ? 23.65 mL NaOH

0.1 M solution ? water q.s.p 100 mL), pH 4.0 (9.35 mL

0.2 M Na2HPO4 ? 10.65 mL 0.1 M citric acid ? water

q.s.p. 100 mL), pH 1.2 (50 mL buffer solution H3BO3�KCl

0.1 M ? 97 mL 0.2 M HCl solution ? water q.s.p.

200 mL) with and without 1 % sodium lauryl sulfate (SLS),

until a saturated solution was obtained (with aliquots

removed at 24, 72, and 168 h).

Determining the physical and particle characteristics

The diffractograms for the drug were obtained using a SIE-

MENS� diffractometer (X-Ray Diffractometer, D-5000),

equipped with a copper anode. The samples were analyzed at

the 2h angle interval of 2–60 at a digitalization speed of 0.02�2h s-1. The samples were prepared on glass support with a

fine layer of powder material without solvent.

The crystal morphology of IVC was examined using an

electron sweep microscope Jeol� JSM-5900, after being

fixed on double-sided carbon tape and metalized with gold

for 15 min (Metalizer Baltec� SCD 050). The electromi-

crographs were obtained using a camera with an excitation

tension of 15 kV.

L. A. Rolim et al.

123

The distribution of particle sizes was determined by

sieving, using standardized stacked sieves (0, 75, 90, 150,

250, 425, 500 lm), mounted on a base equipped with

magnetic vibration (Tamizador Bertel�) for 20 min, in

triplicate.

The density of the powder was determined by an assay

with 10 g of the samples in an automatic compactor (Tap

Density, Varian�) equipped with a standard test tube, in

triplicate [7]. The initial volume occupied by the product

was measured, and then 10 compactions were carried out to

accommodate the powder. Then, further 1,000 consecutive

compactions were carried out until no change was observed

in the volume. The relation between the mass of the sam-

ples and the volume occupied by the powder before and

after compaction determined the apparent density (dAP)

and the compacted density (dCP). The compaction capacity

of the powder (10 g) was evaluated using the Hausner

Index (HI) and the Carr Index (CI) using the following

equations: HI = dCP/dAP e CI (%) = (dCP - dAP)/

dCP 9 100, respectively. The angle of repose was mea-

sured by the cone of powder formed by running the drug

through a funnel of standard dimensions onto a flat surface,

with 10 g of IVC. The flowing time was determined by the

average time needed for a pre-established quantity of drug

to run through the funnel, using a digital stop watch.

Thermal characterization of IVC

The thermal characterization of IVC was carried out using

differential scanning calorimetry(DSC) and thermogravi-

metric analysis (TG). The DSC curves were obtained using a

Shimadzu� DSC-60 calorimeter connected to Shimadzu�

TA-60WS software in a nitrogen atmosphere of 50 mL min-1

at various heating rates (5, 10, 15, and 20 �C min-1) in the

temperature range of 25–300 �C. The samples were placed in

a hermetically sealed aluminum sample holder with masses of

2 mg (±0.2) of sample, in triplicate. Indium and zinc were

used to calibrate the equipment in terms of the temperature

scale and enthalpy response.

The TG analyses were carried out using a Shimadzu�,

TGA Q60 thermoscale, in a nitrogen atmosphere flowing at

50 mL min-1, with a sample mass of around 5 mg (±0.4),

processed in a platinum bowl for the temperature range of

25–600 �C at a heating rate of 10 �C min-1. Prior to the

assays, the thermoscale was checked using hydrated cal-

cium oxalate.

The kinetic investigation of the non-isothermal degrada-

tion of IVC was obtained using TG data collected by

applying the Ozawa method. The heating rates used were 2.5,

5.0, 10, 20, and 40 �C min-1, within a temperature range of

30–600 �C, in platinum bowls with approximately 5 mg of

sample in a dynamic N2 atmosphere (50 mL min-1).

Study of drug-excipient compatibility

The fluxogram presented in Fig. 1 shows the protocol used

to carry out the study. In the first stage, IVC analyses were

carried out separately and in BM by DSC and TG, with

values referring to the initial melting point (Ti) and the

initial temperature of decomposition (Td) adopted as

indicative of the reaction between the drug and the

excipient.

The TG analyses used to calculate the degradation

kinetics, and FT-IR followed the same parameters descri-

bed for the analyses of IVC isolated.

Results and discussion

Chemical identification of IVC

Absorption spectrometry in the infrared region (IR) is

nowadays one of the main resources for structural identi-

fication of organic substances, and it is of extreme

importance in the control of the quality of raw materials

and excipients, as comparing spectra obtained from the raw

material with reference substances may reveal early deg-

radation, or characterize technological processes as in the

case of micro- and nano-particles which should normally

bond weakly with drugs to help improve their solubility or

vectorization. These bonds can be seen in the FT-IR

spectra.

The two isomers present in IVC raw material are dif-

ferentiated by a CH3 group in the structure as illustrated in

Figs. 2 and 3. Organic compounds with ether groups nor-

mally display characteristic bands related to C–O bonds, as

is the case of the IVC molecule which is composed only of

Binary Mixtures

DSC / TG

Interaction Signals?

Yes

No

FT-IR-ATRDegradationStudies Ozawa

Compatible

Fig. 1 Flowchart of study of drug-excipient compatibility

Preformulation study of ivermectin raw material

123

atoms of carbon, oxygen, and hydrogen, forming rings and

hydroxyl groups, ethers and a single ester group. In the

spectra obtained (Fig. 8), it was possible to identify 6

regions as inherent characteristics of IVC: at 3,500 cm-1,

relating to axial deformation of O–H; 2,964.99 and

2,937.33 cm-1, characteristics of methyl groups, axial

deformation of C–H; 1,728.92 cm-1, characteristic of sat-

urated aliphatic ketone; 1,675.79 cm-1 indicating unsatu-

rated lactones with a double bond adjacent to the –O–

group, owing to the C=C group; between 1,383.87 and

1,313.84 cm-1 showing moderate absorption of ketones, in

consequence of the axial and angular vibrations; 1,198.96

and 1,182.48 cm-1 indicating absorption of esters by lac-

tones; between 1,142.13 and 1,021.91 cm-1 shows the

absorption more characteristic of aliphatic ethers, owing to

the asymmetric axial deformation of C–O–C; 982.11 and

970.57 cm-1 and the two symmetrical angular deformation

bands outside the =C–H plane of the terminal alkenes;

950.78 and 929.74 cm-1 show angular deformation outside

the O–H plane; 904.25–832.48 cm-1 these intense bands in

the low-frequency region derived from angular deforma-

tion outside the plane of the C–H bonds of the ring;

807.96–706.11 cm-1 indicate angular deformation outside

the C–H plane; and 686.98 and 661.13 cm-1 angular

deformation outside the C=C plane of the rings.

The mass spectrum of IVC is very peculiar, thus it was

not possible to detect the raw mass of the drug as a char-

acteristic peak, the strongest peak was found at 897 m/z,

which corresponds to an H2B1a ? Na? dimer, followed by

secondary peaks at 284, 478 m/z, related to the fragmen-

tation of ionization, and 913 m/z (H2B1a ? K?). If IVC did

not have this capacity to form complexes the mass spec-

trum should have two peaks with different intensities

according to the quality of the raw material: 874 m/z

(CH2CH3–H2B1a) and 860 m/z (CH3–H2B1b) [9].

Content determination and solubility study (HPLC-

DAD)

IVC is obtained from the biosynthesis of microorganisms

such as S. avermectinius, which synthesize not only iver-

mectin but also other avermectins, and the pharmaceutical

raw material thus passes through a series of purification/

recrystallization steps in which the specification of purity

to be used as the pharmaceutical raw material is not less

than 80 % of H2B1a and no more than 20 % of H2B1b

calculated for an anhydrous sample, free of ethanol, and

dimethylformamide [10, 11].

The result of the average content study using HPLC–

DAD for the raw material was 93 ± 0.9 % of H2B1a in

relation to the standard, which is suitable for use, according

to official compendia [3, 12].

The absorption of drugs in solid orally administered

pharmaceutical forms depends on their release and

absorption under physiological conditions and the perme-

ability of the biological membranes. Based on these con-

siderations, it can be said that the dissolution of the drug in

H3C H3C

H3C

H3COH3CO

HO O

O

O

CH3

CH3

CH3

CH3

CH3

CH3

CH3H

O

O

OO

OH

OH

R

O

H

R =

R =

Bla

Blb

Fig. 2 Chemical structure of

IVC

100

80

60

40

20

0

200 400 600 800 1000 1200m/z

%

Positive ionization

897,48

913,47

478,27

284,30

Fig. 3 Mass spectrum of IVC

L. A. Rolim et al.

123

an aqueous medium is a limiting stage. The result of the

solubility study for IVC revealed low solubility in water

(Table 1), confirming the results already reported in the

literature [3].

For a more detailed evaluation a quantitative study of

solubility in aqueous systems was carried out using HPLC–

DAD, the results of which are described in Table 2.

It can be inferred from the results obtained that the

aqueous solubility of IVC is below the quantification limit

of the method (QL = 6.58 lg mL-1), calculated using the

standard linearity curve (y = 3568.51x - 30518.48),

which corroborates the finding in the literature of

4 lg mL-1 [13]. The other samples were analyzed at 72 h,

since the media used were already saturated in this time.

In addition, regarding the results obtained from the

solutions with different pH, it was observed that the solu-

bility increases with the increasing of the pH of the solu-

tion. This result was expected once the molecule is

extremely apolar, with a pKa value around 6.5, what means

that the molecule must be in a solution with a very alkaline

pH to be ionized, what may lead to the increase of the

solubility. These results were important to predict the

solubility and consequently the permeation of IVC in

biological fluids, which can help to understand the

absorption profile, and thus the effectiveness.

Determination of physical and particle characteristics

X-ray diffraction is a highly versatile and rapid technique

for the application of polycrystalline samples, such as

monitoring samples in the development of pharmaceutical

products in the laboratory and industrial quality control,

providing information on the size and structure of crystals.

This technique is based on the principal that when a

material is exposed to monochromatic X-rays, for a per-

fectly aligned crystal, in which the atoms are regularly

packed and the distance between the crystallographic

planes is defined by the physical characteristics of the

sample, which can be used precisely to measure the spaces

in the crystalline reticle.

This assay established the diffraction pattern of the IVC

powder, revealing the presence of a series of peaks, with a

distinct peak at 2h around 9.32�, apart from other sec-

ondary lower intensity peaks at 9.04�, 12.36�, 13.16�,

13.56�, 18.12�, 18.7�, and 27.32� (Fig. 4), showing the

typically crystalline behavior of the drug, which may be

relevant to its solubilization and fluidity.

Table 1 Result of semi-quantitative solubility study of IVC

Solvent Solubilization

volume/mL

Classification Descriptive

term

Ethyl acetate 10 Easily

soluble

From 1 to 10

parts

Acetonitrile 30 Soluble From 10 to 30

parts

Acetone 100–1,000 Poorly

soluble

From 100 to

1,000 parts

Water [10,000 Practically

insoluble

[10,000 parts

Ethyl alcohol 30–70 Slightly

soluble

From 30 to 100

parts

Methyl alcohol 30 Easily

soluble

From 10 to 30

parts

Dichloromethane 10 Practically

insoluble

From 1 to 10

parts

Ethylic ether 30 Practically

insoluble

From 10 to 30

parts

H2O2 3 % [10,000 Practically

insoluble

[10,000 parts

HCl 0.2 M [10,000 Very poorly

soluble

[10,000 parts

NaOH 0.2 M [10,000 [10,000 parts

N-Hexane 1,000–10,000 From 1,000 to

10,000 parts

Table 2 Quali-quantitative determination of solubility in aqueous

systems

Samples Time/h Content (lg mL-1)

H2O 168 –

H2O ? SLS 1 % 72 10.43

pH 1.2 72 819.41

pH 1.2 ? SLS 1 % 72 61.86

pH 4.0 72 11,403.30

pH 4.0 ? SLS 1 % 72 259.01

pH 6.8 72 21,747.73

pH 6.8 ? SLS 1 % 72 23,756.06

6000

5000

4000

3000

2000

1000

0

5 10 15 20 25 30 35 40 45 50

2 θ/°

Inte

nsity

/%

13,1618,12

18,7

9,32

Fig. 4 Diffractogram and electromicrograph of IVC crystalline

particles

Preformulation study of ivermectin raw material

123

The granulometric distribution carried out (Fig. 5)

showed that the particles of the raw material used for the

study tend to cluster in the 90- to 250-lm interval.

In the light of these results, an evaluation was carried

out of the retention in relation to the passage of particles in

the intervals under study. It was shown that the mean size

of the particles is approximately 198 lm. The raw material

used can thus be classified as a semi-fine powder [6]. This

knowledge is of great importance, given that the speed of

dissolution is directly proportional to the surface area of

particles and, as dissolution is a critical factor for this drug,

determining the size and morphology of IVC particles

allows these factors to be correlated.

The physical and mechanical properties of the com-

paction and rheology of IVC were determined, and the

results are shown in Table 3.

The data showed that the density of IVC was accordingly

with those reported in the literature. However, the Carr

Index expressed in the form of a percentage the compaction

capacity of the powder under analysis, where values up to

10 % are considered to demonstrate excellent flow and

compaction, as observed in the case of IVC. Furthermore,

the Hausner Index, which is similar to the CI, lies between

1.00 and 1.11 demonstrating that the flow is excellent and

there is no need for the addition of lubricants to improve the

flow, while higher values demonstrate poor flow.

Likewise, by determining the angle of rest, it was

observed that IVC flows freely (angle of rest = 28� ± 3�),

at 2 ± 1 s, when tested a mass of 10 g. This behavior

diverges from that expected given the crystalline mor-

phology of the particles (Fig. 6), which favors interaction

between the particles. The drug can therefore be processed

by way of direct compression, since, despite the crystalline

structure of IVC, it has a good flow and good

compressibility.

Thermal characterization of IVC

The thermo-analytic methods outlined in the fifth edition of

the Brazilian Pharmacopeia of 2010 have been widely

applied to the study of drugs. Many studies have used these

methods as alternatives for the characterization and quality

control of pharmaceutical materials.

The DSC curves obtained to confirm the melting range

of IVC (Fig. 6) at heating rates of 5, 10, 15, and

20 �C min-1 showed an endothermic peak in the

152.96–164.2 �C temperature range (mean), characteristic

of the melting of the drug, although the melting peak was

different when analyzed at different rates, occurring sooner

the smaller the heating rate used. According to the Merck

Index, 1999 [13], the melting band of IVC is 155–157 �C,

and the most appropriate heating rate for thermal evalua-

tion of IVC is 10 �C min-1 which allows a melting point of

157.4 ± 0.7 �C to be determined.

The purity of IVC was also calculated using Van’t Hoff

equation in the linearization of the melting event with

analysis obtained at a rate of 0.5 �C min-1, confirmed in

triplicate (and HPLC–DAD analysis, as described above).

In this model, purity is determined through the deviation

from linearity of the melting point, which occurs because

of the presence of impurities. Knowledge of the deviation

from linearity allows the correction factor in linearization

of the straight line to be inferred. The purity of IVC was

found to be around 98.4 %, with a calculated correction

factor for impurities of 8.36 %, which does not concur with

the content found using HPLC–DAD, as this technique is

inappropriate for determining the purity of IVC, probably

because of the enthalpy involved in the process being

related not only to the melting of the drug, but also to the

evaporation of residual solvents as will be discussed below.

The TG curves provided information on the composition

and thermal stability of IVC, thereby making it possible to

determine the initial decomposition temperature for the raw

material and the stages in the degradation of IVC, tracking

any possible dehydration, oxidation, combustion or

decomposition reactions.

The TG curves for the raw material and the analytical

pattern showed that IVC has three characteristic thermal

events, the first loss of mass occurs when the drug melts,

between 153 and 164 �C (Dm = 5.04 %), a second stage of

decomposition between 312 and 327 �C (Dm = 58.54 %),

30

25

20

15

10

5

00 100 200 400 500

μm

%

Fig. 5 Distribution of frequency of IVC particle size

Table 3 Results for compaction properties of sample under analysis

Parameters Values obtained

dap/g mL-1 0.67

dcp/g mL-1 0.74

IH 1.10

CI 9.46

L. A. Rolim et al.

123

followed by a third and last degradation stage between 341

and 427 �C (Dm = 29.35 %) related to the carbonization

of IVC (Fig. 7), events determined by the first TG

derivative.

The IVC raw material has formamide and ethanol in its

crystalline network, which are solvents used for purifica-

tion/recrystallization of the mixture of avermectins pro-

duced by biosynthesis of microorganisms. According to the

specifications contained in the official compendia, analysis

of these residual solvents should be carried out by way of

gaseous chromatography (GC) with a maximum of 5 %

ethanol and 3 % dimethylformamide [3]. However, it was

found that TG analysis is also capable of quantifying the

total residual solvent content, corresponding mostly to the

first event involving loss of mass the IVC raw material

undergoes.

To prove that the first event involving loss of mass in

IVC is not properly related to degradation, but a desolva-

tion of ethanol and dimethylformamide, one TG analysis

was carried out on a sample without pre-heating (sample 1)

and another with an IVC sample pre-heated to the melting

point, with subsequent re-cooling and re-heating of the

sample to 600 �C (sample 2), curves shown in Fig. 7.

The analysis of the TG curves obtained shows that, on

the first heating, the solvents evaporate (Tboiling etha-

nol = 78.4 �C and Tboiling dimethylformamide = 153 �C)

[14], which may justify the decreasing in the melting point

of IVC (157 �C) in the second heating, once this temper-

ature, coincides, approximately, with the boiling point of

the dimethylformamide, corresponding to 6.5 % on aver-

age, a value similar to that calculated by CG for the ana-

lytical standard (sum of the data provided by Sigma

Aldrich� relating to the quantity of dimethylformamide

and ethanol in the batch used in this study) added to the

water content by Karl Fisher.

DSCmW

0.0

–10.0

–20.0

50.0 100.0 150.0 200.0 250.0 300.0 350.0 400.0

Temperature/°C

Endo 160.03C20 °C/min

15 °C/min

10 °C/min

5 °C/min154.52C

157.23C

158.29C

Fig. 6 DSC curves for

establishing heating rate for

IVC

100

80

60

40

20

–0

–0 50 100 150 200 250 300 350 400 450 500 550 600

Temperature/°C

40.0

20.0

0.0

–20.0

–40.0

Hea

t flo

w/m

W m

g–1

Mas

s/%

TG sample 1

TG sample 2

DTA sample 2

DTA sample 1

Endo

Fig. 7 TG curves for IVC

without pre-heating (sample 1)

and with pre-heating to 157 �C

(sample 2) in an atmosphere of

N2 at 50 mL min-1

100

90

80

70

60

50

40

304000 3500 3000 2500 2000 1500 1000

cm–1

T/%

Sample 1

Sample 2

Fig. 8 FT-IR spectrum of samples of IVC without pre-heating

(sample 1) and with pre-heating to 157 �C (sample 2)

Preformulation study of ivermectin raw material

123

FT-IR analysis was also carried out on a sample pre-

heated until the melting point of IVC (sample 1) and a

sample without pre-heating (sample 2) as shown in Fig. 8,

revealing the permanence of the characteristic bands of IVC

even after heating, with no signs of degradation of the drug.

TG analysis of IVC is thus characterized as an auxiliary

technique for determining the quantity of residual solvents

resulting from purification/recrystallization of this drug.

As the first event involving loss of mass on the TG curve

does not constitute a decomposition event, non-isothermic

kinetic analysis of IVC was carried out to evaluate only the

second mass loss event (310–330 �C) [14, 15].

In these analyses the TG curves shift to higher temper-

atures as the heating rate increases (2.5, 5, 10, 20, and

40 �C min-1), allowing the application of the Ozawa

method, through good correlation between the five heating

100

70

50

20

0.0100 200 300 400 500

Temperature/°C

Rz 10 °C min–1

Rz 5 °C min–1

Rz 2,5 °C min–1Rz 40 °C min–1

Rz 20 °C min–1

Mas

s lo

ss/%

LOG A2.00

1.50

1.00

0.50

0.00

1.42 1.52 1.62× 10–3

× 10–18

10.00

8.00

6.00

4.00

0.00 1.00 2.00 3.00 4.00

Reduced time/min

Kinetic EnergyOrderFrequency Factor

217.47 KJ mol–1

2.01.539 × 1016 min–1

1/T/K

(a)

(b)

(c)

Fig. 9 TG curves and Ozawa

graph for IVC obtained for five

heating rates under a dynamic

nitrogen atmosphere using a

non-isothermic method

L. A. Rolim et al.

123

rates. For this thermal decomposition, the Ae was calcu-

lated to be 217.47 kJ mol-1, with a second-order degra-

dation reaction and a factor of frequency of collisions

between IVC molecules of 1,539 9 1,016 min-1 (Fig. 9).

Drug-excipient compatibility study

The behavior of binary mixtures showed that, with all

excipients, there was a significant decrease in the enthalpy

involved in the melting process, when compared with the

melting enthalpy of the drug alone, as well as small

changes in the shape of the peak, with few variations in the

melting temperature, suggesting no incompatibility in most

cases. The data from the TG and DSC curves obtained in

the compatibility study are shown in Tables 4 and 5.

In the DSC curves of the BM of IVC with lactose ex-

cipients (Tabletose� and Flowlac�), there was a reduction

in the melting temperature of 1–3 �C from the start of the

melting temperature, which may indicate the occurrence of

a drug-excipient interaction. Furthermore, the initial deg-

radation temperature (the second event involving loss of

mass) decreased by 10 �C in the BM with lactose and

amide (Starch�, Tabletose�, Flowlac�, and Starlac�),

according to the TG/DTG curves.

Tabletose� was therefore selected to represent excipi-

ents containing lactose and Starch� for those containing

amide and the non-isothermic kinetics was evaluated at

rates of 10, 20, and 40 �C min-1 for the IVC raw material

and the BM containing these excipients.

Table 4 Thermoanalytical data for IVC and BM with excipients

Samples Tonset/�C Tmelting/�C DH/J g-1 Tonset degradation/�C % Degradation

IVC 144.04 159.42 -134.58 305.29 -53.59

IVC ? PVP K-30 149.54 160.01 -36.24 307.49 -27.62

IVC ? Talc 144.12 157.8 -152.8 307.06 -50.02

IVC ? CMC 147.18 159.53 -33.18 310.12 -34.59

IVC ? Starch� 145.47 161.28 -33.92 297.83 -63.88

IVC ? Tabletose� 142.9 149.9 -129.33 292.77 -59.6

IVC ? Croscarm. 144.57 156.57 -26.37 305.69 -51.92

IVC ? Flowlac� 141.27 149.35 -137.44 284.99 -50.74

IVC ? Starlac� 144.98 150.91 -99.4 293.66 -60.69

IVC ? Glycolate 144.06 156.5 -35.45 306.3 -52.4

IVC ? Aerosil� 147.9 162.63 -59.58 308.36 -27.36

Table 5 Activation energy (Ae), frequency factor (A) of BM

obtained by non-isothermic kinetics (Ozawa method)

Sample Activation energy (Ae) Frequency factor (A)

IVC 216.03 kJ mol-1 1.620 9 1016 min-1

IVC/Tabletose� 180.21 kJ mol-1 2.985 9 1013 min-1

IVC/Starch� 200.44 kJ mol-1 7.632 9 1014 min-1

IVC/TABLETOSE

TABLETOSE

IVC/Starch

STARCH

IVC

4000 3500 3000 2500 2000 1500 1000

cm–1

Fig. 10 FT-IR spectrum of IVC, Tabletose�, Starch�, IVC/Table-

tose� binary mixture, IVC/Starch� binary mixture

Preformulation study of ivermectin raw material

123

The activation energy of the IVC/Tabletose� binary

mixture decreased by more than 15 % compared with IVC

alone, while the IVC/Starch� mixture presented a reduc-

tion of 7 % in the activation energy, corroborating the

results of the thermal analysis, which produced evidence of

interaction.

The FT-IRIV absorption spectra for IVC and the BM

with the selected excipients are shown in Fig. 10. In all the

spectra, there is only superposition of the characteristic

bands of IVC in isolation and of the excipients, some of

these having bands relating to the excipients superposed on

them, although it should not be considered incompatibility,

and the FT-IR technique was thus not selected to provide

evidence of possible incompatibilities between IVC and the

excipients under evaluation.

Conclusions

Physico-chemical characterization confirmed data reported

in the scientific literature, insuring the authenticity of the

material under analysis. Quantification of the purity of the

drug showed a good correlation between the data obtained

using chromatographic and thermo-analytic techniques,

showing that thermoanalytical techniques can be a pow-

erful tool for evaluation of the purity of the substance. The

Ozawa method showed the second-order kinetic behavior

for decomposition of the drug, and the frequency factor for

collisions between IVC molecules of 1,539 9 1016 min-1.

The compatibility studies using DTG and TG showed that

the drug-excipient association between IVC, Tabletose�

and Starch� produces an increase in the percentage

decomposition, as well as decrease in the Ae, seen in the

analyses of non-isometric kinetics.

The results obtained are thus of fundamental impor-

tance, as they have enabled the determination of the prin-

ciple physical and chemical properties of IVC, providing

relevant information on the quality of raw material used

and its behavior under a range of different techniques

frequently used to characterize pharmaceutical products,

which have been being developed for this drug as a way of

overcoming this characteristic which represents a techno-

logical and therapeutic barrier in the case of IVC, an anti-

helminth drug used to combat filariasis.

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