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Journal of Pharmaceutical and Biomedical Analysis

Volume 41, Issue 2, 3 May 2006, Pages 533539

Determination of clopidogrel metabolite (SR26334) in humanplasma by LCMS


Hanna Ksycinska , ,
Piotr Rudzki,
Mirosawa Bukowska-Kiliszek
Department of Pharmacology, Pharmaceutical Research Institute, 8 Rydygiera Street,
01-793 Warsaw, Poland
http://dx.doi.org/10.1016/j.jpba.2005.11.035, How to Cite or Link Using DOI
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Abstract
A new, sensitive, specific and
reproducible
method
for determination
of clopidogrel metabolite (SR26334) in human plasma has been developed. After liquid
liquid extraction on Chem Elut cartridges with dichloromethane, samples were quantified
using reversed-phase high performance liquid chromatography with mass detection. The
determination was performed on a Luna C18, 3 m (75 mm 4.6 mm i.d.) column with an
acetonitrile-water-formic acid mixture (60:40:0.1, v/v/v) as a mobile phase. The flow rate was
set at 0.2 mL/min. Repaglinide was chosen as an internal standard and the time of analysis
was 12 min. For SR26334 the limits of detection and quantification were 7.5 ng/mL and
20 ng/mL, respectively, and the calibration curve was linear up to 3000 ng/mL. The
extraction recovery of SR26334 from plasmawas within the range of 8590%. The method
has been successfully used to study clopidogrel metabolite pharmacokinetics in healthy
volunteers.

Journal of Pharmaceutical and Biomedical Analysis


Volume 48, Issue 4, 1 December 2008, Pages 12191224
Short communication

Quantitative determination of clopidogrel active metabolite in human plasma by LC


MS/MS
Makoto Takahashia, , ,
Henrianna Pangb,
Kiyoshi Kawabatac,
Nagy A. Faridd,
Atsushi Kuriharaa

Drug Metabolism & Pharmacokinetics Research Laboratories, Daiichi Sankyo Co.,


Ltd., 1-2-58 Hiromachi, Shinagawa-ku, Tokyo 140-8710, Japan
b
NoAb BioDiscoveries Inc., 2820 Argentia Road, Unit 8, Mississauga, Ontario L5N
8G4, Canada
c
Research Department II, Daiichi Sankyo RD ASSOCIE Co., Ltd., 3-10-2
Kitashinagawa, Shinagawa-ku, Tokyo 140-0001, Japan
d
Lilly Research Laboratories, Eli Lilly and Company, Lilly Corporate Center,
Indianapolis, IN 46285, USA
http://dx.doi.org/10.1016/j.jpba.2008.08.020, How to Cite or Link Using DOI
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Abstract
A quantitative method for the determination of clopidogrel active metabolite (AM) in
human plasma was developed and validated using liquid chromatographytandem mass
spectrometry (LCMS/MS).Clopidogrel AM contains a thiol group, thus requiring
stabilization in biological samples. The alkylating reagent 2-bromo-3-methoxyacetophenone
was used to stabilize clopidogrel AM in blood. An analog of the derivatized clopidogrel AM
was used as the internal standard (IS). The derivatized samples were subjected to solid-phase
extraction with a C2 disk plate and the overall procedure exhibited good reaction (more than
90%) and recovery efficiencies (from 85% to 105%). The derivative of clopidogrel AM (MPAM) and IS were separated on an ODS column and quantified by tandem mass spectrometry
with electrospray ionization. No significant endogenous peaks corresponding to MP-AM or
IS were detected in blank human plasma samples, and no significant matrix effect was
observed for MP-AM and IS in humanplasma samples (from 102% to 121%). The calibration
curve ranged from 0.5 to 250 ng/mL with good linearity, and extended by validation of a 50fold dilution. In the intra- and inter-assay reproducibility tests, the accuracy and precision
were within 12% relative error and 6% coefficient of variation, respectively. The derivatized
MP-AM was stable in human plasma for 4 months at 80 C. The validated method was
successfully used to analyze clinical samples and determine the pharmacokinetics
of clopidogrel AM.

Research Article

1.
2.
3.
4.

Quantification of clopidogrel in human plasma by sensitive liquid


chromatography/tandem mass spectrometry
Ramakrishna V. S. Nirogi*,
Vishwottam N. Kandikere,
Manoj Shukla,
Koteshwara Mudigonda,

5.
6.

Santosh Maurya,
Ravikumar Boosi
Article first published online: 24 APR 2006
DOI: 10.1002/rcm.2497
Copyright 2006 John Wiley & Sons, Ltd.
Issue
Rapid Communications in Mass Spectrometry
Volume 20, Issue 11, pages 16951700, 15 June 2006
Abstract
A simple, sensitive and rapid high-performance liquid chromatography/positive electrospray
ionization tandem mass spectrometry method was developed and validated for the assay of
clopidogrel in human plasma. Following liquid-liquid extraction, the analytes were separated
using an isocratic mobile phase on a reversed-phase column and analyzed by mass
spectrometry in the multiple reaction monitoring mode using the respective [M+H]
+
ions, m/z 322/212 for clopidogrel and m/z 264/154 for the internal standard. The assay
exhibited a linear dynamic range of 56000pg/mL for clopidogrel in human plasma. The
lower limit of quantification was 5pg/mL with a relative standard deviation of less than 8%.
Acceptable precision and accuracy were obtained for concentrations over the standard curve
range. A run time of 2.5min for each sample made it possible to analyze more than 400
human plasma samples per day. The validated method has been successfully used to analyze
human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence
studies.

Journal of Chromatography B
Volume 848, Issue 2, 1 April 2007, Pages 344354

The validation of a bioanalytical method for the determination ofclopidogrel in


human plasma
A. Robinsona, , ,
J. Hillisa,
C. Neala,
A.C. Learyb
a
HFL Ltd., Newmarket Road, Fordham, Cambridgeshire CB7 5WW, UK
b
Shandon Clinical Trials Ltd., 9 John Redmond Street, Cork, Ireland
http://dx.doi.org/10.1016/j.jchromb.2006.10.076, How to Cite or Link Using DOI
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Abstract
A fast, sensitive and specific LCMS/MS bioanalytical method for the determination of
unchanged clopidogrel in human plasma has been developed and validated over the range of
1012,000 pg mL1 (r20.9993) by the Contract Research group at HFL. Samples (0.3 mL)
were buffered (pH 6.8), extracted using diethyl ether and 10 L of the sample extract was
injected onto the LCMS/MS system. Analysis was performed using a C8 column
(temperature controlled to 50 C) by gradient elution at a flow rate of 0.9 mL min1 over a
3 min run time. Retention times of 1.61 and 1.59 min were observed for clopidogrel and 2H3clopidogrel (I.S.), respectively. Detection was achieved using a Sciex API 4000, triple
quadrupole mass spectrometer, in positive TurboIonspray (electrospray) ionisation mode.
Ion transitions were monitored using MRM (multiple reaction monitoring)
for clopidogrel (m/z 322212) and for2H3-clopidogrel (m/z 327217). This validated method
was used to support a pharmacokinetic study in healthy volunteers.
Rapid LCESI-MSMS Method for the Simultaneous Determination of Clopidogrel
and its Carboxylic Acid Metabolite in Human Plasma
Oxford Journals
Mathematics & Physical Sciences
Journal of Chromatographic Science
Volume 46, Issue 10
Pp. 867-875.
1.
2.
3.
4.
5.

Nitesh K. Patel1,2,
Gunta Subbaiah2,2,
Hiten Shah2,2,
Mohan Mohan2,2 and
Pranav S. Shrivastav*,1
+Author Affiliations
1. 1Chemistry Department, School of Sciences, Gujarat
University, Navrangpura, Ahmedabad 380 009, India
2. 2Bioanalytical Laboratory, Torrent Research Center, At Village Bhat,
Gandhinagar 382 428, India
1.
*Author to whom correspondence should be addressed:
emailpranav_shrivastav@yahoo.com.

Received September 1, 2007.

Revision received January 28, 2008.


Abstract
A high throughput liquid chromatography.electrospray ionization tandem mass spectrometry
(LC.ESI-MSMS) method is developed for the simultaneous estimation of clopidogrel
(SR25990C) and its carboxylic acid metabolite (SR26334) in human plasma using

glimepiride as internal standard. The extraction of SR25990C, its metabolite, and IS from the
plasma (0.3 mL) involves treatment with phosphoric acid followed by solid-phase extraction
(SPE). Sample preparation by this method yields clean extracts with quantitative and
consistent mean recoveries of 98.05%, 85.45%, and 105.72% for SR25990C, SR26334, and
IS, respectively. The SPE eluate without drying and reconstitution is analyzed by LCMS
MS, operating in the positive ion and selective reaction monitoring mode. The injection
volume is 2 L with a total chromatographic run time of 5.0 min. The method response is
linear over the dynamic range of 0.25 to 25.0 ng/mL for SR25990C and 50.0 to 6000.0 ng/mL
for SR26334, with correlation coefficients of r 0.9989 and 0.9984, respectively. The
method is validated to demonstrate its specificity, linearity, accuracy, precision, recovery,
matrix effect, dilution integrity, and stability studies. It is applied to study the bioavailability
of 75 mg clopidogrel mesylate tablets in 16 human subjects with satisfactory results.

Short Communication
Determination of clopidogrel in human plasma by liquid chromatography/tandem
mass spectrometry: application to a clinical pharmacokinetic study
1.
Beom Soo Shin1,
2.
Sun Dong Yoo2,*
Article first published online: 2 MAY 2007
DOI: 10.1002/bmc.850
Copyright 2007 John Wiley & Sons, Ltd.
Issue
Biomedical Chromatography
Volume 21, Issue 9, pages 883889, September 2007

Keywords:
clopidogrel;
LC/MS/MS;
pharmacokinetics
Abstract
A rapid and sensitive LC/MS/MS assay was developed and validated for the determination of
clopidogrel in human plasma. Clopidogrel was extracted by single liquidliquid extraction
with pentane, and chromatographic separations were achieved on a C18 column. The method
was validated to demonstrate the specificity, linearity, recovery, lower limit of quantification
(LLOQ), stability, accuracy and precision. The multiple reaction monitoring was based
on m/z transition of 322.2 211.9 for clopidogrel and 264.1 125.1 for ticlopidine
(internal standard). The total analytical run time was relatively short (3 min), and the LLOQ
was 10 pg/mL using 0.5 mL of human plasma. The assay was linear over a concentration
range from 10 to 10,000 pg/mL (r > 0.999). The intra- and inter-day accuracies were 101.3

108.8 and 98.4103.5%, respectively, and the intra- and inter-day assay precisions were 1.9
5.5 and 4.48.1%, respectively. The developed assay method was applied to a
pharmacokinetic study in human volunteers after oral administration of clopidogrel at a dose
of 150 mg. Copyright 2007 John Wiley & Sons, Ltd.

Determination of Clopidogrel in Human Plasma by a Validated LC-MS/MS Method


Dobrin A Svinarov & Lilia V Kasabova
Central Lab of TDM & Clin Pharmacol, Faculty of Medicine, Medical University, Sofia,
Ouside US/Canada, Bulgaria
http://www.ascponline.org/ascp-static/planner_public/abstract_popup.php?abstractno=272
Clopidogrel (CPD) is prodrug converted endogenously to an unstable active metabolite and
an inactive carboxylic acid metabolite. Therefore, CPD is the required analyte in
bioequivalence and bioavailability studies. Development and validation of an LC-MS/MS
determination of CPD was performed with the aim to be applied for pharmacokinetic
analysis. CPD and ticlopidine (TPD, internal standard) were extracted from 200 L of human
plasma with ethylacetate. Chromatographic separation was performed on a C18 analytic
column with isocratic elution, using a mobile phase consisting of methanol and ammonium
acetateformic acid buffer. Positive-ion electrospray ionization and selected reaction
monitoring were used to follow the predominant transitions: collision energy (CE) = 17 at
m/z 322 212 for CPD, and CE = 17 at m/z 264 154 for TPD. Selectivity was assessed
with 6 different individual sources of human plasma and confirmed with matrix effect (ME)
averaging 88% to 89% for CPD, 98% to 102% for TPD, and relative ME of 86% to 91% for
CPD. Accuracy and precision (within-run and between-runs) were all within 12%. Extraction
recoveries averaged 69% to 79% (CPD) and 73% to 79% (TPD); linearity range was 5.4
2,160 ng/L, R2 > 0.99. Freeze-thaw stability was determined for 3 cycles each lasting 24
hours, postpreparative stability was documented for 72 hours at 8oC, short-term stability at
ambient temperature was proven for 12 hours in the dark and for 2 hours at daylight; stock
solution stability and long-term stability in plasma - for 122 days at 20oC. With run time of
less than 2.5 minutes, a throughput of more than 150 samples per working day can be
achieved. Lack of back-conversion of CPD carboxylic acid metabolite to CPD was assured
by avoidance of any alcohols in the procedure and confirmed by analysis of samples spiked
with that metabolite. The method was validated according to FDA requirements and allows
the accurate and precise determination of CPD in human plasma.

Short Communication
Ultra-performance LC MS/MS method for quantification of clopidogrel active
metabolite
1.
Xavier Delavenne1,2,*,
2.
Thierry Basset1,2,

3.
4.
5.
6.

Paul Zufferey2,3,
Nora Malouk2,
Silvy Laporte2,4,5,
Patrick Mismetti2,4
Article first published online: 7 JUN 2010
DOI: 10.1002/jssc.201000115
Copyright 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Issue

Journal of Separation Science


Volume 33, Issue 13, pages 19681972, July 2010
Abstract
A sensible ultra-performance LCMS method was developed and validated for the
quantification of clopidogrel active metabolite in human plasma, with clopidogrel D4 as
internal standard. Plasma pretreatment involved a one-step protein precipitation with
acetonitrile. The separation was performed by reverse-phase chromatography on a C8
column. The method was linear over the concentration range of 1150ng/mL. The intra- and
inter-day precision values were below 17% and accuracy was from 1.7 to 7.5% at all quality
control levels. The lower LOQ was 0.8ng/mL. Sample analysis time was reduced to 5min
including sample preparation (limited to protein precipitation). The present method was
successfully applied to a clopidogrel active metabolite pharmacokinetic study following oral
administration to healthy volunteers.

Short Communication
Determination of clopidogrel in human plasma by liquid chromatography/tandem
mass spectrometry: application to a clinical pharmacokinetic study
1.
Beom Soo Shin1,
2.
Sun Dong Yoo2,*
Article first published online: 2 MAY 2007
DOI: 10.1002/bmc.850
Copyright 2007 John Wiley & Sons, Ltd.
Issue
Biomedical Chromatography
Volume 21, Issue 9, pages 883889, September 2007

Abstract
A rapid and sensitive LC/MS/MS assay was developed and validated for the determination of
clopidogrel in human plasma. Clopidogrel was extracted by single liquidliquid extraction
with pentane, and chromatographic separations were achieved on a C18 column. The method
was validated to demonstrate the specificity, linearity, recovery, lower limit of quantification
(LLOQ), stability, accuracy and precision. The multiple reaction monitoring was based
on m/z transition of 322.2 211.9 for clopidogrel and 264.1 125.1 for ticlopidine
(internal standard). The total analytical run time was relatively short (3 min), and the LLOQ
was 10 pg/mL using 0.5 mL of human plasma. The assay was linear over a concentration
range from 10 to 10,000 pg/mL (r > 0.999). The intra- and inter-day accuracies were 101.3
108.8 and 98.4103.5%, respectively, and the intra- and inter-day assay precisions were 1.9
5.5 and 4.48.1%, respectively. The developed assay method was applied to a
pharmacokinetic study in human volunteers after oral administration of clopidogrel at a dose
of 150 mg.

J Chromatogr B Analyt Technol Biomed Life Sci. 2010 Nov 15;878(30):3134-42. Epub 2010
Oct 1.
Development and validation of an HPLC-MS/MS method to determine
clopidogrel in human plasma. Use of incurred samples to test backconversion.
Silvestro L, Gheorghe MC, Tarcomnicu I, Savu S, Savu SR, Iordachescu A, Dulea C.
Source
3S-Pharmacological Consultation & Research GmbH, Koenigsbergerstrasse 1, 27243
Harpstedt, Germany. cristina.vilciu@3spharma.ro
Abstract
Quantitative methods using LC-MS/MS allow achievement of adequate sensitivity for
pharmacokinetic studies with clopidogrel; three such methods, with LLOQs as low as 5
pg/mL, were developed and fully validated according to the well established FDA 2001
guidelines. The chromatographic separations were performed on reversed phase columns
Ascentis RP-Amide (15 cm x 2.1 mm, 5 m), Ascentis Express C8 (10 cm x 2.1 mm, 2.7 m)
and Ascentis Express RP Amide (10 cm x 2.1 mm, 2.7 m), respectively. Positive
electrospray ionization in MRM mode was employed for the detection and a deuterated
analogue (d3-clopidogrel) was used as internal standard. Linearity, precision, extraction
recovery, matrix effects and stability tests on blank plasma spiked with clopidogrel and stored

in different conditions met the acceptance criteria. During the analysis of the real samples
from the first pharmacokinetic study, a significant increase (>100%) of the measured
clopidogrel concentrations in the extracts kept in the autosampler at 10 C was observed.
Investigations led to the conclusion that most probably a back-conversion of one or more of
the clopidogrel metabolites is occurring. The next methods were optimized in order to
minimize this back-conversion. After a series of experiments, the adjustment of the sample
preparation (e.g. processing at low temperature and introducing a clean-up step on Supelco
HybridSPE-Precipitation cartridges) has proven to be the most effective in order to improve
the stability of the extracts. Incurred samples of real subjects were successfully used in the
validation of the last two analytical methods to evaluate the back-conversion, while tests
using only the known metabolites could not detect this important problem.

CHROMATOGRAPHIA
Volume 70, Numbers 11-12 (2009), 1581-1586, DOI: 10.1365/s10337-009-1349-8
ORIGINAL
Simultaneous Determination of Clopidogrel and Its Carboxylic Acid Metabolite (SR26334) in
Human Plasma by LCESIMSMS: Application to the Therapeutic Drug Monitoring of
Clopidogrel
Jian-Jun Zou, Hong-Wei Fan, Da-Qing Guo, Ying-Bin Li, Song Lin, Yu-Bing Zhu, Cui-Xia
Yu, Jie Zhou, Jiang-Hui Liu and Yun-Fang Hu
Abstract
A sensitive and specific liquid chromatography-tandem-mass spectrometry method was
developed and validated for the simultaneous determination of clopidogrel and its carboxylic
acid metabolite (SR26334) in human plasma using nateglinide and pioglitazone as internal
standards. Analytes were extracted from 0.50 mL of plasma using diethyl ethern-hexane
(4:1, v/v). Chromatographic separation was performed on a Teknokroma C18 column with a
mobile phase of methanolwater (containing 0.1% formic acid) (80:20, v/v) at a flow rate of
0.20 mL min1 within 5.6 min. Linearity was established over the concentration range of
0.0055 ng mL1 for clopidogrel and 202,500 ng mL1 for SR26334. Intra- and inter-batch
standard deviations were less than 9.2% and the accuracy of this assay was found to fall
within an acceptable range 10.0%. The method was successfully applied to the therapeutic
drug monitoring of clopidogrel.
Thromb Haemost. 2011 Apr;105(4):696-705. Epub 2011 Feb 8.
An improved method for specific and quantitative determination of the
clopidogrel active metabolite isomers in human plasma.
Tuffal G, Roy S, Lavisse M, Brasseur D, Schofield J, Delesque Touchard N, Savi P, Bremond
N, Rouchon MC, Hurbin F, Sultan E.

Source
sanofi-aventis R&D, Drug Disposition, Disposition Safety and Animal Research, Montpellier,
France. gilles.tuffal@sanofi-aventis.com
Abstract
Pharmacokinetic analyses of clopidogrel are hampered by the existence of multiple active
metabolite isomers (H1 to H4) and their instability in blood. We sought to retest the
pharmacodynamic activities of the four individual active metabolite isomers in vitro, with the
ultimate aim of determining the isomers responsible for clopidogrel activity in vivo. In vitro
activity was evaluated by measuring binding of [P]-2-methylthio-ADP on P2Y-expressing
Chinese hamster ovary (CHO) cells and human platelets in platelet-rich plasma (PRP). A
stereoselective method that used reverse-phase ultra high-performance liquid chromatography
(UHPLC) and tandem mass spectrometry (MS) was developed to measure individual
concentrations of the stable 3'-methoxyacetophenone (MP) derivatives of H1-H4. The new
method was used to analyze plasma samples from clopidogrel-treated subjects enrolled in a
phase I clinical trial. In vitro binding assays confirmed the previously observed biological
activity of H4 (IC: CHO-P2Y: 0.12 M; PRP: 0.97 M) and inactivity of H3, and
demonstrated that H1 was also inactive. Furthermore, H2 demonstrated approximately half of
the biological activity in vitro compared with H4. Optimisation of UHPLC conditions and
MS collision parameters allowed the resolution and detection of the four derivatised active
metabolite isomers (MP-H1 to MP-H4). The stereoselective assay was extensively validated,
and was accurate and precise over the concentration range 0.5-250 ng/ml. Only MP-H3 and
MP-H4 were quantifiable in incurred clinical samples. Based on in vitro pharmacodynamic
data and found concentrations, the active metabolite isomer H4 is the only diastereoisomer of
clinical relevance for documenting the pharmacokinetic profile of the active metabolite of
clopidogrel.

Ther Drug Monit. 2008 Feb;30(1):84-9.


Determination of clopidogrel main metabolite in plasma: a useful tool for
monitoring therapy?
Mani H, Toennes SW, Linnemann B, Urbanek DA, Schwonberg J, Kauert GF, Lindhoff-Last
E.
Source
Department of Internal Medicine, Division of Vascular Medicine, University Hospital
Frankfurt/Main, Germany. helenmani@gmx.de
Abstract
This study was performed to determine whether analysis of clopidogrel and its main
carboxylic acid metabolite in plasma provides additional information about the wide
variability of platelet aggregation inhibition in clopidogrel-treated patients with peripheral

arterial occlusive disease. Consecutive outpatients (n = 56) with stable peripheral arterial
occlusive disease treated with 75 mg clopidogrel daily, without co-administration of aspirin,
were investigated. With use of a standardized questionnaire, the time of drug intake was
documented. Blood sampling was performed within 24 hours after the most recent drug
intake. Platelet function was measured by optical aggregometry using adenosine diphosphate
(ADP) (2 mumol/L) as the agonist. Plasma concentrations of clopidogrel and its main
metabolite, clopidogrel carboxylic acid, were quantitated using high-performance liquid
chromatography analysis coupled to mass spectrometry. In 95% (53/56) of patients,
clopidogrel carboxylic acid was detected. In 40% (22/56) of patients, the ADP-induced
aggregation response was within the normal range despite clopidogrel treatment. In 14%
(3/22) of these patients, neither clopidogrel nor its main metabolite could be detected. Two of
these patients agreed to ingest 75 mg/d clopidogrel under observation and to undergo blood
sampling after 2, 12, and 24 hours. Clopidogrel carboxylic acid and a significant inhibition of
platelet aggregation were detected even after 24 hours in both patients, confirming
noncompliance as the reason for the lack of inhibition of ADP-induced platelet aggregation
observed in the initial measurements. In the subgroup of patients who had taken clopidogrel
within 4 hours before blood sampling, a large range of carboxylic acid concentrations was
detected, indicating a high variability of drug metabolism among patients. In conclusion,
determining clopidogrel metabolite plasma concentrations could be a useful tool for
identifying poor compliance and variable metabolism in clopidogrel-treated patients.
Nevertheless, in the majority of clopidogrel-treated patients, the variability of platelet
response is not caused by noncompliance.

Determination of clopidogrel in human plasma


by liquid chromatography/tandem mass
spectrometry: application to a clinical
pharmacokinetic study.
(PMID:17472221)

Abstract
Citations
BioEntities
Related Articles

Shin BS, Yoo SD

College of Pharmacy, Catholic University of Daegu, 330 Geumnak 1-ri, Hayang-eup, Gyeongsan-si,
Gyeongbuk, 712-702, Korea.
Biomedical Chromatography : BMC [2007, 21(9):883-889]
Type: Journal Article, Research Support, Non-U.S. Gov't
DOI: 10.1002/bmc.850
Abstract

Highlight Terms
Species(1)

Chemicals(3)

A rapid and sensitive LC/MS/MS assay was developed and validated for the determination
of clopidogrelin human plasma. Clopidogrel was extracted by single liquid-liquid extraction with pentane, and
chromatographic separations were achieved on a C(18) column. The method was validated to demonstrate the
specificity, linearity, recovery, lower limit of quantification (LLOQ), stability, accuracy and precision. The
multiple reaction monitoring was based on m/z transition of 322.2 --> 211.9 forclopidogrel and 264.1 --> 125.1
for ticlopidine (internal standard). The total analytical run time was relatively short (3 min), and the LLOQ was

10 pg/mL using 0.5 mL of human plasma. The assay was linear over a concentration range from 10 to 10,000
pg/mL (r > 0.999). The intra- and inter-day accuracies were 101.3-108.8 and 98.4-103.5%, respectively, and the
intra- and inter-day assay precisions were 1.9-5.5 and 4.4-8.1%, respectively. The developed assay method was
applied to a pharmacokinetic study in human volunteers after oral administration of clopidogrel at a dose of 150
mg.

Arzneimittelforschung.

2011;61(12):681-4.

Bioequivalence study of two clopidogrel film-coated tablet


formulations in healthy volunteers.
Setiawati E, Yunaidi DA, Handayani LR, Santoso ID, Setiawati A, Tjandrawinata RR.

Source
PT Equilab International, Bioavailability and Bioequivalence Laboratory, Jakarta, Indonesia. effi@equilab-int.com

Abstract
The present study was performed to compare the bioavailability of two clopidogrel 75 mg film-coated tablet formulations (test
formulation and reference formulation). This study was a randomized, single-blind, two-period, two-sequence cross-over study
which included 24 healthy adult male and female subjects under fasting condition. The pharmacokinetic parameters were
assessed based on the concentrations of clopidogrel (CAS 120202-66-6) parent compound. In each of the two study periods
(separated by a washout of one week) a single dose of test or reference drug was administered. Plasma concentrations of the
drug were determined by LC-MS/MS method. The pharmacokinetic parameters assessed in this study were area under the
plasma concentration-time curve from time zero to 24 h (AUCt), area under the plasma concentration-time curve from time zero
to infinity (AUCinf), the peak plasma concentration of the drug (Cmax), time needed to achieve the peak plasma concentration
(t(max)), and the elimination half life (t1/2). The geometric mean ratios (90% CI) of the test drug/reference drug
for clopidogrel parent compound were 95.19% (81.63-110.90%) for AUCt, 95.55% (80.50-113.42%) for AUCinf, and 100.18%
(80.87-124.09%) for Cmax. The 90% confidence intervals calculated for AUCt and Cmax ofclopidogrel parent compound were
within the standard bioequivalence range (80-125% for AUC and Cmax). It was concluded that the two clopidogrelfilm-coated
tablets (test and reference drug) were bioequivalent in terms of the rate and extent of absorption.