TY - JOUR
T1 - Validation of a method for quantitation of the clopidogrel active metabolite, clopidogrel, clopidogrel carboxylic acid, and 2-oxo-clopidogrel in feline plasma
AU - Lyngby, Janne G
AU - Court, Michael H
AU - Lee, Pamela M
N1 - Copyright © 2017 Elsevier B.V. All rights reserved.
PY - 2017/8
Y1 - 2017/8
N2 - INTRODUCTION: The clopidogrel active metabolite (CAM) is unstable and challenging to quantitate. The objective was to validate a new method for stabilization and quantitation of CAM, clopidogrel, and the inactive metabolites clopidogrel carboxylic acid and 2-oxo-clopiodgrel in feline plasma.ANIMALS: Two healthy cats administered clopidogrel to demonstrate assay in vivo utility.MATERIALS AND METHODS: Stabilization of CAM was achieved by adding 2-bromo-3'methoxyacetophenone to blood tubes to form a derivatized CAM (CAM-D). Method validation included evaluation of calibration curve linearity, accuracy, and precision; within and between assay precision and accuracy; and compound stability using spiked blank feline plasma. Analytes were measured by high performance liquid chromatography with tandem mass spectrometry. In vivo utility was demonstrated by a pharmacokinetic study of cats given a single oral dose of 18.75mg clopidogrel.RESULTS: The 2-oxo-clopidogrel metabolite was unstable. Clopidogrel, CAM-D, and clopidogrel carboxylic acid appear stable for 1 week at room temperature and 9 months at -80°C. Standard curves showed linearity for CAM-D, clopidogrel, and clopidogrel carboxylic acid (r > 0.99). Between assay accuracy and precision was ≤2.6% and ≤7.1% for CAM-D and ≤17.9% and ≤11.3% for clopidogrel and clopidogrel carboxylic acid. Within assay precision for all three compounds was ≤7%. All three compounds were detected in plasma from healthy cats receiving clopidogrel.DISCUSSION: This methodology is accurate and precise for simultaneous quantitation of CAM-D, clopidogrel, and clopidogrel carboxylic acid in feline plasma but not 2-oxo-clopidogrel.CONCLUSIONS: Validation of this assay is the first step to more fully understanding the use of clopidogrel in cats.
AB - INTRODUCTION: The clopidogrel active metabolite (CAM) is unstable and challenging to quantitate. The objective was to validate a new method for stabilization and quantitation of CAM, clopidogrel, and the inactive metabolites clopidogrel carboxylic acid and 2-oxo-clopiodgrel in feline plasma.ANIMALS: Two healthy cats administered clopidogrel to demonstrate assay in vivo utility.MATERIALS AND METHODS: Stabilization of CAM was achieved by adding 2-bromo-3'methoxyacetophenone to blood tubes to form a derivatized CAM (CAM-D). Method validation included evaluation of calibration curve linearity, accuracy, and precision; within and between assay precision and accuracy; and compound stability using spiked blank feline plasma. Analytes were measured by high performance liquid chromatography with tandem mass spectrometry. In vivo utility was demonstrated by a pharmacokinetic study of cats given a single oral dose of 18.75mg clopidogrel.RESULTS: The 2-oxo-clopidogrel metabolite was unstable. Clopidogrel, CAM-D, and clopidogrel carboxylic acid appear stable for 1 week at room temperature and 9 months at -80°C. Standard curves showed linearity for CAM-D, clopidogrel, and clopidogrel carboxylic acid (r > 0.99). Between assay accuracy and precision was ≤2.6% and ≤7.1% for CAM-D and ≤17.9% and ≤11.3% for clopidogrel and clopidogrel carboxylic acid. Within assay precision for all three compounds was ≤7%. All three compounds were detected in plasma from healthy cats receiving clopidogrel.DISCUSSION: This methodology is accurate and precise for simultaneous quantitation of CAM-D, clopidogrel, and clopidogrel carboxylic acid in feline plasma but not 2-oxo-clopidogrel.CONCLUSIONS: Validation of this assay is the first step to more fully understanding the use of clopidogrel in cats.
KW - Animals
KW - Cats/blood
KW - Chromatography, High Pressure Liquid/methods
KW - Reproducibility of Results
KW - Sensitivity and Specificity
KW - Tandem Mass Spectrometry/methods
KW - Ticlopidine/administration & dosage
U2 - 10.1016/j.jvc.2017.03.004
DO - 10.1016/j.jvc.2017.03.004
M3 - Journal article
C2 - 28602635
SN - 1760-2734
VL - 19
SP - 384
EP - 395
JO - Journal of Veterinary Cardiology
JF - Journal of Veterinary Cardiology
IS - 4
ER -