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Characterization of xylazine metabolism in rat liver microsomes using liquid chromatography–hybrid triple quadrupole–linear ion trap–mass spectrometry
Author(s) -
Lavoie David StGermain,
Pailleux Floriane,
Vachon Pascal,
Beaudry Francis
Publication year - 2013
Publication title -
biomedical chromatography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.4
H-Index - 65
eISSN - 1099-0801
pISSN - 0269-3879
DOI - 10.1002/bmc.2875
Subject(s) - chemistry , chromatography , xylazine , selected reaction monitoring , metabolite , mass spectrometry , microsome , electrospray ionization , formic acid , tandem mass spectrometry , ion trap , high performance liquid chromatography , biochemistry , ketamine , in vitro , medicine , anesthesia
Xylazine is an α 2 ‐adrenoceptor agonist and it is widely used in veterinary anesthesia in combination with ketamine. There is limited information on the metabolism of xylazine. A quantitative method for the determination of xylazine by HPLC‐ESI/MS/MS was developed. The method consisted of a protein precipitation extraction followed by analysis using liquid chromatography electrospray tandem mass spectrometry. The chromatographic separation was achieved using a Thermo Betasil Phenyl 100 × 2 mm column combined with an isocratic mobile phase composed of acetonitrile, methanol, water and formic acid (60:20:20:0.4) at a flow rate of 300 μL/min. The mass spectrometer was operating in selected reaction monitoring mode and the analytical range was set at 0.05–50 μ m . The precision (%CV) and accuracy (%NOM) observed were 2.3–7.2 and 88.2–96.4%. In vitro metabolism studies were performed in rat liver microsomes and results showed moderate cytochrome P450 affinity ( K m = 10.1 μ m ) and a low metabolic stability of xylazine with a half‐life of 4.1 min in rat liver microsomes. Five phase 1 metabolites were observed. The main metabolite observed was an oxidation of the thiazine moiety at m / z 235 and, to a lesser extent, we observed the formation of N ‐(2,6‐dimethylphenyl)thiourea at m / z 181 and three distinctive hydroxylated metabolites at m / z 237. Further experiments with ketamine and ketoconazole strongly supported that the metabolism of xylazine to its main metabolite is mediated by CYP3A in rat liver microsomes. Copyright © 2013 John Wiley & Sons, Ltd.