title: Characterization of xylazine metabolism in rat liver microsomes using liquid chromatography-hybrid triple quadrupole-linear ion trap-mass spectrometry.
abstract: Xylazine is an α2 -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 μm. 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 (Km = 10.1 μm) 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.
authors: Groupe de Recherche en Pharmacologie Animal du Québec (GREPAQ), Département de biomedicine vétérinaire, Faculté de Médecine Vétérinaire, Université de Montréal, Saint-Hyacinthe, Québec, Canada.
authors: David St-Germain
authors: DS
authors: Lavoie
authors: Floriane
authors: F
authors: Pailleux
authors: Pascal
authors: P
authors: Vachon
authors: Francis
authors: F
authors: Beaudry
mesh_terms: D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods*; D008862:Microsomes, Liver / Q000378:metabolism*; D017711:Nonlinear Dynamics; D051381:Rats; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014991:Xylazine / Q000032:analysis / Q000378:metabolism
chemical_list: D014991:Xylazine