Characterization of in vitro metabolism of capsazepine,a vanilloid transient receptor potential channel antagonist,by liquid chromatography quadrupole ion trap mass spectrometry

Capsazepine is an antagonist of the transient receptor potential channel vanilloid 1 (TRPV1), which is known to play an important role in the regulation of pain and inflammation. A selective and sensitive quantitative method for the determination of capsazepine by HPLC‐ESI/MS/MS was developed. The method consisted of a protein precipitation extraction followed by analysis using liquid chromatography electrospray quadrupole ion trap mass spectrometry. The chromatographic separation was achieved using a 100 × 2 mm C₁₈ Waters Symmetry column combined with a gradient mobile phase composed of acetonitrile and 0.1% formic acid aqueous solution at a flow rate of 220 µL/min. The mass spectrometer was operating in full‐scan MS/MS mode using two‐segment analysis. An analytical range of 10–5000 ng/mL was used in the calibration curve constructed in rat plasma. The inter‐batch precision and accuracy observed were 10.1, 6.4 and 6.1% and 100.8, 98.5 and 106.2% at 50, 500 and 5 000 ng/mL, respectively. An in vitro metabolic stability using rat, dog or mouse liver microsomes was performed to determine the intrinsic clearance of capsazepine. The results suggest a very rapid degradation with T_1/2 ranging from 2.6 to 4.3 min and a high clearance, suggesting that drug bioavailability is considerably reduced following extravascular administrations, consequently affecting drug response. Three metabolites were identified by HPLC‐MS/MS. S‐hydroxylation (M + 16), oxidative desulfuration (M − 16) and desulfuration (M − 32) metabolites of capsazepine were observed following exposure to rat, dog and mouse microsomes. Copyright © 2010 John Wiley & Sons, Ltd.