Determination of Methyl p-Hydroxy-benzoate, the Metabolite of the Candidate Vaginal Contraceptive Drug 4߰-Carbomethoxyphenyl 4-Guanidinobenzoate Mesylate, in Rabbit Plasma: Application of a rapid and sensitive liquid chromatography-tandem mass spectrometry method

Abstract

A rapid, sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the determination of methyl p-hydroxybenzoate (CAS 99-76-3), the metabolite of the candidate vaginal contraceptive drug 4′-carbomethoxyphenyl 4-guanidinobenzoate mesylate (4′-CMGB) in rabbit plasma. The samples were prepared by liquid-liquid extraction with ethyl acetate after spiking with the internal standard, ethyl p-hydroxybenzoate (PHBA-Et). Chromatographic separation was achieved on a C18 column (4.6 mm × 150 mm, 5 µm) with a mobile phase composition of methanol-water (55:45, v/v) delivered at 1.0 ml/min. LC-MS/MS analysis was carried out on a LC-triple-quadrupole mass spectrometer using atmospheric pressure chemical ionization (APCI) and negative multiple reaction monitoring (MRM). Ions monitored were m/z 151 → 92 for PHBA-Me and m/z 165 → 92 for PHBA-Et with argon at a pressure of 0.8 Pa and collision energy of 32 eV for collision-induced dissociation (CID). The assay was linear over the range of 2.0–100 ng/ml for PHBA-Me. The average recovery was 103.9%. The established method has been applied in a pharmacokinetic study of 4′-CMGB gel in rabbit.