Quantification of metronidazole in healthy subjects' feces using an LC–MS/MS method

• Published in: Biomedical chromatography Vol. 32; no. 9; pp. e4265 - n/a
• Main Authors: Vanol, Pravin G., Sanyal, Mallika, Shah, Priyanka A., Shrivastav, Pranav S.
• Format: Journal Article
• Language: English
• Published: England 01.09.2018
• Subjects: healthy subjects; human feces; LC–MS/MS; metronidazole; metronidazole‐d4; sensitive; LC-MS/MS; metronidazole-d4; metronidazole; sensitive; human feces; healthy subjects
• ISSN: 0269-3879; 1099-0801; 1099-0801
• DOI: 10.1002/bmc.4265

A highly selective and sensitive liquid chromatography–tandem mass spectrometry (LC MS/MS) method was developed for the quantification of metronidazole (MTZ) in human feces. The analyte was recovered from feces after liquid–liquid extraction with ethyl acetate and separated on Waters Symmetry® C18 (100 × 4.6 mm, 5μm) column using 0.1% formic acid in water and acetonitrile (40:60, v/v) as the mobile phase. A stable‐deuterated internal standard metronidazole‐d4 (MTZ‐d4) was used in the study. Mass analysis was performed on a triple quadrupole mass spectrometer in the positive electrospray ionization mode. A linear response function of MTZ was established in the concentration range of 0.50–250 ng/g, based on dry mass. The mean extraction recovery of MTZ (97.28%) and MTZ‐d4 (96.76%) from spiked feces samples was consistent at higher as well as lower concentrations. Post‐column infusion analysis showed no ion‐suppression/enhancement effects and the mean IS‐normalized matrix factor ranged from 0.986 to 1.013. Spiked feces samples stored at −20 and − 70°C for long‐term stability were stable for at least 3 months, while extracted samples (dry and wet extracts) were stable up to 24 h. The method was applied to determine MTZ in feces of 12 healthy Indian subjects.