Enantioselective determination of ornidazole in human plasma by liquid chromatography-tandem mass spectrometry on a Chiral-AGP column

A rapid, sensitive, and enantioselective method was developed and validated for determination of ornidazole enantiomers in human plasma by liquid chromatography-tandem mass spectrometry. Ornidazole enantiomers were extracted from 100 mu l of plasma using ethyl acetate. Baseline chiral separation (R-s = 2.0) was obtained within 7.5 min on a Chiral-AGP column (150 mm x 4.0 mm, 5 mu m) using an isocratic mobile phase of 10 mM ammonium acetate/acetic acid (100/0.01, v/v). Stable isotopically labeled R-(+)-d(5)-ornidazole and S-(-)-d(5)-ornidazole were synthesized as internal standards. Acquisition of mass spectrometric data was performed in multiple reaction monitoring mode via positive electrospray ionization, using the transitions of m/z 220 -> 128 for ornidazole enantiomers, and m/z 225 -> 128 for d(5)-ornidazole enantiomers. The method was linear in the concentration range of 0.030-10.0 mu g/ml for each enantiomer. The lower limit of quantification for each enantiomer was 0.030 mu g/ml. The relative standard deviation values of intra- and inter-day precision were 1.8-6.2% and 1.5-10.2% for R-(+)-ornidazole and S-(-)-ornidazole, respectively. The relative error values of accuracy ranged from -4.5% to 1.2% for R-(+)-ornidazole and from -5.4% to -0.8% for S-(-)-ornidazole. The validated method was successfully applied to a stereoselective pharmacokinetic study of ornidazole after oral administration of 1000 mg racemic ornidazole. (C) 2013 Elsevier B.V. All rights reserved.