title: Parallel achiral-chiral determination of oxybutynin, N-desethyl oxybutynin and their enantiomers in human plasma by LC-MS/MS to support a bioequivalence trial.
abstract: A parallel achiral and chiral determination of oxybutynin, its pharmacologically active metabolite N-desethyl oxybutynin and their enantiomers in human plasma is described using LC-MS/MS. Both the methods were developed and validated using deuterated analogues as internal standards. Achiral analysis of racemic oxybutynin and N-desethyl oxybutynin was carried out on Phenomenex Gemini C18 (150mm×4.6mm, 5μm) column under isocratic conditions using acetonitrile-5.0mM ammonium acetate, pH 4.0 (90:10, v/v) as the mobile phase. Separation of (S)- and (R)-enantiomers of the analytes was performed on Phenomenex Lux Amylose-2 (150mm×4.6mm, 3μm) chiral column using a mixture of solvent A [acetonitrile:10mM ammonium bicarbonate, 80:20 (v/v)] and solvent B [2-propanol:methanol, 50:50 (v/v)] in 20:80 (v/v) ratio as the mobile phase. Plasma samples were prepared by liquid-liquid extraction with ethyl acetate-diethyl ether-n-hexane solvent mixture. A linear range was established from 0.025 to 10.0ng/mL and 0.25 to 100ng/mL for the enantiomers of oxybutynin and N-desethyl oxybutynin respectively. The extraction recovery varied from 96.0 to 105.1%, while the IS-normalized matrix factors ranged from 0.96 to 1.07 for all the enantiomers. The validated method was applied for a pilot bioequivalence study with 5mg oxybutynin tablet formulation in 8 healthy subjects. The pharmacokinetic profiles showed that the plasma concentration of (R)-oxybutynin was lower than that of (S)-oxybutynin, while a reverse trend was observed for the enantiomers of N-desethyl oxybutynin. The reproducibility in the measurement of study data was demonstrated by reanalysis of 20 incurred samples.
authors.affiliation: Department of Chemistry, School of Sciences, Gujarat University, Ahmedabad 380009, India; Bioanalytical Research Department, Veeda Clinical Research, Ambawadi, Ahmedabad 380015 Gujarat, India.
authors.forename: Primal
authors.initials: P
authors.lastname: Sharma
authors.forename: Daxesh P
authors.initials: DP
authors.lastname: Patel
authors.forename: Mallika
authors.initials: M
authors.lastname: Sanyal
authors.forename: Swati
authors.initials: S
authors.lastname: Guttikar
authors.forename: Pranav S
authors.initials: PS
authors.lastname: Shrivastav
mesh_terms: D001774:Blood Chemical Analysis; D002138:Calibration; D002845:Chromatography; D002853:Chromatography, Liquid; D004355:Drug Stability; D064368:Healthy Volunteers; D006801:Humans; D007194:India; D059625:Liquid-Liquid Extraction; D008333:Mandelic Acids / Q000097:blood*; D010865:Pilot Projects; D011786:Quality Control; D015203:Reproducibility of Results; D012997:Solvents; D013237:Stereoisomerism; D013607:Tablets; D053719:Tandem Mass Spectrometry; D013810:Therapeutic Equivalency
chemical_list: D008333:Mandelic Acids; D012997:Solvents; D013607:Tablets; C032681:desethyloxybutynin; C005419:oxybutynin
keywords: (S)- and (R)-N-Desethyl oxybutynin; (S)- and (R)-Oxybutynin; Achiral; Chiral; LC–MS/MS