Department of Chemistry, School of Sciences, Gujarat University, Ahmedabad 380009, India; Bioanalytical Research Department, Veeda Clinical Research, Ambawadi, Ahmedabad 380015 Gujarat, India.
J Pharm Biomed Anal. 2014 Jan;88:81-91. doi: 10.1016/j.jpba.2013.08.032. Epub 2013 Aug 30.
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.
一种在人血浆中同时测定奥昔布宁、其具有药理活性的代谢物 N-去乙基奥昔布宁及其对映异构体的非对映和对映选择性分析方法，采用 LC-MS/MS 进行。两种方法均使用氘代类似物作为内标进行开发和验证。对映体分析中，外消旋奥昔布宁和 N-去乙基奥昔布宁在等度条件下，于 Phenomenex Gemini C18（150mm×4.6mm，5μm）柱上，以乙腈-5.0mM 乙酸铵，pH4.0（90:10，v/v）为流动相进行分析。对映体分析中，采用 Phenomenex Lux Amylose-2（150mm×4.6mm，3μm）手性柱，以溶剂 A[乙腈：10mM 碳酸氢铵，80:20（v/v）]和溶剂 B[2-丙醇：甲醇，50:50（v/v）]的 20:80（v/v）混合物为流动相进行分离。血浆样品采用乙酸乙酯-二乙醚-正己烷溶剂混合物进行液-液萃取。奥昔布宁和 N-去乙基奥昔布宁对映体的线性范围分别为 0.025-10.0ng/mL 和 0.25-100ng/mL。提取回收率为 96.0-105.1%，而内标归一化基质因子在所有对映体中均为 0.96-1.07。该方法已应用于 5mg 奥昔布宁片剂制剂在 8 名健康受试者中的初步生物等效性研究。药代动力学研究结果表明，（R）-奥昔布宁的血浆浓度低于（S）-奥昔布宁，而 N-去乙基奥昔布宁的对映体则呈现相反的趋势。通过对 20 个剩余样品的重新分析，证明了研究数据测量的重现性。
