Determination Of Risperidone And Tiopronin In Human Plasma By LC-MS/MS And Its Application

There are a large number of samples during bioequivalence study, the matrix ofwhich is very complicated. LC-MS/MS is very suitable for the determination of drugconcentration in plasma owning to its high selectivity and good sensitivity. It can alsodepress the interference from endogenous substances and greatly shorten the run timeof samples.In the first part, we established an LC-MS/MS method for the determination ofrisperidone in human plasma with diazepam as the internal standard, which wassuccessfully applied to the bioequivalence in30healthy volunteers. Positive ionelectrospray ionization with multiple reaction-monitoring mode （MRM） wasemployed by monitoring the transitions m/z411.2/191.0and m/z285.0/154.0forrisperidone and for the IS. Risperidone was extracted by liquid-liquid extraction andseparated on a Ultimate XB-C18（5m,2.1mm150mm） using methanol and5mmol/L ammonium acetate（80:20, v/v）. The calibration curve was linear over therange of0.100²4.150ng/mL and the LLOQ was0.100ng/mL. The90%confidenceintervals were82.5%～102.9%,81.5%～102.2%,77.1%～93.7%for AUC_0-24, AUCi,Cmax, respectively. T_max was estimated by Wilcoxon test and the results indicatedthat there was no significant difference between the test and reference preparations.An LC-MS/MS method was constructed to determine the concentration oftiopronin in human plasma using DL-Thiorphan as the internal standard andsuccessfully applied to a bioequivalence in22healthy volunteers in the second part.Under negative ion electrospray ionization, the transition ion pairs were m/z：161.9/105.0and m/z：251.8/217.9for tiopronin and the IS, respectively. L-Cysteineand1,4-dithio threitol were added as the reducer and stabilitizer before the plasmasamples were precipitated with acetonitrile. The analytes were separated on aUltimate XB-CN （5m,2.1mm50mm） with the mobile phase constituted ofacetonitrile and water including0.1%formic acid （60:40, v/v）. The run time of eachsample was3.0min and the inject volume was5μL. The LLOQ of the method is3.888ng/mL, which can reach our requirements. The pharmacokinetic parameters oftiopronin were calculated by DAS software3.0. The bioequivalence was determinedby the basis of AUC_0-96, AUCi, Cmax ANOVA and group equivalence analysis after logarithmic transformation, the results of two one-sided t-test and [1-2α]%confidenceinterval were also calculated and evaluated. The90%confidence intervals for AUC_0-96,AUCi, Cmax were97.9%～118.7%,97.1%～117.7%,95.2%～116.9%, respectively.The results revealed that the two pharmaceutical preparations were bioequivalent inboth the absorption rate and extent of absorption.