A LC-ESI-MS method for the simultaneous determination of madecassoside and its metabolite madecassic acid in rat plasma:comparison pharmacokinetics in normal and collagen-induced arthritic rats

To develop a simple and highly sensitive high performance liquid chromatography with electrospray ionization mass spectrometric(LC-ESI-MS) method for the simultaneous determination of madecassoside and its major metabolite madecassic acid in rat plasma,and compare the pharmacokinetics of the two compounds in normal and collagen-induced arthritis(CIA) rats.Glycyrrhetinic acid was used as the internal standard(IS).Chromatographic separation was accomplished on an Inertsil ODS-3 column,using a gradient elution with the mobile phase composed of acetonitrile and water acidified with 0.1%(V/V) formic acid.Detection was achieved by ESI-MS under the negative selected ion monitoring(SIM) mode.In normal and CIA rats,madecassoside(30 mgkg-1) was orally administered for 21 consecutive days from the day of arthritis onset.For madecassoside,the linear range was 10-1 000 ngmL-1 with the square regression coefficient(r) of 0.998 9,while for madecassic acid,the linear range was 10-500 ngmL-1 with the square regression coefficient(r) of 0.996 1.The lower limit of quantification was 10 ngmL-1 for both analytes.The intra-and inter-day precision ranged from 1.78% to 13.42% for madecassoside and 2.30% to 14.90% for madecassic acid,and the accuracy was between-0.95% and 6.30% for madecassoside and between-1.48% and 5.34% for madecassic acid.The average recoveries of madecassoside,madecassic acid and IS from spiked plasma samples were81%.The developed method was successfully applied to the pharmacokinetic study of madecassoside and madecassic acid in rats after an oral administration of madecassoside.During initial 7 days of dosing,the cmax and AUC of madecassoside were greatly decreased and Vd/F was markedly increased in CIA rats,and no significant difference was observed on the first day of dosing.In contrast,the T1/2,cmax and AUC of madecassic acid were significantly increased,and Ke of madecassic acid was greatly decreased in CIA rats compared with normal rats.Along with repeated administration of madecassoside,the differences of pharmacokinetic parameters of both madecassoside and madecassic acid between CIA and normal rats gradually subsided.The pharmacokinetic characteristics of both madecassoside and madecassic acid in rats were significantly altered by arthritis status,and the differences of pharmacokinetic parameters between arthritis and normal rats coincide with the severity of arthritis.