In-vivo and In-vitro Metabolism Study of Timosaponin B-II Using HPLC-ESI-MS n

Timosaponin B-II (TB-II), a representative furostanol saponin in Rhizoma anemarrhenae , has been used as an emperor herb in many Chinese herbal formulas to treat diabetes and senile dementia. However, its metabolism and tissue distribution had not been investigated so far. In this work, a sensitive and specific high-performance liquid chromatography-electrospray ionization tandem mass spectrometry method was applied for the identification of TB-II and its major metabolites in in-vivo and in-vitro samples. Rat urine, feces, plasma and tissues were collected after oral administration of TB-II at a single dose of 300 mg kg −1 . Furthermore, TB-II was incubated in artificial gastric juice (AGJ) and artificial intestinal juice (AIJ). As a result, 19 metabolites were detected and identified by comparing their HPLC behavior and MS n spectra profile with those of the parent drug. Moreover, the structures of its five metabolites were identified by using the standards prepared by the acid hydrolysis of TB-II. In addition to the parent drug, 14, 12, 6, 1, 1 and 7 metabolites were detected in rat urine, feces, plasma, heart, kidney and liver, respectively, while no metabolites or the parent drug were found in rat brain, spleen and lung. Seven metabolites appeared in AIJ incubation samples, but the parent drug was absent. Nine metabolites along with the parent drug were observed in AGJ incubation samples. The biotransformation pathways of TB-II mainly included dehydration, deglycosylation, hydroxylation, oxidation and E-ring cleavage. This is the first comprehensive investigation of the in-vivo and in-vitro metabolism of TB-II. The result provided important information for further pharmacological research on TB-II.