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Abstract
Ginkgo biloba leaves extract (GBE), one of the most widely used traditional Chinese medicines worldwide, can be used for the treatment of diabetes mellitus (DM). However, its biotransformation in liver is not fully known under the state of DM. In this study, an off-line hydrophilic interaction × reversed-phase two-dimensional liquid chromatography (HILIC × RP 2D-LC) system coupled with diode array detection (DAD) and quadrupole time-of-flight mass spectrometry (q/TOF-MS) was established for the qualification and quantification of the biotransformation of GBE in normal and diabetic rat liver microsomes (RLMs). 6 metabolites were tentatively identified according to the exact molecular weights and the characteristic fragment ions provided by q/TOF-MS data. The results of metabolic stability showed that the metabolic ratio of four target compounds including quercetin, genistein, kaempferol and isorhamnetin in diabetic RLMs were significantly enhanced when comparing with normal RLMs. The results of enzyme kinetics showed that compared with normal RLMs, the Michaelis-Menten constant (Km) value of genistein was obvious increased while its maximal velocity (Vmax) and intrinsic clearance (CLint) values were significantly decreased by diabetic RLMs, and the Vmax and CLint values of kaempferol and isorhamnetin were notably enhanced while their Km values were markedly reduced. For the half-time (t1/2) values of four target compounds and the Km, Vmax and CLint values of quercetin, there were not statistically significant changes between normal and diabetic RLMs. The results suggest that the developed off-line 2D LC-DAD-q/TOF-MS method is an easy and accurate approach for the study of GBE biotransformation in RLMs and may provide the essential data for further pharmacological and clinical studies of GBE.
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