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Multiple On-line HPLC Coupled with Biochemical Detection Methods for Screening Effective Components in Ginkgo biloba Extract

??OBJECTIVE To develop on-line high performance liquid chromatography-biochemical detection (HPLC-BCD) methods to screen effective components in Ginkgo biloba extract for treatment of Alzheimer's disease. METHODS Radical scavengers and AchE inhibitors in G.biloba leave extract were screened by HPLC-UV-DPPH/ABTS and HPLC-UV-AchE methods, respectively. The stability and repeatability of the on-line HPLC-UV-AchE method were investigated using galanthamine as a positive reference. The main active ingredients in the extract were identified by ultra high performance liquid chromatography linear ion trap/orbitrap high resolution mass spectrometry (UPLC-LTQ/orbitrap MS) method. RESULTS Fourteen antioxidants in G.biloba extract were detected by the HPLC-UV-DPPH/ABTS method, while three AchE inhibitors were found by the HPLC-UV-AchE method. The three AchE inhibitors were tentatively identified by UPLC-LTQ/orbitrap MS as 4-O-beta-glucopyranosyl-cis-coumaric acid?? ginkgolide A and 3-O-2-O-(β-D-Glucosyl)-α-L-rhamnosyl]quercetin, respectively. CONCLUSION The proposed on-line HPLC-BCD method shows high sensitivity, good repeatability and stability. It can be used to screen antioxidants and AchE inhibitors in G. biloba extract. This will provide an effective, quick and convenient analysis tool to quickly find bioactive ingredients of traditional Chinese medicines for treating related diseases.