Multi-Compartment Electrophoresis Separation Systen Based Method For Screening Bioactive Compounds From Traditional Chinese Medicines

Based on the laboratory’s early development of multi-compartment electrophoresis system, we present herein a method for rapid screening potential a-glucosidase inhibitors from traditional Chinese medicine (TCMs). We chose α-glucosidase as the target enzyme in the experiment, the TCMs extract solution was incubated a-glucosidase for an appropriate period firstly, making active compounds bind to a-glucosidase and form α-glucosidase-ligands complex; then we loaded the incubation mixture in the sample compartment, and applied a certain voltage, the enzyme-ligand complexes migrated to the acceptor compartment and compounds of no affinity remained immobile in the sample compartment; at last, the specific bound compounds were dissociated from complex using organic reagents and analyzed by HPLC-MS/MS. The migration percentage of α-glucosidase was an important factor influencing screening results, so the factors influencing migration percentage of a-glucosidase including buffer solution, m embrane properties, a pplied voltage and electrophoresis time were optimized. The optimal conditions were phosphate buffered solution (67 mM, pH 7.4), cellulose acetate membrane (0.2 μm) between sample compartment and acceptor compartment, polyethersulfone ultrafiltration membrane (10 KD) between acceptor compartment and electrode compartment,8 V of applied voltage,15 min of electrophoresis time. Then the developed approach was first validated using a known a-glucosidase inhibitor emodin, and then emodin, tanshinone I, tanshinone IIA and cryptotanshinone standards were used to further evaluate the method performance on studying synergistic effect of ligands. The results showed that the binding capacity of the compounds incubated with a-glucosidase together differed from incubated with a-glucosidase individually, we did enzyme inhibitory assay further, the enzyme inhibiton rate of compounds added together according to screened relative molar ratio was higher than individually, which indicated the developed approach can meet the requirements of screening synergistic ligands to enzyme. Afterwards, the established approach was used for screening potential a-glucosidase inhibitors from schizandra chinensis, and schisandrol A, gomisin D, schisandrol B, Tigloylgomisin H, Angeloylgomisin H, Angeloylgomisin Q and Tigloylgomisin P were screened. Our findings suggested that the proposed approach is efficient in screening bioactive compounds of affinity to target protein from extracts of TCMs. It provides a new way for rapidly screening bioactive compounds from traditional Chinese medicines. Moreover, the proposed method has a broad application in screening drug molecules having synergistic effect.