Isolation Of α-glucosidase And PTP-1B Targeted Bioactive Compounds From Laminaria Japonica Root And Identification Of Their Anti-diabetic Effects

According to the fact that kelp root has been a kind of civilian anti-diabetic drug for long, bioactive compounds are expected to be isolated from Laminaria japonica root, and their molecular targets to be identified in Diabetes MellitusⅡ. The crude extract was discovered to dose-dependently inhibit bothα-glucosidase (IC50 1589ug/ml) and PTP-1B(IC50 1271ug/ml), either of which plays important role in DMⅡ。EtOAc part and P.Ether part have inhibiting activity ofα-glucosidase and PTP-1B respectively. Methodology and various techniques of natural product chemistry were subsequently employed to purify and characterize the anti-diabetic components. The purifying process was strictly guided byα-glucosidase and/or PTP-1B inhibiting bioactivity. Sinceα-glucosidase inhibitor was detected in EtOAc part, the bioactive compound was supposed to be purified through silica column chromatography eluted by P. ether: Actone and Dichloromethane: Methanol, and then through Sephadex LH20 filtration chromatography eluted by Dichloromethane: Methanol. Immediately after each step of chromatography enzymatic assays would locate the bioactive part and the IC50 of each step would also be calculated. HPLC was applied to further purification. Chemical structure ofα-glucosidase inhibitor, with IC50 of 3.6ug/ml, was identified by MS and NMR.Bioactivity assay demonstrates thatα-glucosidase inhibitor and PTP-1B inhibitor are different compounds, located themselves in EtOAc part(IC50 380ug/ml) and P.Ether part(IC50 220ug/ml) respectively.α-glucosidase inhibitor in kelp root is light sensitive and high-temperature sensitive. Treatment of 48h in light or 50℃12h results in remarkable reduction of bioactivity. TLC analysis, FeCl3 as chromogenic reagent, shows the labile bioactive compound seems to be polyphenol. Treatment to STZ induced diabetic mice shows that P.Ether and EtOAc part together, has remarkable anti-hyperglycemic activity, at the dose of 1450ug/kg, statistically different with standard group. In conclusion, kelp root has anti-diabetic activity both in vitro and in vivo, and could be developed as a potential new drug.