Inhibitory Activity Of Triterpenoids From Inonotus Obliauus On α-glucosidase And α-amylase And Separation And Identification Of Effective Compounds

In recent years,diabetes has been damaged human’s health due to its high incidence,however,the clinical medication is single and has side effects.It is urgent to discover new antidiabetic drugs likeα-glucosidase/α-amylase from natural products.Inonotus obliquus has been used as a folk medicine to treat diabetes,cardiovascular diseases and cancer in Russia,China and other countries for a long time.However,due to its rare natural resources,the use of artificial techniques to expand cultivation has become a new trend.In this paper,the effects of wild sclerotia and liquid fermentation triterpenoids inhibitingα-glucosidase andα-amylase were compared and studied,and the effectiveness of using liquid submerged fermentation to cultivate active triterpenes was verified.In this paper,triterpenoids from sclerotia and fermented mycelia were chromatographed and identified,and the effective components were initially determined.The main results of this research are summarized as follows:1.Three kinds of triterpenoid monomers were isolated from the sclerotia of I.obliquus,and they were identified as lanosterol,inotodiol and trametenolic acid by HPLC and NMR analysis.Three kinds of triterpenoids were isolated from the liquid fermentation mycelium,and were identified as inoterpene D,lawsaritol,and betulinic acid by HPLC/MS analysis.Inotodiol had the strongest inhibitory activity onα-glucosidase,and inoterpene D had the strongest inhibitory activity onα-amylase,and the effects are better than acarbose.2.Under the guidance of biological activity,threefractions were isolated from the liquid fermentation I.obliquus.IC₅₀ of fraction 4 inhibitingα-glucosidase was 1.33mg/m L,and ofα-amylase was 7.86 mg/m L;IC₅₀of fraction 7 inhibitingα-glucosidase was 1.52 mg/m L,and ofα-amylase was 15.59 mg/m L;IC₅₀ of fraction 8 inhibitingα-glucosidase was 0.26 mg/m L,and ofα-amylase was 8.88 mg/m L.The main substances of fraction 4,7 and 8 were inoterpene D,lawsaritol,and betulinic acid,respectively.3.Acarbose as the positive control inhibitedα-glucosidase with an IC₅₀ of 0.37mg/m L,and the inhibition type of effect was competitive.The IC₅₀ of lanosterol,inotodiol and trametenolic acid was 0.60 mg/m L,0.15 mg/m L,and 0.187 mg/m L,and their inhibition type of action was competitive.The IC₅₀ of acarbose inhibitingα-amylase is 10.51 mg/m L,the inhibition type was a mixed of non-competitive and competitive,the IC₅₀ of lanosterol,inotodiol and trametenolic acid was 9.51 mg/m L,14.31 mg/m L and more than 100 mg/m L,and their inhibition type was competitive.4.Two-way fermentation of medicinal fungi and Chinese herbal medicine is a new type development technology of medicinal material.Adding Eucommia ulmoides to the basic fermentation medium had no effect on the growth and total triterpenoids production.The addition of water extracts from Eucommia ulmoides bark and leaves increased the biomass of I.obliquus by 4.30%and 26.82%,respectively.The former increased triterpenoids total production by 14.44%while the later decreased by45.80%.In short,the triterpenoids in I.obliquus played an important role in lowering blood glucose by inhibitingα-glucosidase andα-amylase,providing a theoretical basis for the development of blood sugar lowering drugs.The composition of sclerotia and liquid fermentation of triterpenoids is different,and the active triterpenoids,inoterpene D,lawsaritol,and betulinic acid produced by liquid fermentation culture were discovered for the first time.In addition,the liquid culture conditions have obvious effects on the biomass and triterpenoids accumulation of I.obliquus.