Preliminary Investigation Of Phenylpropanoid Glycoside From Ligustrum Purpurasens To Effect Carbohydrate Digestion And Metabolism

With the gradual improvement of people’s standard of living,the global incidence of diabetes is rapidly increasing over the past few decades.According to the statistics from the IDF,an estimated 425 million people had diabetes in 2017.China has the largest diabetic population in the world.Compared to traditional anti-diabetic medication,phytochemicals have the benefits of high efficacy,low toxicity,and minimal side effects.As a result,phytohemicals have great potential in treating symptoms related to diabetes.Ligustrum purpurascens is an edible medicinal plant species commonly found in southwestern China.The leaves used in a drink,called small-leaved kudingtea（bitter tea）comes to a bright prospect today.In this article,the main active ingredient of small-leaved kudingtea,phenylpropanoid glucosides（PPGs）,and its main active monomer acteoside were selected as the main objects of the study andα-amylase andα-glucosidase inhibition assay,fluorescence analysis,circular dichroism（CD）,Fourier transform infrared spectroscopy and docking studies were performed to explore the inhibition properties of PPGs at enzymatic level.The inhibition activity onα-amylase andα-glucosidase was examined by high performance liquid chromatography（HPLC）and liquid chromatography-mass spectrometry（LC/MS）with the products obtained from the heat treatment of acteoside and the inhibition effect before and after heating treatment was compared.In order to provide theoretical basis for the development of small-leaved kudingtea resources,we studied acteoside at cellular level as well.L6myc muscle cells were used as a model to study how acteoside affected the glucose uptake function.The related protein expressions in acteoside-stimulated-insulin receptor signaling pathway was detected and the suppressive hyperglycemic effect was evaluated.The main research results are as follows:1.Two critical exoacting glycoside hydrolase enzymes involved in carbohydrate digestion areα-glucosidase andα-amylase.We investigated the properties of PPGs from L.purpurascens for inhibitingα-amylase andα-glucosidase activity in vitro and found IC₅₀values of 1.02 and 0.73 mg/mL,respectively.The patterns of inhibiting bothα-amylase andα-glucosidase were mixed-inhibition type.Multispectroscopy and molecular docking studies indicated that the interaction between PPGs andα-amylase andα-glucosidase altered the conformation of enzymes,with binding at the site close to the active site of enzymes resulting in changed enzyme activity.2.Acteoside acyl migration to isoacteoside with high-temperature.The inhibitory effects of acyl-migrated acteoside and acteoside onα-amylase were investigated,as were their binding interaction withα-amylase.Acteoside was more effective than preheated acteoside and isoacteoside in inhibitingα-amylase activity.Acteoside and isoacteoside binding toα-amylase may induce conformational changes toα-amylase,and the binding site of acteoside and isoacteoside being near the active site pocket ofα-amylase may explain the decreased activity ofα-amylase.The different affinities and binding sites of acteoside and isoacteoside for a-amylase resulted in different inhibition rates,which may be due to structural differences between acteoside and isoacteoside.3.Acteoside could increase the activity of AMPK and the phosphorylation of AS160（TCB1D4）,did not show any toxicity under this condition.The results showed that acteoside might promote glucose uptake via AMPK signaling pathway.Acteoside could not ameliorate insulin-resistance.The molecular mechanism of the glucose-lowering effect of acteoside still needs further research.