Novel Flavoalkaloids From Bai-Mudan Tea With Inhibitory Activity Against Formation Of Advanced Glycation End Products

Tea is one of the three most popular non-alcoholic beverages in the world.In recent years,with the spread of the concept of healthy life,tea with many health functions has been increasing trade volume in the international and domestic market.White tea is one of the six traditional Chinese tea categories,which retains the original flavor of tea,mellow taste,fresh aroma,because of the unique processing technology.White tea can be ranked into four classes according to its quality as the following sequence:Silver Needle（or Baihao-Yinzhen）,White Peony（or Bai-Mudan）,Tribute Eyebrow（or Gong-Mei）,and Longevity Eyebrow（Shou-Mei）.Among them,Bai-Mudan is a popular one with graceful form of green leaf white core.In the folk"white tea one year tea,three years medicine,seven years treasure"is widely circulated.Recent studies have indicated that white tea shows effects on anti-oxidant,anti-obesity,anti-cancer,and so on.The pharmacological functions are closely related to the chemical compositions of tea.The flavoalkaloids is a kind of catechins derivatives with a catechin skeleton.It is difficult to purify and identify flavoalkaloids.Therefore,few researches have been conducted on this kind of compounds.The determination of the absolute configuration of flavoalkaloids from Bai-Mudan tea and their bioactivity provide a theoretical basis for potentiall development of new drugs.The main content of this experiment is on flavoalkaloids in white tea.Different solvents（dichloromethane,ethyl acetate,n-butyl alcohol）were used for the extraction of80%acetone water leaching liquid to obtain n-butanol soluble fraction.Sephadex LH-20column chromatography（CC）,MCI-Gel CHP20P gel CC,Toyopearl CC,silica gel CC,ODS CC,and HPLC were used to get monomeric compounds.¹H-NMR,¹³C-NMR,2D-NMR,CD and IR were used to determine the structure of the compounds.The activities of flavoalkaloids were explored by molecular docking technique and AGEs inhibition assay.In this experiment,15 compounds were isolated from Bai-Mudan tea,including two novel flavoalkaloids（-）-6-（5’’’S）-N-ethyl-2-pyrrolidinone-epigallocatechin-3-O-gallate（ester-type catechins pyrrolidinone A,etc-pyrrolidinone A,1）,（-）-6-（5’’’R）-N-ethyl-2-pyrrolidinone-epigallocatechin-3--O-gallate（etc-pyrrolidinone B,2）and 13 known compounds including（-）-8-（5’’’S）-N-ethyl-2-pyrrolidinone-epigallocatechin-3--O-gallate（3a*）and（-）-8-（5’’’S）-N-ethyl-2-pyrrolidinone-epigallocatechin-3--O-gallate（3b*）,gallicin（4）,（+）-catechin（5）,（-）-epicatechin（6）,gallic acid（7）,kaempferol（8）,procyanidin B2（9）,kaempferol-3-7-O-β-D-dirhamnoside（10）,epigallocatechin-（4β-8）-epicatechin-3-O-gallate（11）,（-）-epigallocatechin-O-gallate（EGCG,12）,1,6-di-O-galloyl-β-D-glucose（13）,1-O-galloyl-4,6—（-）-hexahydroxydiphenoyl-β-D-glucose（14）,1,4,6-tri-O-galloyl-β-D-glucose（15）.The flavoalkaloids（1,2,3a*and 3b*）showed a promising activity of inhibition against the formation of advanced glycation end products（AGEs）with IC₅₀0 values at 25.26,13.48,and 10.33μM,respectively,better than the well known glycation inhibitor.The highlight of this experiment is that two novel compounds（1 and 2）together with new naturally occurring flavoalkaloids were isolated and identified from white tea.The absolute configuration of the novel flavoalkaloids was extensively elucidated by NMR and CD analyses.Inhibition against the formation of AGEs of the flavoalkaloids was studied and showed a promising activity.The flavoalkaloids are expected to be new drugs candidates for diabetes.Furthermore,the interaction of the four flavoalkaloids with trypsin,adenosine5′-monophosphate（AMP）-activatedproteinkinase（AMPKprotein）,α-glucosidase enzyme was studied by molecular docking technique.Further confirmation of these compounds has the potential to develop as diabetic therapeutics.