Studies On Chemical Constituents And Their Hpyerglycemic Activities Of Silybum Marianum

Silybum marianum(L.)Gaertn.,from the family of Asteraceae,is an annual or biennial herbal,native to the Europe,Mediterranean,North Africa,and Asia regions.Its fruits are used to clear heat and remove toxicity,and protect liver.Recent pharmacological study suggested that S.marianum possessed hepato-,neuro-,and cardio-protective and anti-inflammatory,anti-virus,anti-tumor,and anti-diabetic activities.By means of various chromatographic methods such as silica gel,Sephadex LH-20,opening ODS column chromatography,and semi-preparative HPLC,44 compounds were isolated from the 95%ethanol extract of the seeds of Silybum marianum(L.)Gaertn.Their structures were elucidated on the basis of spectroscopic data and physico-chemical methods.The compounds were comprised by 11 flavonolignans,6 amides,4 terpenoids,7 flavonoids,4 lignans,7 phenolic acids,and 5 others.Among them,compounds 12-13,19-21,and 44 were new,and named as mariamides A,B(12-13),12-hydroxyl anhuienosol(19),2-hydroxymethyl-5-(2-hydroxypropan-2-yl)phenol(20),mariaterpenoside A(21),and trideca-5,7,9-triyne-1,2,3,12-tetraol(44),respectively.The known compounds were identified as silybin A(1),silybin B(2),isosilybin A(3),isosilybin B(4),silychristin A(5),silychristin B(6),isosilychristin(7),dehydrosilychristin(8),silydianin(9),dehydrosilydianin(10),silyamandin(11),4-hydroxy-N-{4-[(E)-3-(4-hydroxy-3-methoxyphenyl)prop-2-enamido]butyl} benzami de(14),glochidiatusamide(15),N,N’-1,4-butanediylbis(4-hydroxybenzamide)(16),4,4’-diphenylmethane-bis(methyl)carbamstes(17),2-(3-hydroxy-4-methylphenyl)propan-l-ol(18),3,5,7-trihydroxychromone(22),dihydroquercetin-4’-methyl ether(23),dihydrokaempferol(24),dihydroquercetin(25),kaempferol(26),naringenin(27),naringonin-7-O-β-glucopyranoside(28),dehydrodiconiferyl alcohol(29),dihydrodehydrodiconiferyl alcohol(30),dehydrodiconiferyl alcohol-y’-methyl ether(31),dehydrodiconiferyl alcohol-4-β-D-glucoside(32),cinnamic acid(33),ferulic acid(34),coniferin(35),glucosyl methyl ferulate(36),hydroquinone(37),vanillic acid(38),protocatechuic acid methyl ester(39),uracil(40),allantion(41),β-adenosine(42),and IAA-OMe-N-Glc(43),respectively.Among them,10 and 18 are new natural compounds and compounds 14-18,23-24,26-28,30-39,41,and 43 were obtained from the Silybum genus for the first time.The isolates were evaluated for their antioxidant,NO inhibitory,and antidiabetic activities.The antioxidant effects of all compounds were assessed using the ABTS,DPPH,and FRAP radical scavenging assays.Significant differences were observed in these assays.In the ABTS assay,most of the compounds showed significant antioxidant activity.It could be found that only amide compounds showed moderate DPPH radical scavenging activity.However,in the FRAP assay,amides,flavonolignans,flavonoids,and lignans exhibited moderate to strong activity against FRAP.The isolated flavonolignans,lignans,and terpenoids were further tested for their inhibitory activities on LPS-induced NO production in murine microglial BV-2 cell line by the Griess reaction.All the tested isolates exhibited inhibitory effects on LPS-stimulated NO production.Among them,lignans and terpenoids showed moderate inhibitory effect,which was comparable to the positive control SMT.However,flavonolignans showed no inhibitory effect in effective concentration.We further evaluated the α-glucosidase and PTP1B inhibitory activities of the isolated compounds.The results showed that flavonolignans and amides exhibited significantα-glucosidase inhibitory activities while flavonolignans and flavonoids exhibited significant PTP1B inhibitory activities.The binding effect of flavonolignans and catalytic active site was evaluated by molecular docking for understanding the significant inhibitory activities of flavonolignans on PTP1B.The results suggested flavonolignans could well inhibit the activities of PTP1B by hydrogen bonds with key amino acid residues such as cys215,lys120,arg221,and arg24 and so on.We evaluated the protective mechanism of silychristin A on STZ-and HG-induced pancreatic INS-1 cells oxidative damage and its hypoglycemic effect in type I diabetic rats to understand the preventive and protective effect of silychrisitn A in diabetic mellitus.The results suggested silychristin A could increase insulin secretion by protecting pancreatic islet β cells from oxidative damage to make hyperglycemic effect.This paper revealed the chemical constituents of Silybum marianum(L.)Gaertn.and their in vitro and in vivo bioactivity,which will provide a basis for further development and utilisation of flavonolignans in diabetic mellitus.