Structural Characterization Of Phenolic Compounds In Clausena Lansium And Its Protective Effects On Alcoholic Liver Injury

Wampee(Clausena lansium),a characteristic fruit in south China.Modern researches demonstrated that wampee was beneficial to liver,the water extracts of wampee have protective effect on liver injury caused by both acute alcoholism and chronic alcoholic liverinjury in mice.However,the variety and structure of the active ingredients in wampee are still not clear.The detailed research content and results are as follows:(1)The protective effect of wampee phenols on alcoholic liver damage was evaluated in basis of mice model of alcoholic liver injury induced by Lieber-DeCarli alcohol liquid.The mice were intervened with different doses(100 and 300 mg/kg bw)of WP for 8 weeks.The results showed that the liver index,glutamic-pyruvic transaminase(ALT)and glutamic oxalacetic transaminase(AST)activities of WP group were significantly lower than those of the model group.The results of HE staining showed that both low and high doses of WP could alleviate the liver lipid aggregation caused by alcohol,indicating that different doses of WP had significant protective effects on alcohol-induced liver injury in mice.Compare to the model group,the level of TG,TC in serum and MDA content in both serum and liver were significantly decreased,and the activity of SOD,CAT and GSH-PX in serum and liver were significantly increased when treated with different doses of WP.All of these results demonstrated that WP might inhibit alcoholic liver injury in mice by controlling the stress of oxidization and lipid accumulation.(2)The WP extracts were isolated by solvent extraction,liquid distribution combined,and further purified by passed through normal silica gel column chromatography,middle-pressure column chromatography,gel column chromatography and highperformance liquid chromatography,the structures of isolated monomeric compounds were elucidated by various spectroscopy techniques such as UV,high-resolution mass spectrometry and NMR.As a result,total phenols and total flavonoids with purity of 88.7%and 85.6%respectively were obtained,and 22 monomeric components were isolated from WP,including 13 flavonoids:Quercetin-3-O-(4-O-β-D-glucoside)-α-L-rhamnoside(1),Isorhamnetin-3-O-(4-O-β-D-glucosyl)-α-L-rhamnoside(2),Myricetin 3-glucoside(4),Quercetin 3-rutoside(5),Epigallocatechin(6),Gallocatechin(7),2-Hydroxynaringin 5-O-β-D-glucopyranoside(8),Maesopsin-6-O-D-glucopyanoside(9),Myricetin(12),2-Methoxy-4-(2-propenyl)phenyl-O-β-D-g-lucopyranoside(19),Syringin 3-O-β-DGlucoside(20),Quercetin 3-O-L-rhamnoside(21),5,7,4’-trihydroxy-3’,5’Dimethoxyflavone 3-O-α-L-rhamnopyranoside(22);6 phenylpropanoids:6-β-D-glucosyl6,7-dihydroxy-5-benzofuranpropanoic acid metal ester(3),Syringin(10),Scopoletin(13),Ferulic acid(15),Trans-dehydrodiferulate dimethyl ester(14),Methyl 3-(2-O-β-D-Glucopyranosyl-4-hydroxyphenyl)propanoate(11)and 3 sesquiterpenes:Corionioside C(16),Trans-2,6-dimethyl-6-hydroxyocta-2,7-dienoic acid β-glucopyranosyl ester(17),Brucea Javanica B(18).Among which compounds 1、2 and 3 were new compounds,except for compound 5,the other 18 compounds were isolated from wampee fruit for the first time.(3)The antioxidant activities of wampee phenols were evaluated by DPPH+free radical scavenging method(DPPH),ABTS+free radical scavenging method(ABTS)and oxidation free radical absorption capacity(ORAC)method.The results showed that flavonoid compounds 6,7,9,21 and phenylpropanoid compounds 3,15 had better ABTS and DPPH free radical scavenging activity than ascorbic acid,the positive control.Flavonoid compounds 1,2,6,7,9 and 21 also showed good ability of oxygen-free radical scavenging,and their ORAC value was significantly higher than gallic acid,the positive control.Therefore we concluded that flavonoid compounds 1,2,6,7,9 and 21 and phenylpropanoid compounds 3 and 15 are the main active compounds contribute to antioxidant activity.It can been deduced that the 3’,4’ o-diphenol hydroxyl substituted B ring in flavonoid is an important antioxidant active functional group,and methoxy substitution or changes in the number of hydroxyl groups will significantly reduce the antioxidant activity.The antioxidant activity of the glucosyl substitution at the 3-position of flavonoids is significantly better than that of rhamnosyl substitution,while the type of substitution of the disaccharide at the position has no significant effect on the antioxidant activity.