Study On The Active Constituents From Tripterospermum Chinense For Anti-H5N1Avian Influenza Virus

Highly pathogenic avian influenza（HPAI）causes a major threat to humans.Screening safe and effective anti-HPAI compounds in Chinese herbal medicinesprovides a large opportunity for the development of anti-HPAI drugs. In the dissertation,we selected66Chinese herbal medicines for anti-HPAI in vitro, employing the methodof MDCK cell cultures infected with H5N1subtype and using cytopathic effect (CPE) asactivity-evaluating method. Seven Chinese herbal medicines were found to have anti-HPAI activity during our experiments. Among them, the extract of Tripterospermumchinense showed the most prominent anti-HPAI activity, so our studies on chemicalconstituents and anti-HPAI activity of T. chinense has been processed deeply.Fifty constituents were separated through AB-8macroporous resin,recrystallisation, chromatography of silica gel, RP C18silica gel, Sephadex LH-20andPHPLC, and41of their structures were elucidated with physical and chemicalconstant, MS,1D and2D NMR (1H-1H COSY、HSQC、HMBC). There were5newcompounds. The structures of41compounds were as follows: tripterospermumcin C(1), tripterospermumcin D (1), tripterospermumcin E (3), tripterospermumcin F (4),tripterospermumcin G (5), tripterospermumcin B (6), sweroside (7), loganic acid (8),8-epi-kingiside (9), bergenin (10), isovitexin (11),7-O-rhamnopyranosyl-isovitexin(12),7-O-rhamnopyranosyl-isoorientin (13), saponarin (14),2′′-O-rhamnopyranosyl-trifoliside (15), trifoliside (16),8-hydroxy-1,2,6-trimethoxyxanthone (17),1,7-dihydroxy-3,8-dimethoxyxanthone (18),1,2,8-trihydroxy-5,6-dimethoxyxanthone(19), lancerin (20), chrysophanol (21), emodin (22), aloe emodin (23), physcion-8-O-β-D-glucopyranoside (24), citreorosein (25), emodin-8-O-β-D-glucopyranoside (26),emodin-8-O-β-D-glucopyranoside (27), aloe emodin-3-(hydroxymethyl)-O-β-D-glucopyranoside (28), oleanolic acid (29), ursolic acid (30), β-daucosterol (31),caffeic acid (32),3,4-dihydroxybenzoic acid (33),2-methoxyphenol (34),4-hydroxybenzoic acid (35), tetratriacontanoic acid (36), thymidine (37), urdin (38), β-sitosterol (39), stigmasterol (40), α-L-oleandrose (41a) and β-D-cymarose (41b).Compounds1,2,3,4and5were new compounds, including two iridoid tetramerswith four glucosides, two iridoid dimers with two glucosides, one β-lactone with aglucoside. Compounds9-10,12-16,21-29,31-38, and40-41were isolated from thegenus Tripterospermum for the first time. Compounds7-8,20, and30were isolatedfrom Tripterospermum chinense (Migo) H. Smith。for the first time.From the perspective of chemical structures,41compounds include8iridoidglycosides,8flavones,8anthraquinones,15other structural types. Compounds1and5are the first isolation of secoiridoid tetramers with four glucosides. Compound3is the first isolation of β-lactone with a glucoside. Compound4is the first isolation ofsecoiridoid dimers with two glucosides, linkage to one alkaloids. These types ofcompounds increase structures of natural products. The NMR data of compounds12and13were reported for the first time.Anti-H5N1activities of compounds11and X has been discovered by CPEactivity assay. At the effective dose of37.5μg/mL, compound X did not showapparent toxicity to cell viability, metabolism or proliferation, which exhibited apotent anti-influenza virus activity in MDCK cell cultures infected with H5N1subtyp.Compound X might become a new leading natural compound for the systhesis anddevelopment of new anti-HPAI drugs. At the effective dose of150μg/mL, compound11did not show apparent toxicity to cell viability, metabolism or proliferation, whichexhibited a potent anti-influenza virus activity in MDCK cell cultures infected withH5N1subtype.