Studies On Constituents, Biological Activities And Quantitative Analysis Of Iridoids In The Roots Of Gentiana Straminea Maxim

The roots of Gentiana straminea Maxim.(Gentianaceae) has the effect of rheumatism, clearing heat and easing bitongkang, which has been used for the treatment of rheumatic arthritis, icteric hepatitis, constipation, pain and hypertension. Little is known about constituents and biological activities of Gentiana straminea Maxim.23 compounds were isolated from the roots of Gentiana straminea Maxim on column chromatography of silica gel, Sephadex LH-20, ODS and polyamide. Among them,22 compounds were identified on the basis of chemical and spectral methods as:7 (S)-n-butyl morroniside (1), 7(R)-n-butyl morroniside (2),6'-O-acetyl gentiopicroside (3), 6'-O-(2-hydroxy-3-O-β-D-glucopyranosyl-benzoyl)-sweroside (4), gentiopicroside (5), swertiamarin (6), sweroside (7), logannic acid (8), 6'-O-β-D-glucosylgentiopicroside (9),2'-O-(2,3-dihydroxyl-benzoyl)-sweroside (10), macrophyiioside D (11), (-)-syringaresinol-4,4'-bis-β-O-D-glucopyranoside (12), gentiaphyllide-D (13), gentiaphyllide-E (14), erythrecentaurin (15), roburic acid (16), uvaol (17), daucosterol (18),β-sitosterol 19), gentianose (20),β-glucose (21) andα- glucose (22). Among these compounds,1,3 and 4 were new compounds; 2 was isolated from this genus for the first time; 6-9,11-16 and 20 were isolated from this plant for the first time.Antioxidation bioactivity of five secoiridoids (4,5,6,7,9) isolated from the roots of Gentiana straminea Maxim were tested. On the basis of experiment data, the link of structure and activity was discussed. We found that these compounds suppressed the superoxide generation in human neutrophils induced by N-formyl-methionyl-leucyl-phenylalanine (fMLP) in a concentration-dependent manner, especially compound 9 showed the strongest effect. Compound 4 and 5 suppressed the superoxide generation in human neutrophils induced by Arachidonic acid (AA) and Phorbol-12-myristate-13-acetate (PMA) and compound 4 showed the stronger effect. The effect of the five secoiridoids on lipid peroxidation of erythrocyte membrane ghosts by hydroxyl radicals was also studied and we found these compounds exhibited almost no effect on lipid peroxidation level. It was found that the five secoiridoids showed no hemolytic effect even at high concentrations, which was much higher than the concentrations sufficient for the suppression of superoxide generation. We also studied the effect of compound 4 and 5 on auricula edema in mice by induced dimethylbenzene. Two compounds showed inhibit activity and compound 4 exhibited stronger effect.In this study, a new HPLV-UV/MS method was developed for simultaneous determination of ten iridoid and secoiridoids in the roots of G. straminea. Separations were performed on a Kromasil-C18 column by gradient elution using methanol and water with phosphoric acid. The velocity, column temperature and detect wavelength were 1.0 mL/min, 25℃and 254 nm, respectively. Analytes were also identified by HPLC coupled with electrospray ionization mass spectrometry experiments. The quantitative method was validated for linearity, precision, accuracy, and limits of detection and quantification.