| Kalimeris indica (L.) Sch-Bip) belongs to family Asteraceae, genus Kalimeris. The whole herb of it has been used as Chinese medicine to treat heat clearing, detoxifying, scattered stasis, hemostatic, warming the lung and relieving cough, dispelling cold and relieving pain, rich in natural resources, folks always eat lots as liver and eyes health food.Traditional Chinese medicine use it treat hepatitis B significant clinical effect. Kalimeris integrifolia Turcz.ex DC is the related species of Kalimeris indica, bitter in taste and cold-nature, with detoxification, relieving cough, stasis to stop bleeding effect.Objective:study material basic of Kalimeris indica (L.) Sch-Bip and its related species about anti immunoiogical liver injury, and lay the foundation for search the lead compound with anti immunoiogical liver injury activity.Methods:The dried and crushed aerial part of Kalimeris integrifolia Turcz.ex DC. (13 Kg) was reflux extracted with 85%(v/v) ethanol in a water bath. The solution was evaporated under reduced pressure to obtain an extract (2.56Kg). Using normal phase silica gel section first, and then the integrated use of positive phase silica gel, reverse phase silica gel, Sephadex LH-20 glucan gel preparation, medium pressure and high liquid phase preparation chromatography for purification and separation, and using NMR, MS, UV, IR and other modern spectrum technology for separation of the structures of the monomer compounds;And respectively to study the response surface analysis to optimize extraction technology of total flavonoids of Kalimeris indica and macroporous resin purification process of total flavonoids of Kalimeris indica.Results:Twenty compounds were isolated and identified as friedelan-3-ol (1), stigmast-7,22-ene-3-dione (2), friedelin (3), syringaresinol (4), a-spinasterol (5), ciwujia-tone (6), syringic acid (7), Kalinturflavone A (8), Kalinturflavo-ne B (9), scopoletin (10), apocynin (11), 1-(3-hydroxy-4,5-Dimethoxyphen-yl)ethan-l-one (12), apigenin (13),5-hydroxymethylfurfural (14), stigmasterol-3-O-?-D- glucopy-ranoside (15), Kalinturoside A (16), Bidenoside C (17), citrusin (18), Iirioresinol A (19), syringaresinol-4-O-?-D-glucopyranoside (20). Compounds 8,9, 16 are new compounds, the rests were isolated from Kalimeris integrifolia for the first time.The response surface methodology results indicated that the optimum extraction conditions were as follows:40% ethanol concentration,16.5:1 liquid-solid ratio,2 h of extraction time,3 times of extraction times. In this condition, the predicted and experimental values were 162.538 and 156.867 mg·g-1. And it is concluded that the factors, in turn, affects the quantity of flavonoids extraction size for ethanol concentration> solid-liquid ratio> extraction times> extraction time.After concentration of the sample, elution rate, sample volume, ethanol concentration and amount of the elution process parameters were investigated stars macroporous resin purification of total flavonoids Kalimeris indica best process parameters:adsorption HPD-300 was 5.64 mg·mL-1, parsing was 82.54%,8 macroporous resin were the highest value; optimum sample concentration of 0.1 g·mL-1, the best on a sample flow rate of 2 ml·min-1, the largest the sample volume is 0.239 g·mL-1, the best elution ethanol concentration and dosage were 90% and 2 column volumes.Animal experiment showed that compared with normal group, model group index data were extremely significant, and between groups of data statistical analysis found that TFK and TFK.T can reduce high immunological liver injury in mice serum ALT, AST, NO, TBiL, AKP, MDA, TNF-a, IFN-y, IL-4, iNOS level, against the SOD activity decreased, increase GSH-Px levels, means that TFK and TFKT protective effect of immunological liver injury in mice. |
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