Research On Chemical Constituents Of Citrullus Lanatus Vine And Pharmacodynamic Screening

Part Ⅰ. Study on Chemical Constituents of the Citrullus Lanatus Vine Experiment1:Preliminary Tests of Chemical Components for the Four Different Species Citrullus Lanatus VineObjective:To identify the chemical components of the four different species citrullus lanatus vine using comprehensive preliminary tests. Methods:The mixture was extracted from the four different species citrullus lanatus vine with purified water95%ethanol, and petrol ether was studied using test tube and chromatography method. The chemical components of the four different species citrullus lanatus vine were preliminarily demonstrated using precipitation reaction and color reaction of multiple indicators. Results:The four different species citrullus lanatus vine might contain sugar, organic acid, sapaomin, flavonoids, alkaloids, steride. Conclusion: The research provides foundation for further study and application of citrullus lanatus vine.Experiment2:Study on Chemical Constituents of Citrullus Lanatus Vine (I) Objective:To study the chemical constituents of citrullus l anatus vine in extracting with petroleum ether. Methods:Compounds wer e isolated and purified by polyamide column chromatography. Their chemi cal structures were elucidated on the basis of physicochemical properties, and spectral data. Results:Ten compounds were isolated from citrullus la natus vine andelucidated as β-sitosterol,daucosterol, stigmasterol, monomyri stin, palmitic acid, monopalmitin, monobehenin, ursolic acid, monoheneicos anoin, stearic acid. Conclusion:Compounds of1-10were obtained from c itrullus lanatus vine for the first time.Experiment3:Study on Chemical Constituents of Citrullus Lanatus Vine (Ⅱ)Objective:To study the chemical constituents of citrullus lanatus vine in extracting with ethyl acetate. Methods:Compounds were isolated and purified by polyamide column chromatography. Their chemical structures were elucidated on the basis of physicochemical properties, and spectral data. Results:Ten compounds were isolated from citrullus lanatus vine and elucidated as pentadecanoic acid, monopentadecanoin,2,3-Dihydroxypropyl nonadecoate, lignocericacid-2,3-dihydroxy-propanenyl ester, lancerebroside5, salicylic acid,4-Hydroxybenzoic acid, hydroquinone, succinic acid, vanillic acid. Conclusion:Compounds of1-10were obtained from citrullus lanatus vine for the first time.Part Ⅱ. Experimental Study on Anti-inflammation and Analgesia Effect and Acute Toxicity of Extract from Citrullus Lanatus VineObjective:To study the effect of extract from citrullus lanatus vine (EWRL) on anti-inflammation and analgesia and acute toxicity. Methods: The models of mice ear edema induced by xylene, carrageenin and cotton pellet granuloma in rats to observe the anti-inflammation effect of EWRL via oral administration. The effect of EWRL on analgesia was tested by measuring the latent period licking hind foot with the hot plate method and counting body twisting induced by acetic acid in mice. The acute toxicity of EWRL was measured by the method of Bliss. Results:In acute toxic reactions which were observed for14days, all mice were survived, and no toxic reaction was shown during the observed period. The maximum administration dosage was87.0g/kg of EWRL given to mice per kilogram. EWRL could significantly inhibit the ear edema caused by xylene in mice, granuloma hyperplasia caused by cotton in rats, paw oedema caused by carrageen in rats. It could significantly prolong the pain threshold on hot-plate in mice, reduce the writhing times in mice. Conclusion: EWRL has obvious effect on anti-inflammation and analgesia.Part Ⅲ. Study on Screening and Mechanism of Anti-inflammatory Activity of Different Fractions from Citrullus Lanatus VineObjective:To screening the extract parts of citrullus lanatus vine on anti-inflammatory activity and study its mechanism. Methods: Inflammatory model of LPS-stimulated RAW264.7cells were used, the level of impact on the release of NO as a symbol, and the petroleum ether, ethyl acetate, n-butanol,50%ethanol and soluble parts of anti-inflammatory activity on citrullus lanatus vine wered screeninged. The concentration of proinflammatory mediators TNF-α, IL-6, IL-1β, iNOS and COX-2HO-1by ELISA were determined. The activation nuclear translocation of NF-κB by immunofluorescence assay were detected, Real-time PCR for the determination level of IL-1β, IL-6, TNF-α, COX-2, iNOS and HO-1mRNA, and explored its anti-inflammatory mechanism. Results:Extract parts of citrullus lanatus vine significantly inhibited NO release of the LPS-stimulated RAW264.7cells inflammatory model, and the petroleum ether part was the strongest. The petroleum ether fraction significantly inhibited the overexpression of TNF-a, IL-6, IL-1β, iNOS, COX-2, but have no effect on the levels of HO-1in inflammatory model. It could also inhibit the activation of NF-κB nuclear translocation, and reduce the level of IL-1β, IL-6, TNF-α, COX-2, iNOS and HO-1mRNA. Conclusion:Petroleum ether fraction of the watermelon vine was the anti-inflammatory active site, which may achieve the anti-inflammatory role by inhibiting the activation of NF-κB nuclear translocation.Part IV. The Screening of Hypoglycemic Activities of Different Fractions from Citrullus Lanatus VineExperiment1:The Hypoglycemic Effects of Different Fractions from Citrullus Lanatus Vine on Diabetic Model Induced by Alloxan in MiceObjective:To investigate the hypoglycemic effects of different fractions from citrullus lanatus vine on diabetic model induced by alloxan in mice. Methods:Healthy mices were used to establish diabetic models by peritoneal injection administration of alloxan. The different fractions from citrullus lanatus vine was applied as raw materials for potential marine drug. The levels of fasting blood glucose(FBG)、 fasting blood insulin (FBI) and glucose tolerance were detected by automatic blood glucose device. The concentrations of total cholesterol (TC)、 total glycerin (TG) nitric oxide (NO) and nitric oxide synthase (iNOS) in blood were respectively measured. The concentrations in liver homogenate of nitric oxide (NO) and nitric oxide synthase (iNOS) were determined. Pathology of pancreas was observed. Results:After treatment, ethyl acetate high dose group and Petroleum ether high dose group of citrullus lanatus vine all significantly decreased the levels of TC、 TG、 NO and iNOS. Meanwhile, blood glucose level and glucose tolerance were improved. Insulin levels was also improved. Conclusion:Ethyl acetate group and Petroleum ether group of citrullus lanatus vine shows significant hypoglycemic effect on diabetic model induced by alloxan in mice. The mechanism of its hypoglycemic effect may be promoted the insulin release and decreased the levels of NO and iNOS.Experiment2:The Protective Renal Tubular Effects of Ethyl Acetate Extract from Citrullus Lanatus Vine in Rats with DNObjective:To investigate the protective renal tubular effects of ethyl acetate extract from citrullus lanatus vine in rats with DN. Methods: Healthy rates were used to establish DN models by peritoneal injection administration of streptozotocin. The ethyl acetate extract from citrullus lanatus vine was applied as raw materials for potential marine drug. The levels of fasting blood glucose(FBG)、fasting blood insulin (FBI)、serum creatinine (Scr)、urine creatinine (Ucr) and urinary albumin(UAlb) were detected. Pathology of live was observed. Results:After treatment, ethyl acetate high dose group and medial dose group of citrullus lanatus vine all significantly decreased the levels of GLU、FBI、UAlb、KWI and TBI.Meanwhile, Ccr was also improved. Conclusion:The ethyl acetate extract from citrullus lanatus vine can protect renal tubular in rats with DN.Part Ⅴ. Antimicrobial Activity of the Extracts from Citrullus Lanatus VineObjective:To study antimicrobial activity in vitro of citrullus lanatus vine extract. Methods:Agar dilution method was applied to detect antimicrobial effect of citrullus lanatus vine extract in vitro; the mice celiac injection of staphylococcus aureus method was applied to detect the antimicrobial effect of citrullus lanatus vine of ethyl acetate extraction in vivo. Results:Citrullus lanatus vine extract had different inhibition effects on staphylococcus aureus, escherichia coli, bacillus aeruginosus, salmonellatyph, bacillus subtilis and klebsiella pneumoniae, but it showed weak inhibition effect on chain coccus. Effects of80%alcohol extract and ethylacetate extract were better than others on staphylococcus aureus. The minimum inhibition concentration (MIC), respectively is4.2mg·mL-1,8.4mg·mL-1. Conclusion: Citrullus lanatus vine extract have antibacterial action, and provide a basis to exploit new antimicro-bic drug.Part VI. Anti-tumor Active of Lancerebroside V from Citrullus Lanatus Vine in VitroObjective:To study the anti-tumor active of Lancerebroside Ⅴ extracted from citrullus lanatus vine in vitro. Methods:MTS method was used to test inhabitation of Lancerebroside Ⅴ from citrullus lanatus vine with five kinds of human tumor cells in vitro and IC50values were calculated respectively. Results:IC50values of these LancerebrosideV tested with five cancer cell lines were3.9～8.1μg·mL-1. Conclusion:To some extent, Lancerebroside V from citrullus lanatus vine have positive effect on inhabiting those human tumor lines in vitro.