The Study Of Preparation And Biological Activity Of Ophiurasaponin From Ophiopholis Mirabilis

In this paper, Ophiopholis mirabilis from the Changdao Archipelago was used as the experimental material. The major nutritional components of Ophiopholis mirabilis was determined.The extraction process of ophiurasaponin was optimized by the orthogonal experiment. The physical and chemical properties of ophiurasaponin were determined. We aiso studied on the extraction, purification and part of the biological activity of ophiurasaponin,including antioxidant activity, antimicrobial activity and antitumor activity.These studies provided the necessary theoretical basis for the development and utilization of Ophiopholis mirabilis.We determined the major nutritional components of Ophiopholis mirabilis(overall), including moisture, ash, protein, total sugar,rude fat, fatty acids, amino acids and inorganic elements. The concentration of moisture, ash, protein, total sugar and rude fat were 4.09%, 25.71%, 18.27%, 3.53% and 1.74%, respectively. The concentration of Ca, Mg, Fe were high among the inorganic elements. Unsaturated fatty acids was accounted for 72.61% of total fatty acids.There were especially rich in essential polyunsaturated fatty acids,such as EPA andαlinolenic acid.And the essential amino acid index (EAAI) was 25.90. Ophiopholis mirabilis had broad potential value,and it can be further considered the development of new functional food or animal feed.Orthogonal experiment has been used to investigate the extraction process of ophiurasaponin. The extraction temperature, extraction time, ratio of the material to water and times of extraction were selected as variable parameters. Their different effects on the yield of ophiurasaponin that extracted from Ophiopholis mirabilis were studied by the L9(34)experimental design method. In addition, a multiple linear regression(MLR) model, in which the yield of ophiurasaponin was as the dependent and the experiment conditions of the orthogonal experiment was as the independent, was firstly constructed to predict the yield of ophiurasaponin of two experiment conditions in the orthogonal experiment. The statistic parameters were R = 0.914, F = 10.585. And the MLR model was used to predict yields of ophiurasaponin of two experiments. The predicted results were in good agreement with the experimental values, indicating that the model can be used to predict the yield of ophiurasaponin.The crude ophiurasaponin was obtained by the traditional solvent extraction.We determined the physical and chemical properties of ophiurasaponin , including color reaction, melting point, hemolytic index, UV absorption spectra and infrared spectra.Polysaccharides and inorganic salts were removed through the AB-8 macroporous resin followed by water, 20% ethanol, 40% ethanol, 60% ethanol, 80% ethanol. Some 40% ethanol elution was through silica gel column chromatography, Sephadex LH-20 silica gel column chromatography, and reverse column chromatography.Ultra-Weak Luminescence Analyzer was used to detect the antioxidant activity of ophiurasaponin.The results showed that the antioxidation capabilities increased linearly with the increasing of the concentration of ophiurasaponin in a certain range. The result showed that ophiurasaponin from ophiopholis mirabilis had strongest inhibition to hydroxyl, weakest to hydroxyl radicals, and mediate to superoxide. Ophiurasaponin from ophiopholis mirabilis had significant antioxidant activity, it may be considered to develop into a new type of antioxidant agents.Using circular filter paper method , the antibacterial activity of ophiurasaponin was detected. The minimum inhibitory concentration (MIC) of ophiurasaponin was determined by modified agar dilution method. The results showed that the antibacterial activity of 40% ethanol elution was enhanced than the crude ophiurasaponin. After the initial silica gel column chromatography, the part of the chloroform had the strongest antibacterial activity, followed by the part of chloroform: methanol: water = 9:1:0.1 and chloroform: methanol: water = 8:2:0.2 . The MIC of Escherichia coli and Bacillus subtilis was 0.0443mg/mL,and it was the 40% ethanol elution through macroporous resin.The MIC of Staphylococcus aureus, Aerobacter aerogenes, Proteus, Penicillium, Mucor, Aspergillus, Saccharomyces cerevisiae was 0.0148mg/mL,and it was the first strip of chloroform. It had no inhibitory effect on Saccharomyces cerevisiae Hansen.Cytotoxic effect of human liver cancer cell NCI-H460 and human lung cancer cell HepG2 were assayed using SRB method in vitro.The results showed that the inhibition rates were 38.03% and 34.57% when the drug reached 1000mg/L.The inhibition rate decreased rapidly when the adminitration was low.In short, Ophiurasaponin had no significant inhibition of tumor cells.