| Agricultural by-product refers to the non main products that have been produced in the process of agricultural production. The reasonable development and utilization of agricultural by-product is of great economic value and practical value,and they have gradually become a hot issue in scientific research.The research of agricultural by-product has developed rapidly in recent years. The research field mainly involves the extraction and identification of natural active substances, the effect and mechanism of active ingredients. Prevention and resistance to aging is a major and important research direction in the study of the effect of action. Aging refers to the growth and development of the biological body after the mature and reach the exuberant period, the physiological function of the growth of the physiological function gradually decreased, the organization and the organs gradually degenerated, the cell metabolism can decline, and eventually to the process of death. Nowadays, the aging of population is becoming more and more serious, and the development and use of the disease, aging mechanism and anti-aging drugs have been included in the scientific research.The extraction and separation of natural active substances and active ingredients are the primary key to the study of the basic theory of food and drug effects and mechanism, and the optimization of the extraction method and extraction technology has been studied. However, due to many reasons, such as a wide variety of plants, active ingredients and content, and extraction methods, the research needs to be improved. In this study,5 kinds active substances of 3 kinds of agricultural by-product (Stigmata maydis, Rape pollen and Ganoderma spore powder) were improved. The efficiency and quality of the extraction were improved. The effects and mechanism of 3 kinds of raw materials and active substances were used to study the effects and mechanism of anti animal cells.In the study of the aging problem, the selection and establishment of the suitable animal (cell)model is a necessary condition for the successful test. Because of ciliates is suitable for scientific research, the research on the report in the literature and preliminary test, try to model ciliate cell aging model was established, on the one hand solves the carrying out agricultural and sideline products anti-aging research of this topic, the animal (cell) model needs a great quantity of problems; On the other hand for other researchers for discussion on the problem of aging provides a good model for the study, provides a new way for anti-aging research. The main research contents and results are as follows:(1) In this study, we optimized the extraction process of the traditional extraction methods such as water extraction and alcohol precipitation method, the ethanol extraction of the flavonoids from corn, the ethanol extraction of rape pollen and the supercritical CO2 extraction of ganoderma spore oil: The optimal solution of Stigmata maydis Polysaccharide water extraction and alcohol precipitation method was:the extraction time is 2 h, the extraction temperature is 100℃, the ratio of material to liquid ratio is 1:35, ethanol volume fraction is 70%. Polysaccharide extraction rate was 4.95%; The optimal solution of Stigmata maydis Flavone alcohol extraction method was:the extraction time is 1.5 h, the extraction temperature is 80℃, the ratio of material to liquid ratio is 1:35, ethanol volume fraction is 70%. Flavone extraction rate was 0.45%; The optimal solution of Rape pollen Polyphenol alcohol extraction method was:the extraction time is 1.5 h, the extraction temperature is 70℃, the ratio of material to liquid ratio is 1:35, ethanol volume fraction is 80%. Polysaccharide extraction rate was 0.45%;④ The optimal solution of ganoderma spore oil supercritical CO2 extraction method was:the extraction pressure was 25 MPa, the extraction temperature was 40℃, the CO2 flux was 15 L/hl, the extraction time was 80 min. Ganoderma spore oil extraction rate was 15.36%.(2) The semi-bionic extraction method (pH method) was optimized, and the pH gradient was set up, and the optimal extraction process was obtained by orthogonal test: ①The optimal solution of Stigmata maydis Polysaccharide semi-bionic extraction method was:extraction time was:Extraction time was 2 h, the ratio of material to liquid ratio was 1:25, the pH value was 2.5, the pH value was 8, the extraction rate was 5.69%; The optimal solution of Stigmata maydis Flavone semi-bionic extraction method was extraction time was:The extraction time was 2 h, the ratio of material to liquid ratio was 1:35, the pH value of simulated gastric juice was 2.5, the pH value of the simulated intestinal fluid was 8, and the extraction rate was 0.50%; The optimal solution of Rape pollen Polyphenol semi-bionic extraction method was:extraction time was:The extraction time was 1.5 h, the ratio of liquid to material ratio was 1:15, the pH value of simulated gastric juice was 2.5, and the pH value of the digestive juice was 9, and the extraction rate was 0.52%.(3) The rape pollen protein and ganoderma spore oil of semi bionic extraction (enzymatic) improvement and optimization. Based on the use of materials of special (broken) attempts to use animal digestive enzymes pretreatment, according to determine the types of substances extracted from kinds of enzyme added, provided suitable enzymes play a role in vivo environment (pH and temperature), the optimum extraction process, effectively improves the extraction efficiency and quality: The optimal solution of rape pollen 4 protein semi-bionic extraction method was:extraction time was:treatment time was 3h, solid-liquid ratio was 1:15; ②The optimal solution of ganoderma spore oil semi-bionic extraction method was:the treatment time is 11:20, the ratio of material to liquid ratio is h, the pH value of simulated gastric juice is 2.5, and the PH value of the digestive process is 9. Ganoderma spore oil extraction rate was 17.02%.(4) The research studied the ciliate sexual reproduction control: ①Optimization of the micro operation method, cell division 2 periods showed high amicronucleate rate:early middle-late 40%, in 5%; ②Screening out two small injury on the cell of conventional drugs, the inhibition of ciliates cell nuclear, get higher amicronucleate rate:norfloxacin on Paramecium caudatum nuclear removal rate was 23.33%; hypochlorous acid on Paramecium caudatum nuclear removal rate is 26.67%; hypochlorous acid to like weaving caterpillars nuclear removal rate is 10%; ③Application of modified carbol fuchsin nuclear staining of ciliates from Department of conjugation of facilitating conditions and nuclear period,1.0g/ ml and papain on Paramecium caudatum self adhesive has good role in promoting, contributed to the rate was 5.8%; Paramecium caudatum system with large nuclear activities for 2-36 h.(5) Studied three ciliates (Paramecium caudatum, Paramecium tetraurelia and Tetrahymena thermophila) non-amicronucleate clones ageing law, and construct a model of aging:①Clarify three ciliates natural growth process and aging law, make clear the relationship between the elimination of oxygen free radicals-age-generation age between; ② Selected for the higher animals cell aging model was constructed and the two drugs (D-galactose and hydrogen peroxide) of 3 species of ciliate cells by different ways of Administration for processing, research on cell of senescence promoting role, confirmed the two drugs on ciliates cell aging model construction; ③Select 5 kinds 13 higher animals(cells) and ciliates commonly used indicators, to quickly build ciliate cell aging model were evaluated to clarify the close relationship between these indexes and cell senescence.(6) We studied the anti ageing effect of 5 kinds of active substances and 3 kinds of agricultural by-products by ciliates non-amicronucleate clones ageing law. The results showed that 3 kinds of agricultural products and 5 kinds of active substances showed different degrees of resistance to cell senescence: ①Effect of Stigmata maydis Polysaccharide on aging cells:The highest decrease of the density of β-galactosidase was 20.16%;The highest rate of Caspase-9 was 20.12%;the expression of CDC-2 mRNA gene was 1.28. The effect of flavonoids was similar to polysaccharide, but the effect was generally lower than that of polysaccharide; ②Rape pollen 4 kinds phenolic acids and 4 kinds proteins were used in Tetrahymena thermophila aging cells,Cell proliferation rate,Growth cycle and expression of RCC-1 gene mRNA had a more significant effect than the control group; ③Ganoderma spore oil maximum proliferation on the natural senescence of Paramecium caudatum cells was 36.67%. The optimal effect of the life cycle of D-Gal promoting high failure model was 7.38 days, added 14.86 generation. Eliminate hydroxyl radical capacity increased by30.77%, and the content of MDA was 22.22%. The lipofuscin content in the cell loss rate was 20.69%. Ganoderma spore oil has a significant effect on the 5 indexes of Paramecium caudatum; ④Ganoderma lucidum spore powder to extend high attenuation dying of Paramecium caudatum’s survival and proliferation effect is remarkable:prolonged survival time was 77.8 hours, added 4.4 generationan.Greatly strengthen the Paramecium caudatum high attenuation the physiological activity of cells:maximal exercise capacity increase rate of 65.02%; feeding ability maximum increase rate 61.90%; excretion and moisture ability to regulate the maximum increase rate was 83.87%; The effect of anti ageing effect of 3 kinds of agricultural by-products is compared:Ganoderma spore powder were significantly higher than those of rape pollen and Stigmata maydis,no significant difference between the corn and rape pollen. |
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