Protective Effect Of Exo-polysaccharides From Inonotus Hispidus On Alcoholic Liver Injury And Its Liquid Fermentation Optimization

So far,alcohol becomes the second leading cause of liver damage following hepatitis virus.Developing safe and effective drugs to prevent and treat alcoholic liver injury is of great significance.Inonotus hispidus is a precious medicinal fungus,which has been widely used in the treatment of various liver diseases and stomach diseases.However,most of these effects are lack of systematic experimental data to support,which greatly limits the further development and utilization of the Inonotus hispidus.Polysaccharide is one of the main active components of Inonotus hispidus,which plays an important role in its biological activity.However,the fruiting bodies of Inonotus hispidus are annual,so it is difficult to obtain the polysaccharide quickly and in large quantities.The liquid fermentation technology can produce polysaccharide rapidly and in large quantities,which is easy to realize industrialization.In order to promote the development and utilization of Inonotus hispidus,a suspected Inonotus hispidus parasitics from jujube tree was identified.Then,the effect of the exo-polysaccharides from liquid fermentation of Inonotus hispidus on alcoholic liver injury in mice and its pharmacological mechanism were analyzed.After clarifying that Inonotus hispidus exo-polysaccharides could alleviate alcoholic liver injury,the fermentation technology and formulation of exo-polysaccharides of Inonotus hispidus were further explored.The main research contents and results are as follows:(1)The identification of the wild suspected Inonotus hispidus was made by morphological characteristics and ITS rDNA sequence analysis.The genetic diversity of ciliates was analyzed based on the length,base composition and variation of ITS1-5.8S-ITS2 sequence.The results showed that the wild strain was Inonotus hispidus,which had abundant genetic diversity in nature.(2)The exo-polysaccharides of Inonotus hispidus was prepared through liquid fermentation.DEAE-52 cellulose column and Sephadex column were used to separate and purify the exo-polysaccharides,and a major polysaccharide component(IHEPS)was obtained.The physicochemical properties of IHEPS were analyzed by HPLC,UV-Vis and FT-IR.The content of polysaccharide and protein in IHEPS was 98.64% and 0.06%,respectively.It mainly composed of glucose,rhamnose,mannose,arabinose,galactose and glucosamine,with the mass percentages of 96.19%,0.53%,0.56%,0.73%,1.75% and 0.25%,respectively.Glucose is the main monosaccharide,of which the skeleton is composed of ?-D glucopyranose.(3)After defining the physical and chemical properties of IHEPS,the protective effect of IHEPS on acute alcoholic liver injury in mice was further explored.C57BL-6 mice were given IHEPS by gavage for 3 weeks,4 hours after the last administration,the mice weregiven alcohol to cause acute alcoholic liver injury.The protective effect of IHEPS on acute alcoholic liver injury in mice was evaluated by recording the time of drunkenness and sobriety of mice,measuring the physiological and biochemical indexes of serum and liver,and analyzing the liver pathological sections and the mRNA expressions of related antioxidant enzymes.The results showed that IHEPS had a good protective effect on acute alcoholic liver injury in mice.Compared with ethanol group,IHEPS could significantly prolong the drunken time and shorten the sobering time of mice(p < 0.05),decrease the liver index,aspartate aminotransferase(AST),alanine aminotransferase(ALT)and malondialdehyde(MDA),and increase the activities of superoxide dismutase(SOD),catalase(CAT),and ethanol dehydrogenase(ADH).The results of pathological sections showed that IHEPS significantly reduced the infiltration of inflammatory cells and lipid accumulation and improved the histomorphology of hepatocytes.The results also showed that IHEPS could activate Nrf2 signaling pathway and increase the expression of downstream antioxidant enzymes,such as NQO1,CAT and Cu-Zn SOD.(4)The fermentation conditions of exo-polysaccharides from Inonotus hispidus were explored and optimized.In the experiment,proper amount of Vernonia amygdalina leaf powder was added to the fermentation medium to increase the yield of Inonotus hispidus exo-polysaccharides,and the fermentation parameters were optimized by the orthogonal experiments design.The results showed that Vernonia amygdalina leaf could significantly increase the yield of Inonotus hispidus exopolysaccharides.Under the optimal conditions,the yield increased about 216% compared with that of control group.In this thesis,the amelioration effect of exo-polysaccharides of Inonotus hispidus on alcoholic liver injury in mice was clarified and its preliminary pharmacological mechanism was explored to give the the pharmacological efficacy of polysaccharides from Inonotus hispidus,which would provide a new idea to develope its further related products.At the same time,the fermentation process of exo-polysaccharides from Inonotus hispidus was optimized.The study provides a theoretical basis for the efficient production and application of exo-polysaccharides from Inonotus hispidus.