Study On The Isolation And Anti-inflammatory Activity Of A Kind Of Wild Ganoderma Lucidum Soaked Water Polysaccharide

Object and Significance:Meat Ganoderma lucidum,also known as"Tai Sui",has a long history in my country.There are related reports and researches that Meat Ganoderma lucidum has antibacterial effects and improves immunity.Meat Ganoderma lucidum from different sources not only has different forms and functions,but also different There are obvious differences in the microbial groups and dominant populations of meat Ganoderma lucidum found in the environment,so their activities are not necessarily the same.In this paper,the soaking water of a wild Ganoderma lucidum with a weight of 900 kg provided by Sichuan Guipu Co.,Ltd.was used as the research object.The immersion water was subjected to physical and chemical determinations in order to understand its effective components,extract and purify it,and explore the meat Ganoderma lucidum.The anti-inflammatory activity characteristics of soaking water are expected to be applied to anti-inflammatory health foods and medicines.Research content:1.The physical and chemical determination of the soaking water of Ganoderma lucidum and the extraction and separation of polysaccharidesPhenol-sulfuric acid method was used to determine the content of crude polysaccharides in the soaking water of Ganoderma lucidum,NBT-Gly method was used to determine the content of pyrroloquinoline quinone(PQQ)in the soaking water of Ganoderma lucidum,elemental analysis was carried out in the soaking water of Ganoderma lucidum.The refined sugar is subjected to elemental analysis and molecular weight determination;the meat Ganoderma lucidum soaked raw sugar is extracted by water extraction and alcohol precipitation,the protein is removed by the Sevag method,and the protein-removed refined sugar is purified and graded by DEAE Sepharose Fast Flow exchange column chromatography for subsequent experiments.The obtained meat Ganoderma lucidum soaking water contains protein,fat,oligopeptides and other active substances,the polysaccharide content is 358.46±14.59?g/m L,the PQQ content is10.3±0.76?g/m L,and the molecular weight of the meat Ganoderma lucidum immersion water refined sugar is 2.6×10~6Da,the meat ganoderma lucidum soaked in water contains N,C,H and S elements.After multiple extraction and purification of the meat Ganoderma lucidum soaking water,five samples were obtained by freeze-drying after dialysis:4 polysaccharide samples(raw polysaccharides,refined sugar,neutral sugar,acidic sugar)of the meat Ganoderma lucidum soaking water and protein,Plus the tissue blocks and soaking water that produce the meat Ganoderma lucidum soaking water,a total of 7 samples.2.Verification of in vitro cellular anti-inflammatory activity of Ganoderma lucidum soaked with hydropolysaccharideLPS(lipopolysaccharide)was used to induce macrophage RAW264.7 to establish an inflammation model.The CCK-8 method was used to determine the effects of seven samples obtained from different treatments of meat ganoderma lucidum on cytotoxicity;the CCK-8method was used to determine the effects of LPS on macrophages The toxicity of cell RAW264.7 and the influence of cell inflammation model were detected by using ELISA to detect the effects of polysaccharides obtained from different treatments of meat Ganoderma lucidum on the NO and inflammatory factors(TNF-?,IL-)in LPS-induced macrophage RAW264.7 medium.1β,IL-10)content.The 7 samples obtained from different treatments of meat ganoderma lucidum soaked in water are non-toxic to macrophages RAW264.7 between 0.01?g/m L-700?g/m L,and the optimal condition for the cytoinflammatory model is 1μg/m L LPS acts on macrophages RAW264.7 for24h;under these conditions,in addition to meat ganoderma lucidum soaking water protein,the obtained meat ganoderma lucidum soaking water polysaccharide samples can significantly reduce the LPS-induced macrophage production in RAW264.7 medium The content of NO,inflammatory factors TNF-?and IL-1βcan significantly increase the content of anti-inflammatory factor IL-10,which proves that the meat ganoderma lucidum soaked in hydropolysaccharide has an anti-inflammatory effect at the cellular level in vitro.3.Verification of the anti-inflammatory activity of meat ganoderma lucidum soaked in water polysaccharides in vivoThe SD rats were divided into a blank control group,a negative control group,a positive control group,and a Ganoderma lucidum soaked hydropolysaccharide experimental group.The rats were modeled with deep second-degree scalds.The scalded rats were measured at 0h,12h,24h,3d,and 7d.Blood indicators,and serum inflammatory factors(TNF-?,IL-1β,IL-10)content.The three polysaccharide samples obtained by different treatment methods of meat ganoderma lucidum soaking water(meat ganoderma lucidum soaking water raw sugar,meat ganoderma lucidum soaking water refined sugar,and meat ganoderma lucidum soaking water)can significantly reduce the inflammatory factor TNF-?,IL-1βcontent,can significantly increase the content of anti-inflammatory factor IL-10,at the same time,can significantly reduce the number of monocytes,neutrophils,significantly increase the number of lymphocytes,3 kinds of meat Ganoderma lucidum soaking water Polysaccharides have anti-inflammatory effects in animals.