The Research On Preparation,structural Identification And Biological Activity Of Polysaccharide From Stropharia Rugosoannulata(SR-1) And Polysaccharide From Tricholoma Lascivum(Fr.) Gillet(TLG-1)

Stropharia rugosoannulata belongs to Stropharia of Strophariaceae,which is one of the edible fungi recommended by FAO.At present,the research on Stropharia rugosoannulata mainly focuses on cultivation techniques and active component analysis.Among them,the research of polysaccharides from Stropharia rugosoannulata mainly focuses on antioxidant activity in vivo and in vitro.The detailed chemical structure and immunocompetence of it is few reports.Tricholoma lascivum(Fr.)Gillet belongs to Tricholoma of Tricholomataceae,which is a rare edible fungi.At present,the research on Tricholoma lascivum(Fr.)Gillet mainly focuses on taxonomic status and morphological characteristics.However,the preparation,structure elucidation and biological activity of polysaccharides from Tricholoma lascivum are few reports.In this study,homogenous polysaccharide of Stropharia rugosoannulata and Tricholoma lascivum(Fr.)Gillet collected from Xiaojin County of Aba of Sichuan Province were obtained by water boiling,alcohol precipitation and de-proteinization;The chemical structure of them was investigated by GC-MS,HPGPC,HPLC,FT-IR and NMR.immunological and antineoplasmic activity in vitro of the obtained polysaccharides were studied by modern biotechnology.On this basis,the antineoplasmic ability of polysaccharide from Tricholoma lascivum(Fr.)Gillet in vivo was further evaluated by establishment of S180 tumor-mice model.Illumina HiScq TM 200 technology was used for transcription sequencing of S180 tumor tissues.The analyses of differentially expressed genes and KEGG pathways of transcriptional datas were analyzed by DESeq2 and cluster profiler software to preliminarily explore the anti-tumor mechanism of polysaccharide from Tricholoma lascivum(Fr.)Gillet in vivo.The main contents and results are as follows:(1)The polysaccharides were isolated and purified from the fruiting bodies of Stropharia rugosoannulata and named SR-1.Its molecular weight was 13,281 Da,and consisted of galactose and glucose with a ratio of 3:1.It has a skeleton structure of(1,6)-α-D-Galp with branches at 2-O and the branches consisted of a(1→6,4)-β-D – Glcp and two →2)-α-D-Glcp.(2)A water-soluble polysaccharide was obtained from the fruiting bodies of Tricholoma lascivum(Fr.)Gillet by water boiling,alcohol precipitation and de-proteinization,and named TLG-1.FT-IR and NMR were used to investigate its structure.The results showed that TLG-1 was a kind of saccharide containing four sugar residues in a repetitive unit.(3)The research for immunological activity in vivo of SR-1 and TLG-1 showed that both polysaccharides possessed certain immunological activity.At a concentration range of 2.5-20 μg/mL,they could distinctly promote the growth of T lymphocyte,B lymphocyte and RAW264.7 cells,augment the phagocytic activity of RAW264.7 cells compared with the blank control group(P<0.01).SR-1 could also distinctly enhance the immunity by significantly promoting B lymphocytes to secrete IgA,IgD and IgG(P<0.05).(4)The research for antineoplasmic activity of SR-1 and TLG-1in vitro indicated that at a concentration of 2.5-20 μg/mL(P<0.01),they had significant inhibitory effects on colon cancer(CT26.WT)cells and gastric cancer cell(MFC)compared with the blank control group,but the degree of inhibition was different.At a concentration of 10 μg/mL,the inhibitory effect of SR-1 was the strongest with the inhibition rates of 34.03% and 33.67%,respectively.The inhibitory effect was better than that of TLG-1.TLG-1 also significantly inhibited the proliferation of S180 cells.(5)The evaluation for antineoplasmic activity in vivo of TLG-1 found that compared with the model group,TLG-1(20 mg/kg)could notablely inhibited the growth of S180 tumor in vivo,and the inhibition rate was 59%(P<0.01).Differential gene expression and pathway analysis of transcriptome revealed TLG-1 inhibited the growth of cancer cells through MAPK and PI3K-AKT signaling pathways.Through these studies,we could find a new way for the high-quality utilization of Stropharia rugosoannulata and Tricholoma lascivum(Fr.)Gillet,and provide theoretical basis and technical support for the further development and utilization of SR-1 and TLG-1.