Extraction And Isolation Of Sweet Potato Glycoprotein SPG-56 And Study On Its Anti-colon Cancer Activity

Objective:In this paper,sweet potato were used as raw materials.The aim is to extract and isolate new active ingredients from them,and to identify the structure of the active ingredients.Then evaluate their anti-tumor activity and mechanism,so as to provide theoretical basis for clinical research and product development.Methods and Results:1.The SPG-56 was obtained by water extraction,isoelectric precipitation,chromatography,electrophoresis and MTT activity tracking.The compositions of sugar and protein in SPG-56 were identified with high performance liquid chromatography and amino acid analyzer,and the antitumor activities were explored using BALB/c nude mice and human colon cancer cells(HCT-116).The results showed that the molecular weight of sweet potato glycoprotein is 56 KD and we named this compound SPG-56.The relative sugar content of SPG-56 is 2.9%.Whereas the relative protein content of SPG-56 is 97.1%.The SPG-56 comprise more than 6 types of sugars and 15 kinds of amino acids.The types of monosaccharides was up to six,consist of mannose,glucuronic acid,galacturonic acid,xylose,galactose,and arabinose.The contents of these monosaccharides were about 0.21%,0.17%,0.20%,0.23%,0.82%and 0.92%,respectively.The essential amino acid was up to 42.9%,while glutamic acid,aspartic acid and valine were abundant in SPG-56,and their contents were about 14.2%,14.0%and 9.5%,respectively.2.In the vitro experiments,SPG-56 was used to study the colon cancer cells HCT-116 cells proliferation inhibiton by MTT assay.Based on the results of MTT assay,flow cytometry was used to detect the apoptosis rate of HCT-116 after SPG-56treatment.Studies showed that SPG-56 can inhibit the growth of tumor cells by inducing apoptosis and 80μg/mL SPG-56 also induced the apoptosis of HCT-116 cells up to 33.6%.3.The antitumor activities were explored using BALB/c nude mice and human colon cancer cells(HCT-116).The results indecated that SPG-56 significantly inhibited tumor growth after gavage at a dosage of 150 mg/kg.And the tumor volume of SPG-56(373.3±25.3 mm~3)was significantly lower than that of the tumor control(827.6±53.4 mm~3).Further more Western blot was used to analyze the effect of SPG-56 on the expression of protein and gene.Studied showed that SPG-56 can promote the expression of Casepase3,Casepase8,Casepase9 and Bax at a dose-dependent behavior.And inhibit the expression of Bcl-2.At the same time,the results of immunohistochemistry were consistent with that of WB.All of these results indecated that SPG-56 would potentially delay the development of colon cancer.4.Based on the above results,anti-colon cancer mechanism of SPG-56 were conducted for the futher study.16s DNA sequence analysis and short chain fatty acid nanlysis were used to study the intestinal microbe of nude mice treated with SPG-56.Animal experiments on male BALB/c nude mice indicated that orally administration of SPG-56(150 mg/kg),Lactobacillus fermentum(2×10~8 cfu/0.2 mL)+SPG-56(150mg/kg)and shot-chain acids(SCFAs 20 mM/0.2 mL)could obviously suppress the tumor growth in contrast with tumor control(TC)group.Results of 16S DNA sequence analysis showed that the microbial species was enriched in SPG-56 group and the abundance of LF in NC group was significantly higher than in TC group.The content of SCFAs in the SPG-56 group was significantly higher than that in other groups,and SCFAs and LF+SPG-56 can significantly inhibit the proliferation of tumor cells.In summary,we consisder that SPG-56(a new glycoprotein isolated from sweet potato)has a certain anti-colon cancer effect,and it can inhibit the growth of animal tumors to some extent,improve intestinal species richness,increase the content of Lactobacillus fermentum,and inhibit the expression of apoptosis related proteins and genes,and induce apoptosis in tumor cells.Therefore,SPG-56 can be used as an anti-tumor drug,and it is worth further study so as to provide a theoretical basis for clinical.