Researches On The Extraction Technology, Property And Anti-tumor Activity Of Polysaccharides From Atractylodes Chinensis

Objective:In this paper, extraction, separation, purification, the monosaccharide composition, anti-tumor activity bioactivity of Atractylis polysaccharide was studied. Partially physical and chemical nature, molecular weight, monosaccharide composition and their connections mode of Atractylis polysaccharide after it was purified. The purpose was to provide theoretical foundation for the industrialized production and provide experimental evidence for clinical application of Atractylis polysaccharide.Methods:1. Content and purity of Atractylis polysaccharide were taken as evaluating norms, studied enzymolysis-assisted(cellulase, hemicellulase, pectinase and complex enzymatic) ultrasonic extraction of Atractylis polysaccharide extraction yield, the best combination of enzymolysis was optimize by orthogonal experimental design.2.We optimize the purification technology including sevage method, trichloroacetic acid(TCA) method, TCA-n-butyl alcohol method and of Atractylis polysaccharide on the basis of the content of polysaccharides, as well as the removal ratio of protein. DEAE-Sepharose Fast Flow column and SephadexG-150 gel column is used to isolate and purify the Atractylis polysaccharide inspect the purity of its separate components.3.The content of uronic acid was determined by the method of m-hydroxydiphenyl. The properties and composition analysis, molecular weight was studied by IR spectra, GPC, and GC. And then MTT and trypan blue was employed to detect inhibition rate of ovarian cancer cell line Skov3, Hela, hepg2 liver cancer cell lines and 7721.Results:1.The best combination of enzymolysis was cellulase and pectinase (ratio of 1:1). The optimal enzymolysis extraction conditions was as follows:temperature was 50 ℃, content was 1.2(g/g), pH was 5, and time was 50 min. Under this conditions, the purity of polysaccharide was 32.29%. Only one component was obtained from DEAE-Sepharose Fast Flow column and SephadexG-150 gel column chromatography, the purity of polysaccharide is 92.68%.2.The refined polysaccharide was nomogoneous component, white powder, which dissolved in water easily, the polysaccharide content was 92.68%, protein content was 0.8%, uronic acid content were 4.36% and 4.07% of ACCPS and ACPS, the molecular weight was 11171 Da. The composition of Atractylis polysaccharide was only fructose, and connected with 2â†'1 way.3.Atractylis polysaccharide had the obvious inhibitory effect on Hela, liver cancer cell line hepg2 and 7721, especially to hepg2, proliferation inhibition rate was reach up to 87.40%.Conclusion:In this study, the different enzymes were used to enzymolysis Atractylodes chinensis, and then polysaccharide was extracted by ultrasonic method. The extraction technology was reviewed for the content of polysaccharide as index. After this step, selected cellulase and pectinase enzyme (ratio of 1:1) to zymolysis Atractylodes chinensis. Studied the impact of enzymolysis temperature, enzyme content, enzymolysis pH, and enzymolysis time on the Atractylis polysaccharide content, optimized the best enzyme hydrolysis. The yield of Atractylis polysaccharide was 32.29% under the optimum conditions-higher than control group which wasn’t adding an enzyme. The stability and reappearance of the optimized technology preparation is good.The deproteinization effect was optimal with TCA method, which has a higher protein extractive coefficient and content of polysaccharides. We obtained one component by DEAE-Sepharose Fast Flow column and SephadexG-150 gel column chromatography, the purity of polysaccharides was 92.68% and has a good anti-tumor activity. The composition of Atractylis polysaccharide was fructose (98.2%)and a small amount of glucose (1.8%), and connected with 2â†'1 way. Comprehensive methylation analysis and NMR spectra, ACPS structure can be expressed as:α-D-G1cp-(1â†'2)-[β-D-Fruf-(1â†'2)-β-D-Fruf-]n-(1â†'2)-β-D-Fruf, Atractylis polysaccharide could inhibit the proliferation of human ovarian cancer cell Lines Skov3, liver cancer cell line hepg2 and 7721, Hela cells in vitro.The inhibitory rate on hepg2 was 87.40%. This study provides the experimental basis and reference forclinical usage of Atractylis polysaccharide.