Optimization of Multi-enzymatic Extraction of Polysaccharide from Anemarrhena asphodeloides Bunge and Its Immunomodulatory Activity

In this study, the extraction process and immune activity of polysaccharide from Anemarrhena asphodeloides Bunge were studied, in order to provide a theoretical basis for its development and utilization. Crude polysaccharide was obtained by complex enzyme method from Anemarrhena asphodeloides Bunge. On the basis of orthogonal experiment to determine compound enzyme ratio, the extraction conditions of Anemarrhena asphodeloides Bunge polysaccharide by enzyme complex were optimized by using response surface methodology and mono-factor experiments, and the optimum technological conditions were obtained. Four polysaccharide components, APSE-0, APSE-1, APSE-2 and APSE-3, were isolated and purified by DEAE-52 cellulose chromatography column. The effects of the polysaccharide fractions on the proliferation of RAW264.7 cells were determined by MTT assay, and the release capacity of nitrogen monoxide NO was determined by the Griess kit. The results showed that compound enzyme ratio was papain 16000 U/g, cellulose 1200 U/g, and pectinase 1600 U/g. The optimum extraction conditions were hydrolysis time of 2 h, liquid-solid of 15:1 (mL/g), temperature of enzymatic hydrolysis of 52 ℃. Under these conditions the extraction yield was (10.58%±0.03%). The results of in vitro immunoassay proved that four polysaccharides could significantly promote the proliferation of RAW246.7 cells, and APSE-0, APSE-2 and APSE-3 significantly induced NO production, among which APSE-2 showed the strongest immunoregulatory activity. APSE-2 could be explored as a potential source for immunomodulatory agents of the functional foods or dietary complementary for people with compromised immune system.