响应面法优化中性蛋白酶提取苦竹花多糖及多糖性质分析

目的:采用响应面法优化中性蛋白酶辅助提取苦竹花多糖的最优条件,并对提取得到的粗多糖进行分离纯化、理化性质及体外抗氧化性测定。方法:在单因素试验基础上,考察提取时间、酶添加量、提取温度对苦竹花粗多糖得率的影响。利用响应面试验建立数学模型,确定最佳提取条件。苦竹花粗多糖经分离纯化,获取2个组分,分别命名为SBH-1和SBH-2,其中以SBH-1为主要组分。通过高效凝胶渗透色谱和红外光谱对SBH-1组分进行单糖组成、分子质量及初步的结构研究。通过与VC对比,研究苦竹花粗多糖SBH-1的体外抗氧化活性,即1,1-二苯基-2-三硝基苯肼(1,1-diphenyl-2-picrylhydrazyl,DPPH)自由基清除率、超氧阴离子自由基清除率及抗脂质过氧化能力的测定。结果:在中性蛋白酶添加量0.83%,提取温度46.68℃,提取时间115.95 min条件下,粗多糖得率为7.63%。经检测苦竹花粗多糖SBH-1组分分子质量为18.3 k D,主要由甘露糖(Man)、葡萄糖(Glc)、半乳糖(Gal)3种单糖组成比例为1.3∶2.3∶1.2。SBH-1对DPPH自由基和超氧阴离子自由基清除EC_(50)分别为2.03 mg/mL和1.038 mg/mL。结论:采用中性蛋白酶辅助提取得到的苦竹花粗多糖简单易行,纯化得到的中性杂多糖SBH-1具有很好的活性,为进一步研究和开发提供理论基础。

Optimization of Neutroproteinase-Assistant Extraction of Polysaccharide from the Stroma of Shiraia bambusicola Henn by Response Surface Analysis and Characterization of the Extracted Polysaccharide

Objective: This study aimed to optimize the neutroproteinase-assisted extraction of polysaccharides from the stroma of Shiraia bambusicola Henn using response surface methodology (RSM) and to investigate the separation, purification, characterization and antioxidant activity of the polysaccharides. Methods: The effects of extraction time, enzyme concentration and extraction temperature on the yield of polysaccharides were examined by using one-factor-at-a-time experiments. These extraction conditions were optimized through mathematical modeling. The crude extract was fractionated into two fractions: SBH-1 and SBH-2, with SBH-1 being among them. The monosaccharide composition, molecular mass and structure of SBH-1 were detected by high performance gel permeation chromatography (HPGPC) and infrared spectroscopy, respectively. The in vitro antioxidant activity was measured in comparison with VC by superoxide anion scavenging, 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging, and lipid peroxidation inhibition assays. Results: The maximum yield of crude polysaccharides of 7.63% was obtained at 115.95 min of extraction at 46.68 ℃ with 0.83% neutral proteinase. The molecular mass of SBH-1 was determined to be 18.3 kD, and its monosaccharide composition consisted of glucose, mannose, and galactose with a ratio of 1.3:2.3:1.2. The EC50 values for scavenging of DPPH and superoxide anion radials were 2.03 and 1.038 mg/mL respectively. Conclusions: The neutroproteinase-assistant extraction procedure was simple and easy to operate. SBH-1 was a neutral heteropolysaccharide with high activity. This study can provides a theoretical basis for further research and development of polysaccharides from S. bambusicola Henn.