败酱草多糖的超声波协同复合酶提取、结构分析及抗菌活性研究

Studies on the Ultrasonic Composite-enzyme Extraction,Structure and Antibacterial Activity of Herba Patriniae Polysaccharide

目的在单因素实验基础上,运用正交试验优选超声波协同复合酶法提取败酱草多糖的工艺,并通过仪器分析和体外抑菌试验初步分析败酱草多糖的结构及其抗菌活性。方法分别考察酶添加量、料液比、超声功率及时间对多糖提取率的影响,采用四因素三水平正交试验进行多糖提取工艺的优化,利用高效凝胶色谱法、气相色谱法及紫外和红外光谱法分析其结构。结果败酱草多糖的最佳提取工艺:提取温度55℃,pH值为6.5,酶添加量2.0%,料液比1∶20,超声功率90 W,超声时间20 min。在此条件下,败酱草多糖的提取率为(25.01±0.15)%,多糖质量分数达(44.12±0.14)%。经凝胶柱层析纯化后紫外光谱分析显示败酱草多糖的纯度较高,红外光谱和气相色谱分析表明败酱草多糖为α-和β-糖苷键的呋喃型糖苷环骨架,主要由葡萄糖、甘露糖、半乳糖、阿拉伯糖、鼠李糖和木糖按照摩尔比1∶0.5∶0.9∶1.1∶0.3∶0.3组成,重均分子量为1.26×10;。抑菌试验显示败酱草多糖对肺炎克雷伯菌和恶臭假单胞菌具有一定的抗菌活性,而对蜡状芽孢杆菌和藤黄微球菌基本没有抗菌作用。结论超声波协同复合酶法提取效率高,多糖含量高,提取的败酱草多糖具有一定的抗菌活性。

OBJECTIVE On the basis of single factor experiment,to optimize the extraction process of polysaccharide from Herba Patriniae by ultrasonic composite-enzyme method by orthogonal experiment.To analyze and evaluate the preliminary structure and the antibacterial activity of Patriniae polysaccharide through instrumental analysis and antibacterial test in vitro,respectively.METHODS The effects of enzyme dosage,solid-liquid ratio,ultrasonic power and time on the extraction rate of polysaccharides were investigated by the four-factor,three-level orthogonal test,while the structure of polysaccharide was analyzed using high-performance gel permeation chromatography,gas chromatography,ultraviolet and infrared spectroscopy.RESULTS The optimum extraction conditions of Patriniae polysaccharides were as follows:extraction temperature 50℃,pH=6.5,enzyme dosage 2.0%,solid-liquid ratio 1∶20,ultrasonic power 90 W,ultrasonic time 20 min.Under the optimum conditions,the extraction yield of Patriniae polysaccharides was(25.01±0.15)%,while the concentration of total carbohydrate was(44.12±0.14)%.Ultraviolet spectroscopy analysis showed that the purity of Patriniae polysaccharide was higher after purification by gel column chromatography.Infrared spectroscopy and gas chromatography analysis showed that Patriniae polysaccharide was a furan-type glycoside ring skeleton withα-andβ-glycosidic bond,and was mainly composed of glucose,mannose,galactose,arabinose,rhamnose and xylose at the molar ratio of 1∶0.5∶0.9∶1.1∶0.3∶0.3,with the average molecular weight 1.26×10~5.Antibacterial test in vitro showed that the Patriniae polysaccharide had certain antibacterial activity against Klebsiella pneumonia(K.pneumonia)and Pseudomonas putida(P.putida),but no antibacterial effect on Bacillus cereus(B.cereus)and Micrococcus luteus(M.luteus).CONCLUSION Ultrasonic-composite enzyme is an efficient extraction method,and the extracts rich in polysaccharide,with certain antibacterial activity.