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目的研究用酸-碱法制备白色念珠菌细胞壁酸碱不溶性β-葡聚糖,并对其物理化学性质和结构进行分析,获得结构明确的酸碱不溶性(1,3)-β-D-葡聚糖。方法用沙氏肉汤培养白色念珠菌,经离心收集和干燥后,先后用NaOH溶液和乙酸进行处理,提取酸碱不溶性葡聚糖,采用薄层层析分析、高碘酸氧化分析和红外光谱法测定多糖的基本结构,并计算用该种方法提取的酸碱不溶性(1,3)-β-D-葡聚糖的提取效率。结果与标准的(1,3)-β-D-葡聚糖红外光谱相比,提取的多糖红外光谱在890 cm-1有特征吸收峰,表明多糖为β-构型;经苯酚-硫酸法鉴定,样品显桔红色,为多糖阳性反应。多糖经薄层层析分析,比较样品与单糖标准品的Rf值,确定多糖的构型为D-型;多糖经高碘酸氧化后,测定消耗高碘酸量和甲酸生成量,从而测定其多糖的主链键型为1,3位键合。经计算酸-碱法提取的酸碱不溶性(1,3)-β-D-葡聚糖效率为20.9%。结论用酸-碱法可以从白色念珠菌细胞壁中提取到高纯度的酸碱不溶性(1,3)-β-D-葡聚糖,为建立新型高特异(1,3)-β-D-葡聚糖的检测试剂提供关键物质基础。
OBJECTIVE To study the preparation of acid-base insoluble β-glucan from cell wall of Candida albicans by acid-base method and to analyze the physicochemical properties and structure of Candida albicans cell wall. The structure-specific acid-base insoluble (1,3) -β-D- Glycan. Methods Candida albicans was cultured with Sabouraud broth, collected by centrifugation and dried. The solution was treated with NaOH solution and acetic acid, and then the acid - base insoluble glucan was extracted. TLC analysis, periodic acid oxidation analysis and FTIR Method to determine the basic structure of polysaccharides and calculate the extraction efficiency of the acid-base-insoluble (1,3) -β-D-glucan extracted by this method. Results Compared with the standard (1,3) -β-D-glucan infrared spectrum, the infrared spectrum of the extracted polysaccharide showed a characteristic absorption peak at 890 cm-1, indicating that the polysaccharide was β-configuration. Identification, the sample was orange, polysaccharide positive reaction. Polysaccharides were analyzed by TLC, comparing the Rf value of the sample with the monosaccharide standard to determine that the polysaccharide was D-type; the polysaccharide was oxidized by periodate, and then the amount of periodic acid consumed and the amount of formic acid generated were measured, The main chain of its polysaccharide 1,3-bond. The calculated acid-base extraction of acid-base insoluble (1,3) -β-D-glucan was 20.9%. Conclusion Acid-base-insoluble (1,3) -β-D-glucan with high purity can be extracted from Candida albicans cell wall by acid-base method. Dextran test reagents provide a key material basis.