高效液相色谱法测定四种植物多糖中单糖方法的建立

Determination of Monosaccharides in Four Plant Polysaccharides by High Performance Liquid Chromatography

摘要: 采用高效液相色谱法测定四种植物多糖的单糖组成。用超声辅助提取大豆、香菇、木耳、枸杞多糖，通过三氟乙酸水解后，用1-苯基-3-甲基-5-吡唑啉酮(PMP)柱前衍生–液相色谱法检测，并选用专属性，精密度，灵敏度，线性范围和加标回收率对此方法进行验证，最后对大豆、香菇、木耳、枸杞多糖中单糖的组成进行分析。结果表明，待测定的各单糖组分在1.03~550 μg/mL范围内呈良好的线性关系，且各组分分离度大于1.5，检出限大于2.01 μg/mL，定量限大于6.43 μg/mL，精密度与稳定性的相对标准偏差均小于4%，加标回收率在98.0%~101.4%之间，验证结果显示该方法简单快速、准确可靠。通过应用此方法测定大豆、香菇、木耳、枸杞多糖的单糖组成，结果显示大豆多糖属于以半乳糖为主的杂多糖，且半乳糖含量为53.40%；香菇和枸杞多糖属于以葡萄糖为主的杂多糖，所含的葡萄糖含量分别为45.74%和21.07%；木耳多糖属于以D-甘露糖为主的杂多糖，且D-甘露糖含量为18.64%。

Abstract: The monosaccharide compositions of four plant polysaccharides were determined by high performance liquid chromatography (HPLC). Polysaccharides from soybeans, shiitake mushrooms, black fungi, and wolfberries were extracted by ultrasonic assistance. After hydrolysis with trifluoroacetic acid, the samples were detected by pre-column derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP) and HPLC. The method was validated using specificity, precision, sensitivity, linear range, and standard addition recovery rate. Finally, the monosaccharide compositions in polysaccharides from soybeans, shiitake mushrooms, black fungi, and wolfberries were analyzed. The results showed that the monosaccharide components to be determined showed a good linear relationship in the range of 1.03~550 μg/mL, and the resolution of each component was greater than 1.5, the detection limit was greater than 2.01 μg/mL, and the limit of quantification was greater than 6.43 μg/mL. The relative standard deviation of precision and stability was less than 4%, and the recovery of standard addition was between 98.0%~101.4%. The verification results show that the method is simple, rapid, accurate and reliable. The monosaccharide composition of soybean, shiitake mushroom, fungus and medlar polysaccharide was determined by applying this method. The results showed that soybean polysaccharide belongs to galactose-based heteropolysaccharide and the galactose content was 53.40%. The heteropolysaccharide contained 45.74% and 21.07%, respectively; the fungus polysaccharide belonged to D-mannose-based heteropolysaccharide, and the D-mannose content was 18.64%.