盐藻多糖单糖组成分析的四种色谱方法比较  

Comparison of four chromatographic methods for monosaccharide composition analysis of Dunaliella salina polysaccharide

作　　者：付海宁[1] 赵峡[1] 于广利[1] 陈娥功[2] 文松松[1] 

机构地区：[1]中国海洋大学医药学院教育部海洋药物重点实验室,山东青岛266003 [2]中国海洋大学兰太药业有限公司,山东青岛266100

出　　处：《中国海洋药物》2008年第4期30-34,共5页Chinese Journal of Marine Drugs

基　　金：国际科技合作资助项目(No.2007DFA30980)

摘　　要：目的通过对几种单糖组成分析的色谱方法进行比较,选择一种准确有效的方法测定盐藻多糖(PDS3)的单糖组成。方法分别采用薄层色谱(TLC)法、糖腈乙酸酯衍生气相色谱(AA-GC)法、离子排阻液相色谱(IEC)法及1-苯基-3-甲基-5-吡唑啉酮柱前衍生高效液相色谱(PMP-HPLC)法测定PDS3的单糖组成。结果TLC法和IEC法无需衍生化处理,操作简便、快捷,但各单糖间的分离效果较差,只适合于单糖组成较为简单的多糖样品的定性分析。AA-GC法测定单糖组成灵敏度高,各单糖具有良好的分离度,并可进行定量分析,与质谱联用还在PDS3中新发现了一种2-甲氧基戊糖,但AA-GC法不适合糖醛酸和氨基糖的分析;PMP-HPLC法不仅灵敏度高,分离效果好,而且还可同时检测PDS3中11种中性、酸性和碱性单糖,该法测得PDS3中各单糖的摩尔比为:Gal:Man:Ara:Glc:Rha:xyl:GlcUA:GalUA:GlcNAc:GlcN=19.3:10.6:6.9:1.6:1.1:1.0:12.9:6.4:4.1:1.6。另外,本实验还确定了PMP-HPLC法的最适衍生条件为:反应时间60min,反应温度70℃,NaOH与单糖的摩尔比为6:1,衍生试剂PMP与单糖的摩尔比为12:1。结论PMP-HPLC法较其它色谱方法更适合于PDS3这类复杂样品的单糖组成分析。Objective To find an accurate and effective method to determine the monosaccharide composition of polysaccharide from Dunaliella salina （PDS3） by comparing several chromatographic methods. Methods Thin layer chromatography （TLC）, aldononitrile acetate precolumn-derivatization gas chromatography （AA-GC）, ion exclusion chromatography （IEC） and 1-phenyl-3-methyl-5-pyrazolone precolumn-derivatization high performance liquid chromatography （PMP-HPLC） were used to determine the monosaccharide composition of PDS3. Results Both TLC and IEC were simple, rapid and easily performed without derivatization, however, the resolutions of monosaccharides were poor. They were only suitable to monosaccharide qualitative analysis of simple polysaccharide. AA-GC had high sensitivity and good resolution of neutral monosaccharides, and it was also suitable to monosaccharide quantitative analysis. A 2-methoxy pentose was found in PDS3 for the first time by GC-MS. However, AA-GC was not suitable to analysis of ketose, aminosaccharide and uronic acid. PMP-HPLC not only had high sensitivity and good resolution, but also it was suitable to simultaneous analyses of neutral, acidic and alkaline monosaccharides. The monosaccharide composition of PDS3 which determined by PMP-HPLC was Gat ： Man ： Ara ： Gle ： Rha ： Xyl ： GlcUA ： GalUA ： GlcNAc ： GlcN = 19.3 ： 10. 6 ： 6.9 ： 1.6 ： 1.1 ： 1.0 ： 12.9 ： 6.4 ： 4.1 ： 1.6. In addition, the optimum PMP derivation conditions were determined as follows： reaction time 60min, incubation temperature 70℃, ratio of NaOH to monosaccharide 6 ： 1, ratio of PMP to monosaccharide 12 ： 1. Conclusion PMP-HPLC is more suitable than other chromatographic methods to complicated monosaccharide composition analysis of polysaecharide such as PDS3.

关 键 词：盐藻多糖 单糖组成 薄层色谱法 糖腈乙酸酯衍生气相色谱法 离子排阻液相色谱法 柱前衍生高效液相色谱法 

分 类 号：Q949.2[生物学—植物学] Q539