柱前衍生化HPLC分析蒲公英多糖的单糖组成

Analysis on Monosaccharide Composition of Taraxacum Polysaccharides by Pre-column Derivatization HPLC

关键词:  柱前衍生化  高效液相色谱法  单糖  1-苯基-3-甲基-5-吡唑啉酮

Key words:  pre-column derivatization  HPLC  monosaccharide  1-phenyl-3-methyl-5-pyrazolone

摘要:

目的 合理优化衍生方法测定蒲公英多糖中单糖的组成及摩尔比。方法 采用1-苯基-3-甲基-5-吡唑啉酮(PMP)柱前衍生，用反相高效液相色谱法进行测定。色谱条件为：ZORBAX-C₁₈柱；流动相：0.1 mol·L^-1的磷酸盐缓冲液(pH 6.7)-乙腈(83∶17)；流速：1 mL·min^-1；检测波长：245 nm；进样量：20 μL；柱温：室温。结果 蒲公英多糖由D-鼠李糖、葡萄糖、D-半乳糖、D-木糖和D-阿拉伯糖组成，以不同方法提取的各单糖含量的摩尔比为：水提取法，Rha∶Glc∶Gal∶ Ara=0.088∶0.500∶0.190∶0.340；超声提取法，Rha∶Glc∶Gal∶Ara=0.050∶0.350∶0.080∶0.320；酶提取法，Rha∶Glc∶ Gal∶Ara=0.270∶0.630∶0.330∶0.460；超声协提取同酶法，Rha∶Glc∶Gal∶Ara =0.078∶0.550∶0.130∶0.170。结论 改进后的衍生化方法和等度洗脱的色谱方法具有操作简单、快速、重复性好等特点，可用于蒲公英多糖中单糖组成和摩尔比的测定。

Abstract:

OBJECTIVE To optimize the derivative method reasonably and detect monosaccharide components and molar ratio of taraxacum polysaccharides. METHODS Pre-column derivative reagent was 1-phenyl-3-methyl-5-pyrazolone(PMP), monosaccharide composition and molar ratio of taraxacum polysaccharides were detected by reversed-phase high performance liquid chromatography (RP-HPLC). Chromatographic conditions: ZORBAX-C18 column; mobile phase: 0.1 mol·L^-1 phosphate buffer solution (pH 6.7)-acetonitrile (83∶17); velocity: 1 mL·min^-1; detection wavelength: 245 nm; injection volume: 20 μL; column temperature: room temperature. RESULTS Taraxacum polysaccharides was composed of D-rhamnose, glucose, D-galactose D-xylose, D-Arabinose. The molar ratio of taraxacum polysaccharides which was extracted by different means was as forrows: water extraction method, Rha∶Glc∶Gal∶Ara=0.088∶0.500∶0.190∶0.340; ultrasonic extraction method, Rha∶Glc∶Gal∶Ara=0.050∶0.350∶0.080∶0.320; cellulose extraction method, Rha∶Glc∶Gal∶Ara=0.270∶0.630∶0.330∶ 0.460; synergistic ultrasonic extraction and cellulose extraction, Rha∶Glc∶Gal∶Ara=0.078∶0.550∶0.130∶0.170. CONCLUSION The improved derivative method and the chromatographic conditions of isocratic elution were simple, rapid and with a good reproducibility, and can be used to detect monosaccharide components and molar ratio of taraxacum polysaccharides.