柱前衍生化HPLC法测定玉屏风复合多糖的单糖组成

Analysis of monosaccharide compositions of YuPingFeng Polysaccharides by HPLC pre-column derivatization

本研究旨在采用HPLC法测定玉屏风复合多糖(Yupingfeng polysaccharides,YPF-P)中的单糖组成。水提醇沉提取,AB-8大孔树脂纯化后的YPF-P样品用三氟乙酸溶液水解成单糖,然后将水解后的YPF-P样品与作为对照品的8种单糖D-甘露糖、鼠李糖、D-葡萄糖、D-核糖、D-半乳糖醛酸、D-半乳糖、阿拉伯糖用1-苯基-3-甲基-5-吡唑啉酮(PMP)柱前衍生化,经Wondasil C18(250 mm×4.6 mm,5μm)色谱柱分离,以磷酸盐缓冲溶液(p H值6.9)-乙腈(85∶15)进行洗脱,柱温40℃;流速1.0 mL/min;进样量20μL检测波长254 nm。结果表明,YPF-P是由D-甘露糖、鼠李糖、D-葡萄糖、D-核糖、D-半乳糖醛酸、D-半乳糖、阿拉伯糖组成的,物质的量比为4.64∶1∶5.88∶2.40∶120∶42.9∶69.5。研究结果表明,该方法灵敏度高、样品用量少、稳定性好、准确可靠,能够很好地分离测定YPF-P的各单糖组分。

This study aims to analyze the monosaccharide compositions of YuPingFeng Polysaccharides（ YPF-P） by HPLC.Yupingfeng polysaccharides were extracted by water and ethyl alcohol and then samples were purified by AB-8 macroporous resin,Yupingfeng polysaccharides were hydrolyzed into monosaccharides with trifluoroacetic acid,and then the derivatized samples and the 7 reference monosaccharides,mannose,rhamnose,glucose,ribose,galactose,glucuronic acid,galactose,and Arabia sugar were biochemcially analyzed using 1-phenyl-3-methyl-5-pyrazolone（ PMP）. The chromatographic separation was carried out on Wondasil C18（ 250 mm × 4. 6 mm,5μm） using phosphoric acid aqueous solution-acetonitrile（ p H6. 9） and acetonitrile（ 85∶ 15） as mobile phases at a flow rate of 1 mL/min kept at 40℃ through gradient elution. The detection wavelength was 254 nm. The results indicated that Yupingfeng polysaccharides was composed of mannose,rhamnose,glucose,ribose,galactose,uronic acid,galactose and Arabia sugar,and their mole ratio was 4. 64∶ 1∶ 5. 88∶ 2. 40∶ 120∶ 42. 9∶ 69. 5. The results also showed that this method had high sensitivity,low sample consumption,good stability and accuracy and reliabibity method. The method is simple and relatively inexpensive for detecting YuPingFeng Polysaccharides.