柱前衍生HPLC法分析烟叶水溶性多糖的单糖组成  

Analysis of Monosaccharide Composition of Tobacco Water-soluble Polysaccharides by Pre-column Derivatization HPLC

作　　者：赵妍 黄延俊 刘秀彩 黄华发 刘江生 叶仲力 张廷贵 刘泽春 谢卫 李耀群[2] ZHAO Yan;HUANG Yanjun;LIU Xiucai;HUANG Huafa;LIU Jiangsheng;YE Zhongli;ZHANG Tinggui;LIU Zechun;XIE Wei;LI Yaoqun(Technology Center of China Tobacco Fujian Industrial Co.,Ltd.,Xiamen 361021,China;College of Chemistry and Chemical Engineering,Xiamen University,Xiamen 361005,China)

机构地区：[1]福建中烟工业有限责任公司技术中心,厦门361021 [2]厦门大学化学化工学院,厦门361005

出　　处：《中国烟草科学》2025年第2期52-59,共8页Chinese Tobacco Science

基　　金：福建中烟工业有限责任公司博士后项目(FJZYKJJH2023BH002)。

摘　　要：建立了1-苯基-3-甲基-5-吡唑啉酮(PMP)柱前衍生-高效液相色谱法测定烟叶水溶性多糖单糖组成及含量的方法,采用完全酸水解法水解烟叶水溶性多糖,水解产物经PMP衍生化反应,采用反相高效液相色谱法测定烟叶水溶性多糖经酸解后的单糖衍生物信号,并将其方法用于探究不同产地及类型烟叶水溶性多糖的单糖组成及含量。经优化后的检测条件如下:色谱柱为安捷伦Eclipse XDB-C18(250 mm×4.6 mm,5μm);0.05 mol/L磷酸盐(KH_(2)PO_4-NaOH,pH 7.0)缓冲液为A流动相,乙腈为B流动相,洗脱条件为84∶16等度洗脱;紫外检测波长为250 nm。结果表明,9种单糖在20.0~800μg/mL范围内线性关系良好,相关系数R~2>0.992 7,检出限(LOD)为0.07~0.17μg/mL,定量限(LOQ)为0.22~0.56μg/mL,平均回收率为82.3%~104.0%,经柱前衍生HPLC法测定烟叶水溶性多糖由甘露糖(Man)、鼠李糖(Rha)、半乳糖(Gal)、阿拉伯糖(Ara)、葡萄糖(Glc)、葡萄糖醛酸(GlcUA)、半乳糖醛酸(GalUA)等组成,其中Glc和GalUA占较高比例,不同产地和类型烟叶水溶性多糖的单糖组成比例存在显著差异。该方法快速简单、灵敏度高、重复性好,可用于烟叶多糖的单糖组成分析,同时为进一步研究烟叶多糖的结构提供基础。A determination method was established for the monosaccharide composition and content of tobacco water-soluble polysaccharides by 1-phenyl-3-methyl-5-imidazolinone(PMP)pre-column derivatization high performance liquid chromatography.Complete acid hydrolysis method was employed to hydrolyze tobacco water-soluble polysaccharides,and the hydrolysis products were subjected to PMP derivatization reaction.The signal of monosaccharide derivatives of tobacco water-soluble polysaccharides after acid hydrolysis was determined by reverse phase high-performance liquid chromatography.The method was used to explore the monosaccharide composition and content of tobacco water-soluble polysaccharides of different regions and types.The optimized detection conditions were as follows:0.05 mol/L phosphate(KH2PO4-NaOH,pH 7.0)buffer(solvent A)and acetonitrile(solvent B)were chosen as the mobile phase of elution system at the isocratic elution condition(84∶16),with Agilent Eclipse XDB-C18 column(250 mm×4.6 mm,5μm);the UV detection wavelength was 250 nm.Results showed that the linear relationship is good for the nine monosaccharides in the concentration range of 20.0-800μg/mL,with the correlation coefficient(R2)>0.9927.The limits of detection and quantification were 0.07-0.17 and 0.22-0.56μg/mL,respectively.The average recoveries ranged 82.3%-104.0%in spiked samples.The tobacco water-soluble polysaccharides were composed of mannose(Man),rhamnose(Rha),galactose(Gal),arabinose(Ara),glucose(Glc),glucuronic acid(GlcUA),and galacturonic acid(GalUA),with Glc and GalUA accounting for a higher proportion.There were significant differences in the monosaccharide composition of tobacco water-soluble polysaccharides from various regions and types.The method is fast,simple,highly sensitive,and has good repeatability,thus can be used for monosaccharide composition analysis of tobacco polysaccharides.At the same time,it provides a basis for further research on the structure of tobacco polysaccharides.

关 键 词：烟叶水溶性多糖 柱前衍生化 液相色谱 单糖组成 

分 类 号：TS411[农业科学—烟草工业]