基于柱前衍生-HPLC法研究焦栀子炮制过程单糖组成及含量变化  

Composition and content change of monosaccharide in Gardeniae FructusPraeparatusduring heat processing based on pre-column derivatization HPLC method

作　　者：乔梦如 王云[2] 贾哲 刘艳萍 宋亚南 张村[1,2] QIAO Meng-ru;WANG Yun;JIA Zhe;LIU Yan-ping;SONG Ya-nan;ZHANG Cun(College of Pharmacy,HenanUniversity of Chinese Medicine,Zhengzhou 450008,China;Institute of Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,China)

机构地区：[1]河南中医药大学药学院,河南郑州450008 [2]中国中医科学院中药研究所,北京100700

出　　处：《中草药》2023年第7期2096-2105,共10页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金项目(81873010);国家自然科学基金项目(82173979);中国中医科学院科技创新工程(CI2021A04204);国家中医药管理局中药炮制技术传承基地项目(国中医药科技中药[2022]59号)。

摘　　要：目的建立柱前衍生-HPLC法测定焦栀子炮制过程样品中8种单糖(甘露糖、鼠李糖、葡萄糖醛酸、半乳糖醛酸、葡萄糖、半乳糖、木糖、阿拉伯糖)的含量测定方法,通过分析比较总单糖、结合型单糖和游离单糖的组成及差异,为揭示焦栀子炒制过程颜色褐变机制奠定基础。方法以Phenomenex Luna C18柱(250 mm×4.6 mm,5μm)为色谱柱,流动相为100 mmol/L乙酸铵-乙腈(83∶17),柱温35℃,体积流量1.0 mL/min的条件下,分别对酸水解糖提取方法、游离糖提取条件进行优化,衍生化后样品进行方法学考察。结果酸水解方法以1.5 mol/L硫酸、100℃水解4.0 h最佳,游离单糖为超声处理15 min。焦栀子炮制过程样品酸水解产物检测出甘露糖、鼠李糖、葡萄糖醛酸、半乳糖醛酸、葡萄糖、半乳糖、木糖、阿拉伯糖共8种单糖;游离单糖检测出甘露糖、葡萄糖和半乳糖。经方法学验证,8种单糖的精密度、重复性、稳定性、加样回收率均符合方法学要求;酸水解得到的总单糖在种类上、含量上均高于游离单糖,总单糖和结合型单糖部分糖变化基本一致,甘露糖含量呈先降后升的趋势,半乳糖醛酸含量呈降低趋势,总单糖葡萄糖含量呈先升后降趋势,结合型葡萄糖含量呈升高的趋势,其他糖的变化幅度不大。随着炒制程度的加重,过程饮片的结合型单糖的组成类别不变,除结合型半乳糖醛酸外,各结合型单糖比例基本不变。游离葡萄糖和游离半乳糖含量随炮制程度的加深呈降低的趋势,其中游离半乳糖在焦栀子中未检出。结论游离葡萄糖、游离半乳糖和结合型半乳糖醛酸是焦栀子炮制过程中变化最明显的单糖,可为栀子炒焦过程颜色褐变机制的反应物提供参考。ObjectiveReversed-phase-high-performance liquid chromatography(RP-HPLC)precolumnderivatization was used to determine the monosaccharide composition and content in Jiaozhizi(Gardeniae FructusPraeparatus,GFP)during heat processing.By analyzing and comparing the composition and difference of total monosaccharides,binding monosaccharides and free monosaccharides,the basis was laid for revealing the mechanism of color browning during the heating process of GFP.Methods Chromatographic separation was carried out on a Phenomenex Luna C_(18)column(250 mm×4.6 mm,5μm)with the mobile phase of 100 mmol/L ammonium acetate-acetonitrile(83:17)at the flow rate of 1.0 mL/minand the column temperature of 35℃.Sample pretreatment conditions were optimized by investigating the effect of extraction solvents and methods on the extraction efficiency of monosaccharide used for HPLC quantification.Results Total monosaccharides was measured after hydrolysis in 1.5 mol/L sulphuric acid at 100℃for 4.0 h and free monosaccharide was measured after ultrasonic in water for 15 min.Eight total monosaccharides,including mannose,rhamnose,glucuronic acid,galacturonic acid,glucose,galactose,xylose and arabinose,athree free monosaccharides,including mannose,glucose and galactose,were detected in the GFPduring heat processing.The developed HPLC method for the determination of eight monosaccharides was validated in terms of linearity,precision,reproducibility,stability,and recovery of the eight monosaccharide.The total monosaccharides obtained by acid hydrolysis was higher than free monosaccharides in terms of variety and content.The change of total monosaccharide and some sugars of conjugated monosaccharides were basically the same.The mannose content showed a trend of decreasing first and then increasing,the galacturonic acid content showed a trend of decreasing,the total monosaccharose glucose content showed a trend of increasingfirst and then decreasing,the bound glucose content showed a trend of increasing,other sugars changed little.With the a

关 键 词：焦栀子 单糖 柱前衍生化 HPLC 甘露糖 鼠李糖 葡萄糖醛酸 半乳糖醛酸 葡萄糖 半乳糖 木糖 阿拉伯糖 

分 类 号：R283.6[医药卫生—中药学]