柱前衍生超高效液相色谱-串联四极杆质谱法测定仙草多糖组成及其含量

摘要

建立了测定仙草多糖组成及其含量的超高效液相色谱-串联四极杆质谱分析方法.仙草样品在碱性条件下用沸水提取,提取液经固相萃取小柱净化后加三氟乙酸在110℃水解,然后采用1-苯基-3-甲基-5-吡唑啉酮(PMP)衍生.以Waters ACQUITY UPLC BEH C18(2.1mmi.d.×50mm,1.7 μm)为分析柱,乙腈和缓冲盐溶液(0.5 mmol/L乙酸铵-0.05％乙酸)为流动相进行梯度洗脱分离,流速0.5 mL/min,电喷雾正离子多反应监测模式检测,内标法定量.结果显示8种单糖在1～ 100 μ mol/L浓度范围内呈良好的线性关系,相关系数r2均大于0.96,方法的回收率为84％ ～ 112％,相对标准偏差(RSD)不高于4.7％.仙草多糖由甘露糖、鼠李糖、核糖、葡萄糖醛酸、半乳糖醛酸、葡萄糖、半乳糖和木糖8种单糖组成,其摩尔百分比为7.4％、5.7％、4.2％、0.9％、28.4％、26.5％、16.4％和10.6％.该法简单、快速、灵敏高、重现性好,可用于仙草多糖的单糖组成分析和含量测定.%An ultra-high performance liquid chromatography - tandem quadrupole mass spectrometric method was developed for the determination of monosaccharide composition and content in Mesona Chinensis Benth polysaccharides. Mesona Chinensis Benth Polysaccharides were extracted with hot water under alkaline environment, cleaned with C18 SPE cartridge, and then hydrolysised with triflu-oroacetic acid at 110℃. The monosaccharaides were derivated with l-phenyl-3-methyl-5-pyrazolone (PMP), and separated at a flow rate of 0. 5 mL/min on a Waters ACQUITY UPLC BEH C18 column (2. 1 mm i. d. ×50 mm, 1. 7 μm) at 35 °C, by using acetonitrile and 0. 5 mmol/L ammonium ace tate - 0. 05% acetic acid buffer as mobile phase by a gradient elution. The analysis of target compounds was performed in multiple reaction monitoring ( MRM) mode via positive electrospray ioniza-tion, and quantified by the internal standard method. The developed method showed good linearities over the concentration range of 1 - 100 jjunol/L for all monosaccharaides with correlation coefficient (r2) more than 0. 96. The spiked recoveries were between 84% and 112%, and the relative standard deviations ( RSD) were no more than 4. 7% . The results showed that Mesona Chinensis Benth polysaccharides consisted of eight monosaccharides, including mannose, rhamnose, ribose, glueu-ronic acid, galacturonic acid, glucose, galactose, xylose, with the molar percentages of 7.4%,5.7%, 4. 2%, 0. 9%, 28.4%, 26. 5%, 16.4% and 10. 6%, respectively. The method is simple, rapid, sensitive and convenient, and could be applied in the analysis of Mesona Chinensis Benth polysaccharides.