1-苯基-3-甲基-5-吡唑啉酮衍生中药寡糖的毛细管区带电泳分离

Separation of 1-phenyl-3-methyl-5-pyrazolone derivatives of traditional Chinese medicine oligosaccharides by capillary zone electrophoresis

建立了一种可用于中药寡糖分离的毛细管区带电泳(CZE)分析方法。分离条件:未涂层熔融石英毛细管柱(48 cm×50μm,有效长度38 cm),紫外检测波长245 nm,采用1-苯基-3-甲基-5-吡唑啉酮(PMP)为寡糖衍生试剂,50 mmol/L磷酸盐溶液(pH 2.5)为运行缓冲液,电压15 kV,重力进样10 cm×2 s。针对中药寡糖实际样品的复杂性,通过添加多种常见单糖进行方法的实用性验证,并且将方法用于板蓝根多糖的控制降解产物的分离。结果表明,板蓝根寡糖组分可按相对分子质量从小到大的顺序分离,分离效果令人满意。该分离方法操作简单、高效,可用于中药寡糖实际样品分析。另外,还对单糖、寡糖PMP衍生物的电泳迁移行为进行了初步的理论探讨。

A simple and practical capillary zone electrophoretic （CZE） method has been developed for the separation of traditional Chinese medicine oligosaccharides according to their rela- tive molecular masses （Mr）. To optimize the conditions of the method, the concentration and pH value of the running buffer, and the applied voltage were evaluated. The optimized condi- tions were as follows： 1-phenyl-3-methyl-5-pyrazolone （PMP） derivatives of the oligosaccha- rides were separated with 50 mmol/L phosphate buffer （pH 2.5） as running buffer and the voltage was 15 kV. The detection was performed with an ultraviolet detector at 245 nm. An uncoated fused-silica capillary of 50 μm i.d. and 38/48 cm length （effective length/total length） was employed, and a hydrodynamic pressure injection （ 10 cm × 2 s） was applied. In order to examine the practicability of the method for the analysis of actual traditional Chinese medicine oligo-saccharide samples, a complex sample consisting of some usual monosaccharides and oligosaccharides degraded from β-cyclodextrin was separated under the electrophoretic conditions. And then, the method was applied to the analysis of the controlled degradation products of Indigowoad Root polysaccharide. The results indicated that the oligosaccharide sample could fully be separated from low to high Mr. This method is efficient and practical. In addition, the separation hypothesis of PMP derivatives of monosaccharides and oligosaccharides in pH 2.5 running buffer is also discussed, which would be helpful for us to understand the unusual migration of the PMP derivatives of rhamnose and mannose.