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以人参皂苷Rg1为配基的亲和色谱介质的制备与初步应用

Preparation and application of affinity chromatography medium with Ginsenoside Rg1 as a ligand

  • 摘要: 通过偶联人参皂苷Rg1(Rg1)与环氧活化琼脂糖凝胶6B(epoxy-activated SepharoseTM 6B,EAS6B),制备以Rg1为配基的亲和介质,以红外色谱法验证其偶联,并采用正交设计方法优化偶联条件为40 ℃,pH 13,每毫升EAS6B添加Rg1 100 μmol;以此条件放大工艺制备的亲和介质偶联率为31.47%,偶联量为31.56 μmol/mL;利用Rg1-EAS6B亲和介质,以系列亲和色谱法,从PC12细胞蛋白裂解液中筛选得到1个Rg1特异性结合蛋白的条带,为制备含有类似活性基团的中药有效成分的亲和色谱介质提供参考依据,也为进一步探讨人参皂苷Rg1的作用机制奠定基础。

     

    Abstract: Ginsenoside Rg1(Rg1) is one major active component of the traditional Chinese herb,Ginseng.It showed remarkable neuroprotective effects in vitro and in vivo.In order to provide new technological support for the identification of the target of Rg1 and further explanation of its activities,the affinity medium Rg1-epoxy-activated sepharoseTM6B(EAS6B) was prepared by coupling Rg1 to EAS6B.The copulation was optimized by a L9()(34) orthogonal design experiment,and verified by IR.The optimal coupling method was determined as 40 ℃,pH 13,n(Rg1)∶V(EAS6B)=100 μmoL∶1 mL according to the orthogonal analysis and practical application.The coupling rate and yield were 31.47% and 31.56 μmol/mL in amplified preparation.Furthermore,a specific protein band was found from PC12 lysate using Rg1-EAS6B medium by serial affinity chromatography from PC12 cell lysate.The study may provide some references for further identifying the target of Rg1 and preparing affinity chromatography resin of other natural compounds with similar active group.

     

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