一步柱层析纯化血浆分离多种血液制品组分

Separation of various plasma components by purification with one step column chromatography

目的采用一步吸附及分步解吸的柱层析技术——扩张床吸附(expanded bed adsorption,EBA)技术,从原料血浆中分离多个有效组分,从而提高血浆的综合利用率。方法以琼脂糖(碳化钨)-DEAE为吸附介质,将原料血浆按一定流速流经层析柱,并使待分离组分吸附于介质上,通过不同的缓冲液组成、洗脱流速、洗脱体积分步分离有效组分,并初步探讨工艺放大过程中的关键因素。结果 EBA技术可从原料血浆中依次分离出白蛋白、免疫球蛋白、纤维蛋白原、α-1蛋白酶抑制剂4种蛋白;该技术在工艺放大过程中,柱高及洗脱流速是影响分离效果的关键因素;该技术可去除部分病毒,使病毒滴度降低约一个数量级。结论一步柱层析技术具有血浆综合利用度高、易操作、操作周期短、成本低、可去除病毒等优势,其有望成为传统低温乙醇工艺的有效替代技术,应用于工业化生产。

Objective To separate various active components from source plasma by expanded bed adsorption,a column chromatography for one-step adsorption and stepwise desorption,so as to improve the comprehensive utilization of plasma.Methods Source plasma was loaded onto chromatographic column at a constant flow rate using agarose（tungsten carbide）-DEAE as an adsorption medium to adsorb the components to be separated. The active components were separated stepwise by elution with buffers of various compositions at various flow rates and elution volumes,based on which the key factors in scale-up of procedure were preliminarily investigated. Results Albumin,immunoglobulins,fibrinogen and α-1proteinase inhibitor were separated in turn from source plasma by EBA. The column height and elution rate were key influencing factors of separation effect during scale-up of the procedure. Partial viruses were removed by the technique,which decreased the virus titer by about one order of magnitude. Conclusion With a high utilization level of plasma and a short operation cycle,the one-step column chromatography technique was easy to handle,low-cost and effective in removal of viruses,which might be used for industrial production instead of traditional cold ethanol fractionation.