有机聚合物整体柱的制备以及对生物大分子的应用
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摘要
整体柱具有渗透性好、制备简单、无需塞子制作等优点，从而避免了由塞子所引起的问题，已引起人们越来越多的关注，已在反相色谱、离子交换色谱、疏水作用色谱、亲和色谱、体积排阻色谱中获得了应用。
本文结合本研究室的工作，对有机聚合物整体柱的制备条件以及该柱对生物大分子的应用作一概述。
本文通过原位聚合制备了一种以甲基丙烯酸缩水甘油酯(GMA)为单体，乙二醇二甲基丙烯酸酯(EDMA)为交联剂的连续棒状色谱柱，并考察了单体、交联剂、制孔剂等条件对柱结构的影响，以此色谱柱为基质进行多方面的研究。
首先通过硫酸水解将整体柱处理为高效疏水色谱柱并建立了蛋白质非线性色谱保留行为的数学模型，利用此种色谱柱对蜗牛酶进行了分离，考察了致孔剂及单体对柱体疏水性及蜗牛酶分离效果的影响。
然后建立了蛋白质非线性色谱保留行为的数学模型，应用此数学模型对梯度洗脱条件下连续棒状疏水色谱柱上不同蛋白质非线性行为的保留时间进行了预测和实验验证。
最后通过键合氨基的方法成功合成连续棒状弱阴离子交换柱，通过优化色谱条件，对IL-18粗提液进行了分离，考察了在不同条件对白介素-18在该色谱柱上分离效果的影响。
In contrast to traditional packed column, monolithic column has the advantage of easy preparation, excellent permeability, low backpressure at chromatographic runs and large capacity for samples. This kind of column has been applied in many fields.
This paper consists of the following parts: preparation of monolithic polymer column, investigation of polymerization conditions and its application to bio-molecules.
First, a continuous rod hydrophobic interaction chromatographic column consisting of poly(glycidyl methacrylate and ethylene dimethacrylate) was prepared by a free radical polymerization and used in the separation of snailase. The influences of polymerization conditions on the hydrophobility of the rod and on the separation of snailase were investigated.
Second, a mathematical model predicting the non-linear retention behavior protein was established. Using this model, the non-linear retention behaviors of different proteins under the gradient elution in high performance hydrophobic interaction chromatographic column have been predicted.
Third, a continuous ion-exchange chromatographic column with glycidyl methacrylate as the functional monomer and ethylene dimethacrylate as the cross linker was prepared by a free radical polymerization and applied to the separation of IL-18. After the factors influencing the separation were investigated, an optimum condition for separating EL-18 from its related proteins was obtained.
本文结合本研究室的工作，对有机聚合物整体柱的制备条件以及该柱对生物大分子的应用作一概述。
本文通过原位聚合制备了一种以甲基丙烯酸缩水甘油酯(GMA)为单体，乙二醇二甲基丙烯酸酯(EDMA)为交联剂的连续棒状色谱柱，并考察了单体、交联剂、制孔剂等条件对柱结构的影响，以此色谱柱为基质进行多方面的研究。
首先通过硫酸水解将整体柱处理为高效疏水色谱柱并建立了蛋白质非线性色谱保留行为的数学模型，利用此种色谱柱对蜗牛酶进行了分离，考察了致孔剂及单体对柱体疏水性及蜗牛酶分离效果的影响。
然后建立了蛋白质非线性色谱保留行为的数学模型，应用此数学模型对梯度洗脱条件下连续棒状疏水色谱柱上不同蛋白质非线性行为的保留时间进行了预测和实验验证。
最后通过键合氨基的方法成功合成连续棒状弱阴离子交换柱，通过优化色谱条件，对IL-18粗提液进行了分离，考察了在不同条件对白介素-18在该色谱柱上分离效果的影响。
In contrast to traditional packed column, monolithic column has the advantage of easy preparation, excellent permeability, low backpressure at chromatographic runs and large capacity for samples. This kind of column has been applied in many fields.
This paper consists of the following parts: preparation of monolithic polymer column, investigation of polymerization conditions and its application to bio-molecules.
First, a continuous rod hydrophobic interaction chromatographic column consisting of poly(glycidyl methacrylate and ethylene dimethacrylate) was prepared by a free radical polymerization and used in the separation of snailase. The influences of polymerization conditions on the hydrophobility of the rod and on the separation of snailase were investigated.
Second, a mathematical model predicting the non-linear retention behavior protein was established. Using this model, the non-linear retention behaviors of different proteins under the gradient elution in high performance hydrophobic interaction chromatographic column have been predicted.
Third, a continuous ion-exchange chromatographic column with glycidyl methacrylate as the functional monomer and ethylene dimethacrylate as the cross linker was prepared by a free radical polymerization and applied to the separation of IL-18. After the factors influencing the separation were investigated, an optimum condition for separating EL-18 from its related proteins was obtained.
引文
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[3] Svec F, Frechet J M. Continuous rods of macroporous polymer as high-performance liquid chromatography separation media Anal. Chem. 1992,64:820-822
[4] 平贵臣，袁湘林，张维冰，张玉奎．整体柱的制备方法及其应用．分析化学．2001．29．(12)：1464-1469
[5] 何炜，雷建都，谭天伟．连续床色谱分离技术 化工进展．2002，21(4)：262
[6] Mihelic I, Koloini T, Podgornik A, Strancar A. Dynamic capacity studies of CIM(convective interaction media) monolithic columns. J. High Resol. Chromatogr.,2000,23(1):39-43
[7] Minakuchi H, Nakanishi K, Soga N, Tanaka N. Performance of an octadecylsilylated continuous porous silica column in polypeptide separations J. Chromatogr. A, 1998, 828: 83～90
[8] Ericson C, Nakazato K, Hjerten S. J. Preparation of continuous beds for electrochromatography and reversed-phase liquid chromatography of low-molecular-mass compounds.J.Chromatogr. A, 1997, 767:33～41
[9] Minakuchi H, Nakanishi K, Soga N, Tanaka N. Effect of skeleton size on the performance of octadecylsilylated continuous porous silica columns in reversed-phase liquid chromatography J. Chromatogr. A, 1997, 762:135～146
[10] Fujimotoj C, Fujise Y, Matsuzawa E. Fritless Packed Columns for Capillary Electrochromatography: Separation of Uncharged Compounds on Hydrophobic Hydrogels. Anal.Chem., 1996, 68:2753-2757
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