Preparation and characterization of alkylated polymethacrylate monolithic columns for micro-HPLC of proteins.
Preparation and characterization of alkylated polymethacrylate monolithic columns for micro-HPLC of proteins.
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DOI：
10.1002/jssc.200401796
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发表时间：
2004-12
影响因子：
3.1
通讯作者：
Yan Li;Jian Zhang;Rong Xiang;Yanhui Yang;C. Horváth
中科院分区：
文献类型：
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作者：
Yan Li;Jian Zhang;Rong Xiang;Yanhui Yang;C. Horváth
以十二醇和甲苯为多孔原，甲基丙烯酸甘油酯（GMA）和乙二醇二甲基丙烯酸酯（EGDMA）原位共聚制备表面烷基化整体柱，采用微高效液相色谱（microhplc）对蛋白质进行分离。首先，用硫酸水解多孔单体表面的缩水甘油酯基团。形成的羟基与正烷基氯反应形成烷基化固定相。比较了C18和C8固定相柱对蛋白质的分离效果。结果表明，十八烷基衍生化的聚（GMA-EGDMA）载体具有较好的分辨率。采用十八烷基化聚（GMA-EGDMA）整体柱分离蛋白质混合物，采集流出峰，采用基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）进行分析。采用扫描电子显微镜（SEM）、多点BET氮吸附/脱附、达西定律对整体柱的形貌、比表面积、介孔尺寸分布、渗透性等物理性质进行了表征。
Protein separations were carried out by micro-high performance liquid chromatography (micro-HPLC) with surface alkylated monolithic columns, which were prepared by in situ copolymerization of glycidyl methacrylate (GMA) and ethylene glycol dimethacrylate (EGDMA) in the presence of dodecanol and toluene as porogens. First, glycidyl groups at the surface of the porous monolith were hydrolyzed with sulfuric acid. The hydroxyl groups thus formed were then reacted with n-alkyl chloride to form alkylated stationary phase. Separation performance for proteins on columns with C18 and C8 stationary phases was compared. The results showed that a poly(GMA-EGDMA) support derivatized with octadecyl moieties could achieve much better resolution than one with octyl groups. A protein mixture was separated with the octadecylated poly-(GMA-EGDMA) monolithic column, and the effluent peaks were collected and analyzed by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). The physical properties of the monolithic columns such as column morphology, surface area, mesopore size distribution, and column permeability were further characterized by scanning electron microscopy (SEM), multipoint BET nitrogen adsorption/desorption, and Darcy's law, respectively.