Beijing National Laboratory for Molecular Sciences, Key Laboratory of Analytical Chemistry for Living Biosystems, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China.
Talanta. 2010 Sep 15;82(4):1332-7. doi: 10.1016/j.talanta.2010.06.041. Epub 2010 Jul 23.
A new kind of immobilized human serum albumin (HSA) column was developed by using the sub-micron skeletal polymer monolith based on poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) [poly(GMA-EDMA)] as the support of high-performance affinity chromatography. Using the epoxide functional groups presented in GMA, the HSA immobilization procedure was performed by two different means. The affinity columns were successfully adopted for the chiral separation of D,L-amino acids (AAs). Then this method was shown to be applicable to the quantitative analysis of D-tryptophan, with a linear range between 12.0 microM and 979.0 microM, and a correlation coefficient above 0.99. Furthermore, it was used for the analysis of urine sample. This assay is demonstrated to be facile and relatively rapid. So it allows us to measure the enzyme catalytic activity in the incubation of D,L-AAs with D-AA oxidase and to study the kinetics of the enzyme reaction. It implied that the affinity monolithic columns can be a useful tool for studying DAAO enzyme reaction and investigating the potential enzyme mechanism requirement among chiral conversion.
一种新型的固定化人血清白蛋白（HSA）柱是通过使用基于聚（甲基丙烯酸缩水甘油酯-乙二醇二甲基丙烯酸酯）[聚（GMA-EDMA）]的亚微米骨架聚合物整体作为高效亲和色谱的支撑来开发的。利用 GMA 中存在的环氧化物官能团，通过两种不同的方法进行 HSA 固定化。亲和柱成功地用于 D,L-氨基酸（AAs）的手性分离。然后，该方法被证明适用于 D-色氨酸的定量分析，线性范围为 12.0 μM 至 979.0 μM，相关系数大于 0.99。此外，它还用于尿液样本的分析。该测定方法简便快速。因此，它允许我们在 D,L-AA 与 D-AA 氧化酶孵育过程中测量酶的催化活性，并研究酶反应的动力学。这表明亲和整体柱可以成为研究 DAAO 酶反应和研究手性转化中潜在酶机制要求的有用工具。
