Because d-amino acids (AAs) play an essential role in the regulation of many processes in living cells, detection of D-AAs and assay of d-amino acid oxidase (DAAO) activity are of vital importance in bioanalytical science. However, the reliability and accuracy of DAAO assays could be interfered due to the facts that DAAO presents broad substrate activity towards different D-AAs and there could be abundant L-AA enantiomers in biological samples. In this study we presented the first application of optically gated capillary electrophoresis with LIF detection (OGCE-LIF) for efficient assay of DAAO activity. High repeatability of the OGCE-LIF assay of amino acids (AAs) was achieved with relative standard deviation (RSD) (n = 15) less than 1.5% and 2.7% for migration time and peak height, respectively. Under the optimal OGCE-LIF conditions, five pairs of D/L-AA enantiomers were efficiently separated in less than 1 min with low limit of detection of 1.3 μM. Enzymatic assays of DAAO were successfully achieved by detection the substrate consumption with OGCE-LIF, for either single or mixed AA substrates. Kinetic analysis of the parallel oxidation reactions of two different substrates was performed, which was in good agreement with the experimental results. Our study indicates OGCE-LIF can perform rapid and efficient separation of mixed pairs of AA enantiomers and is a promising method for quantitatively assaying DAAO catalyzed reaction with the presence of L-AA enantiomers in the sample. Our study would pave the way for accurate determination of D-AAs and DAAO enzymes in complicated biological samples.

因为d-氨基酸（AAs）在活细胞许多过程的调节，D-AAs的检测和d的测定中起着至关重要的作用-氨基酸氧化酶（DAAO）活性在生物分析科学中至关重要。但是，由于DAAO对不同D-AA具有广泛的底物活性，并且生物样品中可能存在大量L-AA对映体，因此DAAO测定的可靠性和准确性可能会受到干扰。在这项研究中，我们介绍了具有LIF检测（OGCE-LIF）的光学门控毛细管电泳在DAAO活性有效测定中的首次应用。氨基酸（AAs）的OGCE-LIF分析具有很高的重复性，相对标准偏差（RSD）（n = 15）分别小于迁移时间和峰高的1.5％和2.7％。在最佳OGCE-LIF条件下，可以在不到1分钟的时间内有效分离出五对D / L-AA对映异构体，其检测下限为1.3μM。通过使用OGCE-LIF检测单个或混合AA底物的底物消耗，成功实现了DAAO的酶法测定。对两种不同底物的平行氧化反应进行了动力学分析，与实验结果吻合良好。我们的研究表明，OGCE-LIF可以快速有效地分离成对的AA对映异构体，并且对于定量分析样品中存在L-AA对映异构体的DAAO催化反应是一种很有前途的方法。我们的研究将为准确测定复杂生物样品中的D-AAs和DAAO酶铺平道路。对两种不同底物的平行氧化反应进行了动力学分析，与实验结果吻合良好。我们的研究表明，OGCE-LIF可以快速有效地分离成对的AA对映异构体，并且对于定量分析样品中存在L-AA对映异构体的DAAO催化反应是一种很有前途的方法。我们的研究将为准确测定复杂生物样品中的D-AAs和DAAO酶铺平道路。对两种不同底物的平行氧化反应进行了动力学分析，与实验结果吻合良好。我们的研究表明，OGCE-LIF可以快速有效地分离成对的AA对映异构体，并且对于定量分析样品中存在L-AA对映异构体的DAAO催化反应是一种很有前途的方法。我们的研究将为准确测定复杂生物样品中的D-AAs和DAAO酶铺平道路。