利用二維高效液相層析儀同時分離乳酸與3-羥基丁酸鏡像異構物

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Title:	利用二維高效液相層析儀同時分離乳酸與3-羥基丁酸鏡像異構物 Development of a two-dimensional HPLC system for the simultaneous determination of lactate and 3-hydroxybutyrate enantiomers
Authors:	劉書伶 Liu, Shu-Ling
Contributors:	許秀蘊李仁愛
Keywords:	乳酸;3-羥基丁酸;鏡像異構物分離;高效液相層析儀 Chiral separation;2D-HPLC;Lactic acid;3-Hydroxybutyric acid
Date:	2014-06-23
Issue Date:	2019-09-24 12:58:39 (UTC+8)
Abstract:	此篇論文主在建立以二維液相層析儀同時分離乳酸 (Lactate)與3-羥基丁酸 (3-Hydroxybutyrate)鏡像異構物。將螢光衍生化試劑 4-(N-Chloroformylmethyl-N-methylamino)-7-nitro-2,1,3-benzoxadiazole (NBD-COCl)加入含有乳酸以及 3-羥基丁酸的溶液，將混合液置於60˚C 下加熱 15 分鐘，進行螢光衍生化反應後取 20 μL 注入液相層析儀。在第一維的液相層析儀中，利用 capillary monolithic ODS 管柱可以成功分離乳酸與 3-羥基丁酸。在第二維的液相層析儀中利用QD-AX、KSAACSP-001S 管柱及不同比例之氰甲烷、甲醇與三氟乙酸所組成的移動相，來進行乳酸與 3-羥基丁酸鏡像異構物的分離。所得之最佳分離係數分別是 1.14 與 1.08，偵測極限(LOD)大約為 2 微毫莫爾 (2 fmol/inj)。利用此方法成功定量了生物檢體，如在健康人的血漿、尿液以及唾液中發現，無論是乳酸或是 3-羥基丁酸的鏡像異構物均存在人體中。未來此方法可望應用在釐清乳酸與 3-羥基丁酸的鏡像異構物與疾病之間的關聯性與其個別的生理意義。 A two-dimensional HPLC system has been established for the simultaneous determination of lactate and 3-hydroxybutyrate enantiomers. Lactate and 3-hydroxybutyrate were derivatized with NBD-COCl, and heated at 60˚C for 15 min. In the first dimension, the lactate and 3-hydroxybutyrate were separated by a capillary monolithic ODS column. In the second dimension, various chiral stationary phases had been tested. The lactate enantiomers can be separated by the QD-AX and the 3-hydroxybutyrate enantiomers can be separated by the KSAASCP-001S. The mobile phases were composed of acetonitrile, methanol and TFA. The separation factors are 1.14 and 1.08 respectively. The detection limit of lactate and 3-hydroxybutyrate are 2 fmol/injection. Now by using the 2D-HPLC system, we can separate lactate and 3-hydroxybutyrate enantiomers at the same time. We also examined this method in the clinical healthy human samples, and we can find that both the lactate and 3-hydroxybutyrate enantiomers exist on healthy human serum, urine and saliva. In the end, we hope this method can be applied to various clinical disease samples and that will help us to clarify the role of these enantiomers.
Description:	碩士指導教授-許秀蘊共同指導教授-李仁愛委員-方嘉佑委員-蔡東湖委員-李宗徽
Data Type:	thesis
Appears in Collections:	[School of Pharmacy] Dissertation/Thesis

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