Selective spectrofluorimetric determination of glutathione in clinical and biological samples using 1,3,5,7-tetramethyl-8-phenyl-(2-maleimide)-difluoroboradiaza-s-indacene
Selective spectrofluorimetric determination of glutathione in clinical and biological samples using 1,3,5,7-tetramethyl-8-phenyl-(2-maleimide)-difluoroboradiaza-s-indacene
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使用 1,3,5,7-四甲基-8-苯基-(2-马来酰亚胺)-二氟硼二氮杂-s-indacene 选择性分光荧光法测定临床和生物样品中的谷胱甘肽
DOI：
10.1016/j.aca.2008.11.028
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发表时间：
2009-02-02
影响因子：
6.2
通讯作者：
Zhang, Hua-Shan
中科院分区：
文献类型：
--
作者：
Guo, Xiao-Feng;Wang, Hong;Zhang, Hua-Shan
这项工作报告了一种选择性、灵敏和快速的荧光分光光度法的开发，用于使用 1,3,5,7-四甲基-8-苯基-(2-马来酰亚胺)-二氟硼二氮杂-s-indacene 在临床和生物样品中存在相对较高水平的半胱氨酸 (Cys) 的情况下测定还原型谷胱甘肽 (GSH) (TMPAB-o-M)。 TMPAB-o-M 的荧光被其马来酰亚胺部分强烈猝灭，但与硫醇反应后，荧光恢复，强度增加 350 倍（荧光量子产率从 0.002 至 0.73）。在H(3)Cit-Na2HPO4缓冲液(pH 7.40)中，37℃下仅需5分钟即可完成衍生化。线性范围为0.005-0.2 mu mol L-1。检测限为1.1 x 10(-10) mol L-1（信噪比=3）。几乎所有氨基酸，包括 Cys，即使以相对较高的浓度存在也不会产生干扰（氨基酸：GSH = 100：1。Cys：GSH = 1：1，摩尔比，C-GSH = 3 x 10(-7) mol L-1）。样品去除蛋白质后无需进一步处理可直接使用。该方法精确，相对标准偏差（R.S.D.）低于5.0%（n = 6），并已应用于人血和猪肝中GSH的测定，回收率在94.4%至105.6%之间。 (C) 2008 年由 Elsevier B.V. 出版
This work reports the development of a selective, sensitive and rapid spectrofluorimetric method for the determination of reduced glutathione (GSH) in the presence of relatively high levels of cysteine (Cys) in clinical and biological samples using 1,3,5,7-tetramethyl-8-phenyl-(2-maleimide)-difluoroboradiaza-s-indacene (TMPAB-o-M). The fluorescence from TMPAB-o-M is strongly quenched by its maleimide moiety, but after reaction with thiol, the fluorescence is restored with a 350-fold intensity increase (fluorescence quantum yield from 0.002 to 0.73). In H(3)Cit-Na2HPO4 buffer (pH 7.40), the derivatization is completed in just 5 min under 37 degrees C. The linear range is 0.005-0.2 mu mol L-1. with detection limit of 1.1 x 10(-10) mol L-1 (signal-to-noise ratio=3). Almost all amino acids, including Cys, impose no interference even if present at relatively high concentrations (amino acids:GSH = 100: 1. Cys:GSH = 1:1, molar ratio, C-GSH = 3 x 10(-7) mol L-1). The sample can be used directly without further treatment after the protein is removed. The developed method is precise with a relative standard deviation (R.S.D.) lower than 5.0% (n = 6) and has been applied to the determination of GSH in human blood and pig's liver with recoveries between 94.4 and 105.6%. (C) 2008 Published by Elsevier B.V.