In vivo monitoring of hepatic glutathione in anesthetized rats by 13C NMR
In vivo monitoring of hepatic glutathione in anesthetized rats by 13C NMR
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DOI：
10.1002/mrm.10244
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发表时间：
2002-09-01
影响因子：
3.3
通讯作者：
James, TL
中科院分区：
文献类型：
--
作者：
Macdonald, JM;Schmidlin, O;James, TL
已经开发出一种在完整麻醉大鼠体内进行肝谷胱甘肽 (GSH) 体内 C-13 NMR 监测的方法。研究使用专为该动物模型设计的三重调谐、手术植入的表面线圈进行。该线圈允许 C-13 NMR 谱完全解耦和足够的分辨率，以监测静脉注射 2-C-13-甘氨酸的肝脏 C-13-代谢物的时间过程，特别是分别在 44.2 ppm 处的谷胱甘肽和在 61.1 ppm 和 57.2 ppm 处的丝氨酸信号。它还允许通过 P-31 NMR 同时监测高能磷酸原和细胞内 pH 值。为了确认体内 NMR 峰归属，我们将肝高氯酸提取物的高分辨率 2D H-1{C-13} 异核多量子相干谱和 1D C-13 光谱与真实标准品进行了比较。肝脏C-13甘氨酸同位素富集分数以1.68 h(-1)的速率呈指数增加，并在2 h内达到81%的平台水平。 GSH的C-13同位素富集分数以0.316 h(-1)的速率呈指数增加，并在2-C-13-甘氨酸输注4小时后达到55%，但没有达到平台期。为了确认 44.2 ppm 处的共振是由 GSH 引起的，给大鼠静脉注射 2-氧代噻唑烷-4-羧酸 (OTC)，这是一种增加细胞内 GSH 的半胱氨酸前体。正如预期的那样，通过 OTC 给药，肝脏 C-13 GSH 至甘氨酸的峰面积增加了七倍以上。 (C) 2002 Wiley-Liss, Inc.
A method for in Vivo C-13 NMR monitoring of hepatic glutathione (GSH) in intact, anesthetized rats has been developed. Studies were conducted using a triple-tuned, surgically implanted surface coil designed for this animal model. The coil permitted complete decoupling and sufficient resolution in the C-13 NMR spectrum to monitor the time course of hepatic C-13-metabolites of intravenously administered 2-C-13-glycine, particularly GSH at 44.2 ppm and serine signals at 61.1 and 57.2 ppm, respectively. It further allowed concomitant monitoring of high-energy phosphagens and intracellular pH by P-31 NMR. To confirm in vivo NMR peak assignments, we compared high-resolution 2D H-1{C-13} heteronuclear multiple quantum coherence and 1D C-13 spectra of hepatic perchloric acid extracts to those of authentic standards. The fractional isotopic enrichment of hepatic C-13 glycine increased exponentially at a rate of 1.68 h(-1) and reached its plateau level of 81% in 2 h. The C-13 fractional isotopic enrichment of GSH increased exponentially at a rate of 0.316 h(-1) and reached 55% after 4 h of 2-C-13-glycine infusion, but without achieving a plateau. To confirm that the resonance at 44.2 ppm resulted from GSH, a rat was given an intravenous dose of 2-oxothiazolidine-4-carboxylic acid (OTC), a cysteine precursor that increases intracellular GSH. As expected, with OTC administration the hepatic C-13 GSH-to-glycine peak area increased more than sevenfold. (C) 2002 Wiley-Liss, Inc.