培养细胞中磷酸己糖旁路活性的气相色谱-质谱分析。

Gas chromatographic-mass spectrometric analysis of hexose monophosphate shunt activity in cultured cells.

作者信息

Mitchell S L, Ross B D, Krick T, Garwood M

机构信息

Department of Radiology, University of Minnesota, Minneapolis 55455.

出版信息

Biochem Biophys Res Commun. 1989 Jan 31;158(2):474-9. doi: 10.1016/s0006-291x(89)80072-5.

DOI:10.1016/s0006-291x(89)80072-5

PMID:2916995

Abstract

A GC/MS method is described for monitoring the relative amount of glucose degraded to lactate via the hexose monophosphate shunt (HMPS) in neoplastic cells. C6 glioma cells were incubated in medium supplemented with [1-13 C]glucose and medium containing [1-13C]glucose with 0.001 mM phenazine methosulfate (PMS). The ratio of the [13C]lactate to [12C]lactate determined from the measurement of the m/z 219/220 and 117/118 ions of the trimethyl silyl derivative, was used to calculate HMPS activity. PMS increased HMPS activity in C6 glioma cells by 3.2 and 4.8 fold at 2 and 12 hours of incubation respectively. GC/MS results were compared with 1H NMR measurements of the [3-13C]lactate/[3-12C]lactate ratio. The GC/MS method was found to require less sample size and yielded better sensitivity than the NMR method.

摘要

描述了一种气相色谱/质谱（GC/MS）方法，用于监测肿瘤细胞中通过磷酸己糖途径（HMPS）降解为乳酸的葡萄糖的相对量。将C6胶质瘤细胞在补充有[1-¹³C]葡萄糖的培养基和含有[1-¹³C]葡萄糖及0.001 mM吩嗪硫酸甲酯（PMS）的培养基中孵育。由三甲基甲硅烷基衍生物的m/z 219/220和117/118离子测量确定的[¹³C]乳酸与[¹²C]乳酸的比率，用于计算HMPS活性。在孵育2小时和12小时时，PMS分别使C6胶质瘤细胞中的HMPS活性提高了3.2倍和4.8倍。将GC/MS结果与[3-¹³C]乳酸/[3-¹²C]乳酸比率的¹H NMR测量结果进行比较。发现GC/MS方法所需的样品量较少，并且比NMR方法具有更高的灵敏度。