Analysis of the nucleoside moiety of cobalamin and cobalamin analogues using gas chromatography-mass spectrometry.
Analysis of the nucleoside moiety of cobalamin and cobalamin analogues using gas chromatography-mass spectrometry.
复制标题
使用气相色谱-质谱法分析钴胺素和钴胺素类似物的核苷部分。
DOI：
10.1016/0003-9861(92)90463-7
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发表时间：
1992
影响因子：
3.9
通讯作者：
Allen,RH
中科院分区：
文献类型：
--
作者：
Sundin,DP;Allen,RH
现有的钴胺素和钴胺素类似物鉴定技术通常在表征过程中使用完整的分子，结果有些模糊。在这项研究中，描述了一种方法，用于鉴定钴胺素下轴配体中的核苷以及各种天然存在的钴胺素类似物，这些类似物与核苷中存在的碱基中的钴胺素不同。使用 R-蛋白-琼脂糖柱通过亲和层析从生物样品中分离钴胺素和钴胺素类似物。然后水解下部轴向配体的核苷，并使用混合床柱通过柱色谱分离。核苷用高碘酸盐氧化并用硼氢化物还原。重新分离后，制备叔丁基二甲基甲硅烷基衍生物，并使用气相色谱/质谱法和选择离子监测进行分析。生物合成生产了钴胺素的稳定同位素内标，并用于定量兔肾中的钴胺素。钴胺素类似物也被证明存在于兔肾中，但它们在下部轴向配体中含有 5,6-二甲基苯并咪唑核苷（α-ribodium），表明这些类似物在分子的咕啉环区域中与钴胺素不同。
Existing techniques for identification of cobalamin and cobalamin analogues generally use the intact molecule during characterization with somewhat ambiguous results. In this study a method is described for the identification of the nucleoside in the lower axial ligand of cobalamin and a variety of naturally occurring cobalamin analogues that differ from cobalamin in the base that is present in the nucleoside. Cobalamin and cobalamin analogues were isolated from biological samples by affinity chromatography using R-protein-Sepharose columns. The nucleosides of the lower axial ligand were then hydrolyzed and isolated by column chromatography using a mixed bed column. Nucleosides were oxidized with periodate and reduced with borohydride. After reisolation, thet-butyldimethylsilyl derivatives were prepared and analyzed using gas chromatography/ mass spectrometry with selected ion monitoring. A stable isotope internal standard of cobalamin was biosynthetically produced and used to quantitate cobalamin in rabbit kidney. Cobalamin analogues were also shown to be present in rabbit kidney, but they contain the 5,6-dimethylbenzimidazole nucleoside (α-ribazole) in the lower axial ligand, indicating that these analogues differ from cobalamin in the corrin ring region of the molecule.