A SENSITIVE ASSAY OF LYSOGANGLIOSIDES USING HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY
A SENSITIVE ASSAY OF LYSOGANGLIOSIDES USING HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY
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DOI：
10.1016/0005-2760(91)90021-9
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发表时间：
1991-01-28
期刊：
影响因子：
--
通讯作者：
GOTO, I
中科院分区：
文献类型：
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作者：
KOBAYASHI, T;GOTO, I
溶神经节苷脂，LGM 1、LGM 2和LGM 3，各自携带单个鞘氨醇碱基（即，C18：1，C18：0，C20：1，C20：0），并开发了使用HPLC的这些脂质的灵敏测定方法。 该方法包括用邻苯二甲醛对分子的游离氨基进行荧光衍生化，使用反相HPLC分离每种lysoganglioside的分子种类，并基于已知量的一种lysoganglioside作为内标进行测定。 用这种方法，溶神经节苷脂可以在5-1000 pmol范围内准确测定。 为了测定组织中的脂质，在荧光衍生化之前需要粗分离程序，包括脂质提取、Folch分配和DEAE-Sepharose和AG 1-X2柱色谱。 在正常人和牛的大脑皮层中，检测到0.4-2.0 pmol/mg蛋白质的LGM 1含有C18：1和C20：1鞘氨醇残基。 在来自Sandhoff病患者的额叶皮质中，注意到LGM 2的异常积累（55-78 pmol/mg蛋白质）。 在各种分子种类中，含C18：1的LGM 2是最丰富的。
Lysogangliosides, LGM1, LGM2 and LGM3, each carrying a single sphingoid base (i.e., C18:1, C18:0, C20:1, C20:0), were prepared and a sensitive assay method of these lipids using HPLC was developed. The method involves fluorescence derivatization of the free amino group of the molecule with o-phthalaldehyde, separation of the molecular species of each lysoganglioside using reversed-phase HPLC and assay on the basis of a known amount of one of the lysogangliosides, as the internal standard. Using this method, lysoganglioside can be accurately assayed in the range of 5-1000 pmol. For assay of the lipid in the tissue, crude isolation procedures including extraction of lipids, Folch's partition and DEAE-Sepharose and AG 1-X2 column chromatographies were required before the fluorescence derivatization. In the normal human and the bovine cerebral cortex, 0.4-2.0 pmol/mg protein of LGM1 containing C18:1 and C20:1 sphingosine residues were detected. In the frontal cortex from a patient with Sandhoff disease, an abnormal accumulation (55-78 pmol/mg protein) of LGM2 was noted. Among various molecular species, LGM2 containing C18:1 was the most abundant.