Usuki S, Nagai Y
Anal Biochem. 1986 Jan;152(1):172-7. doi: 10.1016/0003-2697(86)90137-5.
A new procedure for introducing tritium into the carbohydrate portions of glucosyl- and galactosylceramides was developed using a new catalyst, CrO3-graphite, which specifically oxidizes the primary alcohol group to the aldehyde. About 10% of the glycolipid was converted to the aldehyde and the aldehyde produced was then reduced back to the original form with KB3H4. After methanolysis, more than 96.7% of the radioactivities of [3H]glucosyl- and [3H]galactosylceramides were found to be located in the carbohydrate portions, and the specific activities of the [3H]galactosyl- and [3H]glucosylceramides were 2.08 to 4.30 X 10(4) cpm/nmol, which could be increased greatly by purifying the aldehydes and reducing them with KB3H4. In addition, beta-galactosidase activity was successfully determined with [3H]galactosylceramide as the enzyme substrate; the Km was 18.73 mM and the Vmax was 11.63 nmol/mg/h, indicating that no significant structural modification occurs during the oxidation.
利用一种新型催化剂CrO₃-石墨开发了一种将氚引入葡糖基神经酰胺和半乳糖基神经酰胺碳水化合物部分的新方法，该催化剂能将伯醇基特异性氧化为醛。约10%的糖脂转化为醛，然后用KB₃H₄将生成的醛还原回原始形式。甲醇解后，发现[³H]葡糖基神经酰胺和[³H]半乳糖基神经酰胺的放射性超过96.7%位于碳水化合物部分，[³H]半乳糖基神经酰胺和[³H]葡糖基神经酰胺的比活为2.08至4.30×10⁴ cpm/nmol，通过纯化醛并用KB₃H₄还原可大幅提高。此外，以[³H]半乳糖基神经酰胺为酶底物成功测定了β-半乳糖苷酶活性；Km为18.73 mM，Vmax为11.63 nmol/mg/h，表明氧化过程中未发生明显的结构修饰。
