Nunes Patricia M, Carvalho Eugénia, Jones John G
Center for Neurosciences and Cell Biology, Department of Zoology, University of Coimbra, 3004-517 Coimbra, Portugal.
Carbohydr Res. 2008 Jul 7;343(9):1486-9. doi: 10.1016/j.carres.2008.04.004. Epub 2008 Apr 7.
Glycogen was quantified in rat adipocytes by isolation using conventional KOH digestion and ethanol precipitation, followed by hydrolysis and spectrophotometric assay of the glucose product. A concentration of 0.193+/-0.020 micromol glucosyl units/10(6)cells was recorded. When this procedure was modified by including a 4h incubation with glucose oxidase prior to glycogen hydrolysis, the glycogen concentration was found to be 0.055+/-0.008 micromol glucosyl units/10(6) cells. Therefore in adipocytes, conventional glycogen assays give substantial overestimates due to incomplete removal of glucose during glycogen isolation. Contaminant glucose can be scavenged in a simple manner by incubation with glucose oxidase prior to glycogen hydrolysis.
采用常规的氢氧化钾消化和乙醇沉淀法分离大鼠脂肪细胞中的糖原,随后进行水解,并对葡萄糖产物进行分光光度测定,以此对糖原进行定量分析。记录的糖原浓度为0.193±0.020微摩尔葡萄糖基单位/10⁶个细胞。当在糖原水解前加入葡萄糖氧化酶孵育4小时对该方法进行改进时,发现糖原浓度为0.055±0.008微摩尔葡萄糖基单位/10⁶个细胞。因此,在脂肪细胞中,由于糖原分离过程中葡萄糖去除不完全,常规的糖原检测会产生显著高估。在糖原水解前通过与葡萄糖氧化酶孵育,可以简单地清除污染的葡萄糖。