Palmieri M J, Reynolds R A, Gibson J B, Berry G T, Segal S
Division of Biochemical and Molecular Diseases, Children's Hospital of Philadelphia, PA 19104.
Enzyme Protein. 1993;47(3):105-15. doi: 10.1159/000468666.
We have applied a HPLC method to separate and quantitate UDPgalactose (UDPGal) and UDPglucose (UDPGlu) in human white blood cells (WBCs). Trichloroacetic acid-treated, protein-free filtrates were chromatographed on an anion-exchange column (Dionex CarboPac PA-1) using a gradient of 20-40% potassium phosphate buffer (pH 4.5). Recoveries of UDPGal and UDPGlu ranged from 93 to 106%, and the method was linear over a wide range of WBC protein concentrations. Volumes of blood as low as 2.5 ml (2.2 mg WBC protein) could be used to achieve quantitative recovery of the sugar nucleotides. The mean values and standard deviations of UDPGal and UDPGlu in 33 normal individuals ranging in age from 1 day to 65 years were 12.4 +/- 4.2 and 31.5 +/- 9.3 mumol/100 g protein, respectively. No statistical differences in UDPGal and UDPGlu values were observed between children and adults. No correlation was established between the concentrations of UDPGal and UDPGlu and either total WBC count or the number of lymphocytes obtained from Coulter counter analysis. There was no relationship between the concentrations of UDPGal and UDPGlu in WBCs and RBCs which were prepared from the same blood specimen.
我们已应用高效液相色谱法（HPLC）来分离和定量人白细胞（WBC）中的UDP半乳糖（UDPGal）和UDP葡萄糖（UDPGlu）。经三氯乙酸处理的无蛋白滤液在阴离子交换柱（Dionex CarboPac PA - 1）上进行色谱分析，使用20 - 40%磷酸钾缓冲液（pH 4.5）的梯度洗脱。UDPGal和UDPGlu的回收率在93%至106%之间，并且该方法在广泛的白细胞蛋白浓度范围内呈线性。低至2.5毫升血液（2.2毫克白细胞蛋白）的量即可用于实现糖核苷酸的定量回收。33名年龄从1天至65岁的正常个体中，UDPGal和UDPGlu的平均值及标准差分别为12.4±4.2和31.5±9.3微摩尔/100克蛋白。儿童和成人之间未观察到UDPGal和UDPGlu值的统计学差异。UDPGal和UDPGlu的浓度与通过库尔特计数器分析获得的白细胞总数或淋巴细胞数量之间未建立相关性。在由同一血液样本制备的白细胞和红细胞中，UDPGal和UDPGlu的浓度之间没有关系。
