Qurishi Ramatullah, Kaulich Marko, Müller Christa E
Department of Pharmaceutical Chemistry Poppelsdorf, Pharmaceutical Institute, University of Bonn, Germany.
J Chromatogr A. 2002 Apr 5;952(1-2):275-81. doi: 10.1016/s0021-9673(02)00095-x.
An easy and fast method for the quantitative analysis of nucleotides by capillary zone electrophoresis was developed. The method employing a neutral-bonded capillary and reversed polarity mode provided a good resolution and a short analysis time of less than 5 min. The samples were injected electrokinetically using -6 kV voltage for 30 s and detected by their UV absorbance at 254 nm. Constant current (-45 microA) was applied, and a phosphate buffer, pH 7.4, was used. The detection limits for ATP, UDP, and UTP ranged between 0.14 and 0.28 microM. This method was required for the investigation of the purity of the commercially available nucleotides used in pharmacological studies. In addition, the analytical method was applied to study the metabolism of nucleotides in a cell line, neuroblastoma x glioma hybrid cells (NG108-15), which is used in pharmacological studies with nucleotides, since it contains purine- and pyrimidine-sensitive nucleotide receptors. Furthermore, we used the new method for monitoring enzymatic studies using the enzyme hexokinase to convert nucleotide triphosphates to diphosphates.
开发了一种通过毛细管区带电泳对核苷酸进行定量分析的简便快速方法。该方法采用中性键合毛细管和反相极性模式，具有良好的分离度，分析时间短于5分钟。样品通过 -6 kV 电压电动进样30秒，并通过在254 nm处的紫外吸收进行检测。施加恒定电流（-45 μA），使用pH 7.4的磷酸盐缓冲液。ATP、UDP和UTP的检测限在0.14至0.28 μM之间。该方法对于研究药理学研究中使用的市售核苷酸的纯度是必需的。此外，该分析方法还应用于研究细胞系神经母细胞瘤x胶质瘤杂交细胞（NG108-15）中核苷酸的代谢，该细胞系用于核苷酸的药理学研究，因为它含有嘌呤和嘧啶敏感的核苷酸受体。此外，我们使用新方法监测使用己糖激酶将三磷酸核苷酸转化为二磷酸核苷酸的酶学研究。
