Grune T, Ross G A, Schmidt H, Siems W, Perrett D
Institute of Biochemistry, Medical Faculty (Charité), Humboldt University, Berlin, Germany.
J Chromatogr. 1993 Apr 23;636(1):105-11. doi: 10.1016/0021-9673(93)80062-d.
An optimized separation of the main purine compounds of human serum by capillary zone electrophoresis is presented. Separations were performed in an uncoated silica capillary (44 cm x 75 microns I.D., 37 cm to window) on a SpectraPhoresis 1000 system with UV detection. The separation of adenine (Ade), adenosine (Ado), guanine (Gua), guanosine (Guo), hypoxanthine (Hyp), inosine (Ino), xanthine (Xan) and uric acid (UA) was optimized with respect to pH, temperature, applied potential and hydrodynamic injection time. Optimum conditions were 20 mM borate buffer (pH 9.4), 37 degrees C, 20 kV and 9 s load and detection at 260 nm. Linearity extended from 1 to 125 microM. The sensitivity of the method was 0.5 microM, which is adequate for measuring Ade, Gua, Hyp and UA in plasma samples. Plasma samples from newborns were precipitated with an equal volume of perchloric acid (7%, v/v), the supernatant was adjusted to neutral pH with potassium carbonate and, before injection, the sample was alkalized with sodium hydroxide. The method presented here allows the determination of Ade, Guo, Hyp and UA. The levels of the determined purines were compared in samples from control newborns, preterm babies and newborns with asphyxia or acidic serum pH values.
本文介绍了一种通过毛细管区带电泳对人血清中主要嘌呤化合物进行优化分离的方法。分离在未涂层的石英毛细管（内径75微米，长44厘米，至检测窗口长37厘米）中进行，采用SpectraPhoresis 1000系统并配备紫外检测。对腺嘌呤（Ade）、腺苷（Ado）、鸟嘌呤（Gua）、鸟苷（Guo）、次黄嘌呤（Hyp）、肌苷（Ino）、黄嘌呤（Xan）和尿酸（UA）的分离，针对pH值、温度、施加电压和流体动力学进样时间进行了优化。最佳条件为20 mM硼酸盐缓冲液（pH 9.4）、37℃、20 kV、9秒进样，检测波长为260 nm。线性范围为1至125 microM。该方法的灵敏度为0.5 microM，足以测定血浆样品中的Ade、Gua、Hyp和UA。新生儿血浆样品用等体积的高氯酸（7%，v/v）沉淀，上清液用碳酸钾调至中性pH值，进样前用氢氧化钠碱化样品。本文介绍的方法可用于测定Ade、Guo、Hyp和UA。对正常新生儿、早产儿以及窒息或血清pH值呈酸性的新生儿的样品中所测定嘌呤的水平进行了比较。
