化学
色谱法
尿酸
腺苷
高效液相色谱法
黄嘌呤
核苷酸
次黄嘌呤
检出限
黄嘌呤氧化酶
嘌呤
定量分析(化学)
肌苷
腺嘌呤核苷酸
一磷酸腺苷
磷酸盐
作者
Yanjie Chen,Ji Peng,Guangyin Ma,Zehua Song,Bruce Qing Tang,Tongju Li
摘要
Adenine nucleotides and malondialdehyde (MDA) are key components involved in energy metabolism and reactive oxygen species (ROS) production. Measuring the levels of these components at the same time would be critical in studying mitochondrial functions. We have established a HPLC method to simultaneously measure adenosine triphosphate, adenosine diphosphate, adenosine monophosphate, MDA, and uric acid (UA). The samples were treated with perchloric acid followed by centrifugation. After neutralization, the supernatant was subjected to HPLC determination. HPLC was performed using a C18 chromatographic column, isocratic elusion, and UV detection. The detection and quantification limits for these components were determined with standard solutions. The precision, repeatability, and 24-h stability were evaluated using cellular samples, and their relative standard deviations were all within 2%. The reproducibility and efficiency were confirmed with sample recovery tests and the observed oxidative effects of H2 O2 on Jurkat cells. With this method, we discovered the dependence of energy and oxidative states on the density of Jurkat cells cultured in suspension. We also found a significant correlation between UA in serum and that in saliva. These results indicate that this method has good accuracy and applicability. It can be used in biological, pharmacological, and clinical studies, especially those involving mitochondria, ROS, and purinergic signaling.
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