DETERMINATION OF RIBONUCLEOSIDE TRIPHOSPHATES AND DEOXYRIBONUCLEOSIDE TRIPHOSPHATES IN NOVIKOFF HEPATOMA-CELLS BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY
DETERMINATION OF RIBONUCLEOSIDE TRIPHOSPHATES AND DEOXYRIBONUCLEOSIDE TRIPHOSPHATES IN NOVIKOFF HEPATOMA-CELLS BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY
复制标题
DOI：
10.1016/0003-2697(87)90613-0
复制
发表时间：
1987-01-01
影响因子：
2.9
通讯作者：
AREZZO, F
中科院分区：
文献类型：
--
作者：
AREZZO, F
建立了一种快速、特异、灵敏的测定Novikoff肝癌细胞中核糖核苷和脱氧核糖核苷三磷酸的方法。使用了一个简单的三步程序。(I)用5%的冷三氯乙酸(TCA)提取生物样品；(Ii)用乙醚洗涤去除TCA，冷冻干燥浓缩样品；(Iii)在等规度条件下分离CTP、dCTP、ATP、dATP、UTP、dTTP、GTP、dGTP，并用阴离子交换高效液相色谱法测定它们的含量。通过与纯核糖核苷三磷酸盐(NTPs)或脱氧核糖核苷三磷酸盐(DNTDPs)的共层析，以及与已知的NTP和dNTP标准品的280 nm：254 nm光谱比的比较，所有化合物都得到了鉴定。用[~(32)P]正磷酸标记的Novikoff肝癌细胞也测定了上述核苷酸的比活性。
A rapid, specific, and sensitive method has been developed for the determination of ribonucleoside and deoxyribonucleoside triphosphates in Novikoff hepatoma cells. A simple three-step procedure was used. (i) Extraction of the biological material with 5% cold trichloroacetic acid (TCA); (ii) elimination of TCA by ethilic ether wash and concentration of the sample by lyophilization; and (iii) separation of CTP, dCTP, ATP, dATP, UTP, dTTP GTP, dGTP and their quantitation of anionic-exchange high-performance liquid chromatography under isocratic conditions. All the compounds were identified by comparing their retention times with those of pure compounds, by cochromatography with single pure ribonucleoside triphosphates (NTPs) or deoxyribonucleoside triphosphates (dNTDPs), and by comparing the 280 nm:254 nm spectral ratios of the peaks with those of known NTP and dNTP standards. The specific activity of all the above mentioned nucleotides also was determined in Novikoff hepatoma cells labeled with [32P]orthophosphate.