Kuster H, Vogt M, Joos B, Nadai V, Lüthy R
Department of Medicine, University Hospital, Zurich, Switzerland.
J Infect Dis. 1991 Oct;164(4):773-6. doi: 10.1093/infdis/164.4.773.
An assay to quantify the phosphorylation products of zidovudine (AZT) in peripheral blood mononuclear cells (PBMC) was developed. Extracts of PBMC were separated by high-performance liquid chromatography. Eluted AZT mono- (MP), di- (DP), and triphosphate (TP) were collected in separate portions. Treatment with alkaline phosphatase yielded equimolar amounts of AZT, which after solid-phase enrichment were assayed by radioimmunoassay. Detection limit was 0.1 pmol/10(6) PBMC for each nucleotide. Recoveries of 102%-118% were observed. AZT nucleotides were measured in samples from three patients receiving 250 mg of AZT every 12 h. Intracellular concentrations of AZT-MP after 1-2 h ranged from 0.9 to 1.4 pmol/10(6) PBMC and then declined to 0.3-1.1 pmol/10(6) PBMC after 4 h. AZT-DP and AZT-TP reached concentrations of 0.3-0.5 pmol/10(6) PBMC after 1-2 h and could not be detected after 4 h in any of the three patients. Duplicate determinations deviated by less than 20%.
开发了一种用于定量外周血单核细胞（PBMC）中齐多夫定（AZT）磷酸化产物的测定方法。PBMC提取物通过高效液相色谱进行分离。洗脱的AZT单磷酸（MP）、二磷酸（DP）和三磷酸（TP）分别收集。用碱性磷酸酶处理产生等摩尔量的AZT，经过固相富集后通过放射免疫测定法进行检测。每种核苷酸的检测限为0.1 pmol/10(6) PBMC。回收率为102% - 118%。对三名每12小时接受250 mg AZT治疗的患者的样本进行了AZT核苷酸测定。1 - 2小时后AZT - MP的细胞内浓度范围为0.9至1.4 pmol/10(6) PBMC，4小时后降至0.3 - 1.1 pmol/10(6) PBMC。1 - 2小时后AZT - DP和AZT - TP的浓度达到0.3 - 0.5 pmol/10(6) PBMC，4小时后在三名患者中的任何一人中均未检测到。重复测定的偏差小于20%。
