title: A method for the quantification of intracellular zidovudine nucleotides.
abstract: 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%.
authors.affiliation: Department of Medicine, University Hospital, Zurich, Switzerland.
authors.forename: H
authors.initials: H
authors.lastname: Kuster
authors.forename: M
authors.initials: M
authors.lastname: Vogt
authors.forename: B
authors.initials: B
authors.lastname: Joos
authors.forename: V
authors.initials: V
authors.lastname: Nadai
authors.forename: R
authors.initials: R
authors.lastname: Lüthy
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D002851:Chromatography, High Pressure Liquid; D054306:Dideoxynucleotides; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000737:chemistry*; D010766:Phosphorylation; D011863:Radioimmunoassay; D013942:Thymine Nucleotides / Q000097:blood*; D015215:Zidovudine / Q000031:analogs & derivatives* / Q000097:blood / Q000493:pharmacokinetics*
chemical_list: D054306:Dideoxynucleotides; D013942:Thymine Nucleotides; C064181:3'-azido-3'-deoxythymidine 5'-diphosphate; C057287:3'-azido-3'-deoxythymidine 5'phosphate; D015215:Zidovudine; C052873:zidovudine triphosphate; D000469:Alkaline Phosphatase