Assay of thioinosinic acid, an active metabolite of azathioprine, in human lymphocytes.

Authors:
  • LFL Fletcher
    L Fletcher
  • JMJl Maddocks
    Jl Maddocks
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Abstract
1 A specific assay for the measurement of thioinosinic acid, in human lymphocytes, has been developed with a sensitivity of 50 ng of thioinosinic acid per 5 × 10(6) lymphocytes. 2 Thioinosinic acid is precipitated from purified lymphocytes as the lanthanum salt. Acid hydrolysis results in the formation of 6‐mercaptopurine which, when converted into its phenyl mercury derivative, can be easily extracted into toluene. Back‐extraction of the toluene layer with 0.1N HCl regenerates 6‐mercaptopurine which is then assayed fluorometrically. 3 Blood samples were taken from renal transplant recipients 3 h after an oral dose of 50 mg azathioprine. The results from 5 patients gave a range of 54 to 173 ng of thioinosinic acid per 5 × 10(6) lymphocytes, with a mean of 110 ng. 4 In an in vitro incubation of azathioprine, 1mM with fresh human blood, 160 and 180 ng of thioinosinic acid per 5 × 10(6) lymphocytes was formed after 0.5 h and 5 h respectively. 5 The assay is suitable for the study of the kinetics of thioinosinic acid formation in lymphocytes of patients with kidney transplants. It could also prove useful in the study of thioinosinic acid formation in leukaemia patients undergoing 6‐mercaptopurine treatment.

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