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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

HPLC analysis of azathioprine metabolites in red blood cells, plasma and urine in renal transplant recipients.

Anemia has been frequently reported in renal transplant recipients receiving azathioprine for immunosuppression and enalapril for treatment of hypertension. During the course of a prospective trial in such patients we determined azathioprine metabolites in erythrocytes, plasma, and urine as well as erythropoietin and hemoglobin levels in order to evaluate a potential interaction between these 2 drugs, possibly leading to anemia. Two specific high performance liquid chromatography (HPLC) methods for determination of azathioprine metabolites, both employing a mercurial cellulose resin for extraction, are presented. One method using a strong anion exchange column allows detection of 6-thioguanosine di- and triphosphate (thioguanine nucleotides) in red blood cells (RBC) with a sensitivity of 30 pmol/100 microliters RBC. 6-mercaptopurine (MP) and 6-thiouric acid (TUA) in plasma and urine were analyzed simultaneously by reversed-phase HPLC with a sensitivity of 5 ng/ml. The average (median values are given) steady state concentrations of thioguanine nucleotides in erythrocytes came to 267 pmol/100 microliters RBC (range 53-613) with and to 246 pmol/100 microliters RBC (range 39-629) without concomitant enalapril medication. Mean plasma concentrations of MP and TUA 3 hours after drug intake came to 14.8 +/- 9.9 ng/ml and 398 +/- 262 ng/ml, respectively, during enalapril comedication. Withdrawal of enalapril did not influence these metabolite levels coming to 15.3 +/- 9.1 and 451 +/- 253 after stopping enalapril treatment. Thioguanine nucleotides in RBCs were neither related to the dose of azathioprine given (r = -0.113, p > 0.05) nor to hemoglobin levels (r = 0.278, p > 0.05). However, azathioprine dose/kg body weight seemed to be related to hemoglobin concentration, with and without enalapril comedication. We conclude that enalapril therapy does not influence the measured azathioprine metabolites, the reported cases of anemia may rather be due to a pharmacodynamic interaction as shown by the significant increase in erythropoietin after withdrawal of enalapril. The assays described here are suitable to study the metabolism of azathioprine in patients with various diseases.[1]


  1. HPLC analysis of azathioprine metabolites in red blood cells, plasma and urine in renal transplant recipients. Weller, S., Thürmann, P., Rietbrock, N., Gossmann, J., Scheuermann, E.H. International journal of clinical pharmacology and therapeutics. (1995) [Pubmed]
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