MEASUREMENT OF NITROBENZYLTHIOINOSINE IN PLASMA AND ERYTHROCYTES - A PHARMACOKINETIC STUDY IN MICE

Purpose: Nitrobenzylthioinosine (NBMPR), a potent inhibitor of nucleos ide transport in many cell types, modulates the in vivo disposition of several cytotoxic nucleoside analogs. In this study, a radioligand bi nding assay was developed for measurement of the NBMPR content of plas ma and erythrocytes. Methods: The assay was based on the competition b etween NBMPR and [H-3]NBMPR for high-affinity sites on human erythrocy te membranes. With this assay, we followed in mice changes in the NBMP R content of blood plasma and erythrocytes, following the intraperiton eal injection of the disodium salt of NBMPR 5'-monophosphate (NBMPR-P) , a prodrug form of NBMPR. Results: The radioligand binding assay was able to measure precisely as little as 2.5 pmol of NBMPR, allowing the direct determination of NBMPR concentrations in plasma as low as 16 n M. As few as 8 x 10(3) molecules of NBMPR per cell could be determined in erythrocytes. The NBMPR content of plasma from mice injected with NBMPR-P was maximal at about 20 min after injection and declined to < 0.2% of the peak value by 10 h. Erythrocyte-associated NBMPR was also maximal at 20 min, and declined to 11% of the peak value by 10 h after injection. Time courses for the disappearance of NBMPR from plasma an d erythrocytes were monoexponential and yielded half-life values of 0. 39 h and 0.68 h, respectively, an apparent volume of distribution of 0 .61 1/kg, and a clearance of 1.11/h per kg. Conclusions: The radioliga nd binding assay is a sensitive and facile method for monitoring NBMPR concentrations in mammalian plasma and tissue extracts.