Effects of di-isopropylfluorophosphate and isopropanol on the measurement of plasma renin activity

Abstract

Since the protease inhibitor, di-isopropylfluorophosphate (DFP), inhibits the cryoactivation of inactive renin, its addition to stored plasma should prevent unintended renin activation and thereby minimize this source of inaccuracy in the assay of plasma renin activity. However, we have found that 6 mmol/1 DFP in stored frozen plasma actually reduced plasma renin activity from 4.89 ± 0.13 (S.D.) to 2.48 ± 0.17 ng · ml −1 · h −1 ( p < 0.001). This effect did not occur if DFP was added immediately before assay, suggesting that the decreased renin activity reflected an action of DFP on components of the renin system during storage rather than any interference with the assay method itself. The organic solvent isopropanol, which is required to dilute the DFP, appeared responsible for this phenomenon, for when the isopropanol alone, in concentrations of 20 and 40 μl/ml, was added to stored frozen plasma it decreased plasma renin activity from 3.6 ± 0.3 to 2.2 ± 0.1 and 0.6 ± 0.1 ng · ml −1 · h −1, respectively ( p < 0.001 in each case). Correspondingly, plasma renin substrate concentrations were decreased to 71% and 6% of control, indicating that the renin activity reductions produced by isopropanol were due to its denaturation of substrate. Moreover, addition of exogenous sheep renin substrate (1400 ng/ml) immediately before assay restored plasma renin activity to control. Thus, although DFP effectively prevents inadvertent renin activation in stored frozen plasma, it would seem important that subsequent assays for plasma renin activity be performed in the presence of added exogenous substrate.