The measurement of
hemoglobin-
nitric oxide (NO) adduct (
HbNO) in whole blood by the electron paramagnetic resonance (EPR) method seems relevant for the assessment of systemic NO levels. However,
ceruloplasmin and unknown radical species overlap the same magnetic field as that of
HbNO. To reveal the EPR spectrum of
HbNO, we then introduced the EPR signal subtraction method, which is based on the computer-assisted subtraction of the digitized EPR spectrum of
HbNO-depleted blood from that of sample blood using the software. Rats were treated with
N(omega)-nitro-L-arginine methyl ester (
L-NAME; 120 mg. kg-1. day-1) for 1 wk to obtain
HbNO-depleted blood. When this method was applied to the analysis of untreated fresh whole blood, the five-coordinate state of
HbNO was observed.
HbNO concentration in
pentobarbital-anesthetized rats was augmented (change in [
HbNO] = 1.6-5.5 microM) by infusion of
L-arginine (0.2-0.6 g/kg) but not D-
arginine. Using this method, we attempted to evaluate the effects of
temocapril on
HbNO dynamics in an
L-NAME-induced rat endothelial dysfunction model. The
oral administration of
L-NAME for 2 wk induced a serious
hypertension, and the
HbNO concentration was reduced (change in [
HbNO] = 5.7 microM). Coadministration of
temocapril dose dependently improved both changes in blood pressure and the systemic
HbNO concentration. In this study, we succeeded in measuring the blood
HbNO level as an index of NO by the EPR
HbNO signal subtraction method. We also demonstrated that
temocapril improves abnormalities of NO dynamics in
L-NAME-induced endothelial dysfunction rats using the EPR
HbNO signal subtraction method.