To examine whether or not the levels of NOx (nitrite; NO2- and nitrate; NO3-) in coronary circulating blood reflect endothelial dysfunction due to coronary atherosclerosis, NOx levels in plasma obtained from ostium of left coronary artery and coronary sinus of patients who complained of chest pain were evaluated in relation to their coronary angiographic findings. Prior to the study, a HPLC-Griess system for NOx measurement was critically evaluated. This system has a detection limit of 0.1 microM of NO2- and NO3- by 10 microl of loading and was able to distinguish a difference of 0.1-0.2 microM of these substances. Heparin (1 U/10 microl) did not affect the detective and discriminative abilities. NO3- difference, calculated from sino-arterial difference of NO3-, was almost zero (-0.2 +/- 0.2 microM) in patients with either normal coronary arteries or mild organic coronary stenosis (< or = 20% narrowing), while a significant negative value (-5.9 +/- 1.7 microM) was obtained from patients with significant stenosis (> or = 70% narrowing) in the left coronary arteries. These results demonstrate reliable ability on the HPLC-Griess system in evaluating NO2- and NO3- in biological samples, and that the negative NO3- difference through coronary circulation may reflect endothelial dysfunction in the patients with coronary atherosclerosis with severe organic stenosis.