Implication of steady state concentrations of nitrite and nitrate metabolites of nitric oxide in plasma and whole blood in healthy human subjects.

1. The aim of the present study was to determine whether the steady state NOx concentration reflects NOx formation in vivo. 2. A NO3- load study was performed after achieving NOx steady state. Chronological changes in NOx concentrations in plasma and whole blood samples from nine healthy subjects were determined by the HPLC-Griess system and NOx concentrations in erythrocytes were estimated as a possible NOx compartment influential in regulating plasma NOx concentrations. 3. Analysis was performed using the first-order one-compartment open model and the NOx formation rate was subsequently calculated. 4. The mean (+/-SEM) steady state NOx concentration of plasma (15.5 +/- 1.6 micromol/L), whole blood (12.8 +/- 1.2 micromol/L) and erythrocytes (11.9 +/- 0.7 micromol/L) did not correlate with the NOx formation rate in the compartments (0.50 +/- 0.05, 0.61 +/- 0.04 and 0.91 +/- 0.17 micromol/kg per h, respectively), whereas a significant correlation was found between the steady state NOx concentration and NOx elimination rate (Kel) in plasma (r=-0.69; P=0.04) and whole blood (r=-0.79; P=0.01). 5. Although there was no direct correlation between steady state NOx concentrations and serum creatinine levels, the correlation between half-life and serum creatinine levels was significant (plasma: r=0.60, P=0.02; whole blood: r=0.49, P=0.04). 6. Plasma NOx concentrations correlated significantly with erythrocyte NOx concentrations (r=0.92, P <0.01; erythrocyte NOx=0.66 x plasma NOx). 7. The results of the present study indicate that NOx does not accumulate excessively into erythrocytes at steady state and during a NO3- load and that the steady state NOx concentration in whole blood and plasma preferentially implies NOx elimination (mainly depending on renal function) rather than NOx formation.

A practical procedure for achieving a steady state of NOx concentration in plasma: with special reference to the NOx content of Japanese daily food.

To establish a concrete procedure to achieve a steady state plasma NOx concentration with Japanese daily food, NOx contents of about 200 types of food and beverages consumed daily were measured and NOx concentration in plasma was monitored till steady state after various degree of intake of NOx restricted food. The NOx content was found to be high in dark green leaved vegetables and low in grains, processed food, fresh and processed seafood. Tap water and mineral water were found to contain various amounts of NOx that were drastically reduced by treatment with a reverse osmosis column and remained in trace amounts after ion exchange column treatment. NOx content was low in drinks such as cola, but was extremely high in vegetable juice containing dark green leaved vegetables. The intake of high NOx drinks resulted in elevated plasma NOx concentration, but intake of low NOx drinks did not change the plasma NOx concentration. Based on these findings, a steady state could be achieved by 18 hours fasting after the intake of a moderately NOx-restricted diet (about 370, micromoles/day) and by 13 hours fasting after the intake of an extremely NOx-restricted diet (< 100 micromoles/day). NOx concentrations in randomly collected blood samples without these conditions were sometimes ten times higher than that at steady state. This procedure can be undertaken under normal Japanese daily life and is expected to be applicable even to outpatients.

Decrease in plasma NOx concentration by isosorbide dinitrate, an organic nitrate ester.

It has been suggested that isosorbide dinitrate (ISDN)-induced venodilation could be ascribed to preferential accumulation of the agent in venous tissues, resulting in higher concentrations of nitric oxide (NO). Here, the authors investigated whether the venodilating effect of ISDN is associated with a preferential increase in plasma concentrations of NOx (NO2- and NO3-, stable end-products of NO) in venous blood than arterial blood. Plasma NOx was measured by high-performance liquid chromatography-Griess system with a sensitivity of 0.01 microM for NO2- and 0.1 microM for NO3-. Arterial and venous blood samples were obtained after coronary angiography from the aorta and right atrium of patients with or without ischemic heart disease. Nicardipine, a calcium channel blocker, was used as a non-NO-related arteriovasodilator. At 1 mg i.v., it did not cause any changes in NOx concentration in arterial and venous blood irrespective of hemodynamic changes. However, ISDN (3 mg i.v.) increased NO2- and decreased NO3- in both arterial and venous blood, with concomitant venodilation. Further analysis revealed that plasma NO increased in the pulmonary circulation and this increase was preserved after nicardipine and ISDN, and that ISDN, but not nicardipine, increased plasma NO3- in the pulmonary circulation. The authors did not detect higher concentrations of NOx in venous blood relative to their level in arterial blood. Further studies are necessary to clarify the kinetics of NO and NO-related compounds in the whole body.

Paradoxical decrease in plasma NOx by L-arginine load in diabetic and non-diabetic subjects.

L-arginine, a substrate of nitric oxide synthase, was infused (30 g/300 ml/30 min) to patients with or without type 2 diabetes to examine whether or not endothelial dysfunction expressed as attenuated depressor response to the substrate in diabetic patients may accompany attenuated plasma NOx (NO2- and NO3-; an index of NO formation) elevation. Decrease in blood pressure by L-arginine was significantly smaller in diabetic patients than that in non-diabetic patients, and increase in plasma cGMP level in diabetic patients tended to be smaller and retarded than non-diabetic patients. However, plasma NOx decreased in both groups in a similar degree without changes in urinary NOx excretion, implying that NOx in plasma moved to other compartments. These results indicate that plasma NOx could not be solely used as an index of NO formation by L-arginine load and that this paradoxical decrease in plasma NOx would require further examination extending to other NOx compartments.

Insulin resistance is associated with reduced nocturnal falls of blood pressure in normotensive, nonobese type 2 diabetic subjects.

To assess the relationship between insulin resistance and ambulatory blood pressure (BP) pattern, we determined glucose infusion rate (GIR) as a marker of insulin resistance using a glucose clamp method, and measured 24-h BPs in 25 normotensive, nonobese type 2 diabetic subjects. They were divided into two groups: 11 dippers and 14 nondippers. Clinical characteristics were similar in the two groups except for orthostatic fall in systolic BP. The median GIR level was significantly lower in nondippers than in dippers (P < 0.05). Spearman's rank correlation revealed that the GIRs were negatively correlated with the systolic, diastolic and mean BPs during nighttime (P < 0.05 or less), but not with daytime or whole day BPs. Moreover, based on a logistic regression analysis, the GIR as well as orthostatic fall in systolic BP discriminated independently between dippers and nondippers. Thus, our results suggest that insulin resistance is associated with decreased nocturnal BP fall in type 2 diabetic subjects.