Plasma nitrite was evaluated as an indirect marker of NOx, using Griess reagent following Promega's Instructions for use of the Griess reagent system [36]. First, 200 μl of plasma were mixed with 500 μl of 0.1% of N-1-napthylethylenediamine dihydrochloride (NED) in water and left in the dark for 5 min, then 500 μl of 1% sulfanilamide were added to 5% phosphoric acid and kept in the dark again for 5 min. A slightly pink color was produced with an absorbance reading at 520 nm. Nitrite in plasma was calculated by comparing with the absorbance of standard sodium nitrite (NaNO3) (0-40 μmol/L).