Determination of the nitrite reductase activity

The nitrite reductase activity was determined by measuring the reduction of KNO₂ by methyl viologen (Moir et al., 1993). Sample preparation and processing was performed as specified above for the determination of the nitrate reductase activity. Nitrite reduction was initiated by adding 100 μl of a solution containing 4 mg/ml sodium dithionite, 4 mg/ml sodium bicarbonate and 200 μM KNO₂. After 30 min, the assay was stopped by vigorous shaking until the dark blue color disappeared. The amount of nitrite remaining in the reaction mixture was measured by using the Griess assay (Nicholas and Nason, 1957), and was quantified by comparing the values to a standard curve obtained with nitrite. All experiments were performed in triplicates and repeated at least three times. All results are presented as mean ± standard deviation (SD).