Naoko S Watanabe and Hideo Yamasaki
Nitrite (NO2-) contained in dietary foods has long been recognized for its toxicity as the causative agent of methemoglobinemia and also as a source of mutagenic nitrosamines. Because of these potential toxicities, nitrite as well as nitrate contained in foods and drinks are strictly limited by regulations in many countries. Recent studies have offered us to update our recognition of nitrite; nitrite is an important precursor for Nitric Oxide (NO) that is required for fundamental physiological activities including vasorelaxation. Although it is well established that green vegetables contain high amounts of nitrate, there has been controversy regarding the source of nitrite accumulation in fresh green vegetables. In this study, we investigated the dynamics of nitrite and nitrate contents in spinach leaf extracts to verify the mechanisms of nitrite formation. The time course of nitrite production in leaf extracts showed a reciprocal relationship with nitrate degradation, suggesting a conversion from nitrate to nitrite. The reaction strongly depended on temperature and it was suppressed at a low temperature. Sodium tungstate, a nitrate reductase enzyme inhibitor, was effective to suppress the conversion. Pre-sterilization by autoclaving or filter sterilization completely prevented the formation of nitrite as well as degradation of nitrate. We suggest that previous reports of nitrite accumulation can be attributed to microbial nitrate reductase activities that occur during the degradation spinach leaves.