A new method to measure nitrate/nitrite with a NO-sensitive electrode

Abstract

There are different methods to measure the unstable molecule nitric oxide (NO). We will describe a new sensitive method to measure NO by reconversion of nitrate/nitrite to NO, which will be determined with an amperometric Clark-type electrode. Nitrate and nitrite are the degradation products of NO. First, nitrate is enzymatically converted to nitrite with the use of the nitrate reductase. Second, nitrite is reduced to equimolar NO concentrations by an acidic iodide solution. The detection limit of the electrode in an aqueous solution was 2 nmol/l NO (meaning the threshold was depending on the volume added: 500 μl of a 0.2 μmol/l nitrite solution added to a 10-ml bath). This method provides the ability to assess basal and agonist-stimulated NO releases of different biological models. We measured basal and carbachol-stimulated NO release of native endothelial cells from porcine coronary arteries and porcine aortic endothelial cell cultures. Moreover, it was possible to measure the nitrate/nitrite concentration in the coronary effluent of a guinea pig heart. In conclusion, we present a valid, highly sensitive new method of measuring nitrite/NO in biological systems with a commercially available electrode.