Fluorometric determination of ascorbic acid in the absence of the oxidant in juices of common citrus

A simple and sensitive fluorimetric method for the determination of ascorbic acid (AA) in juices of common citrus is described. The method is based on the condensation reaction between AA and o-phenylenediamine (OPDA) in the abscence of the oxidant. The keto-form of AA reacts with OPDA in alkaline medium and forms an N-heterocyclic compound with large π-bond system, which can emit strong fluorescence. The fluorescence intensity is measured at exitation and emision wavelengths of 330nm and 430nm, respectively. Under optimum conditions, a linear relationship is obtained between the fluorescence intensity and the concentraction of AA in the range 0.05–50µg mL-1. The detection limit (3σ) was found to be 0.54µg mL-1 of AA (σ from 5 determination of 10µg mL-1). A relative standard deviation of 1.4% was recorded for 8 measurements of 1µg mL-1 standard AA solution. The efect of the reaction time is also studied. The fluorescence intensity of the system reached a maximum immediatelly after all the reagens were added and remained stable at least for 45 minutes. This method is simple, sensitive and easy for the determination of AA in juices of common citrus. This is first report that analyses of AA in biological samples were done without oxidant.