High-performance liquid chromatography with fast-scanning fluorescence detection and post-column on-line photoderivatization for the analysis of folic acid and its metabolites in vegetables

Abstract

The photochemical behavior of folic acid and its metabolites, tetrahydrofolic and 5-methyltetrahydrofolic acids has been followed by high performance liquid chromatography, coupled with a fast scanning fluorimetric detector (HPLC-FSFD). The main photoproducts generated were characterized through accurate mass measurement using liquid chromatography-mass spectrometry (HPLC-MS). The effect on photogeneration processes of irradiation time, and the hydrogen peroxide concentration, in presence and in absence of N2 stream, was also studied. A chromatographic method with post-column on-line photoderivatization has been developed for the analysis of folic, tetrahydrofolic and 5-methyltetrahydrofolic acids. Linearity ranges from 50 to 1000ng·mL−1 for folic acid, and from 100 to 1000ng·mL−1 for tetrahydrofolic and 5-methyltetrahydrofolic acids have been established. Detection limits of 7.5ng·mL−1, 15.3ng·mL−1 and 22.9ng·mL−1 were calculated for folic acid, tetrahydrofolic acid and 5-methyltetrahydrofolic acid respectively. The method has been satisfactorily applied to cereals, spinach and tomato analysis. Serum samples analysis was also tested.