Determination of ascorbic and dehydroascorbic acid in lean and fatty fish species by high-performance liquid chromatography with fluorometric detection

Abstract

Ascorbic acid (AA) and dehydroascorbic acid (DHAA) were analysed in fish muscle by high performance liquid chromatography (HPLC) coupled with fluorescence determination using 4,5-dimethyl-ophenilenediamine (DMPD) as a derivative agent. The use of the DMPD in combination with isobutanol extraction eliminated many matrix interferences and increased significantly the specificity and sensitivity of the method. A strong pH dependence in the derivatization reaction was also demonstrated. Satisfactory results in terms of sensitivity, stability and reproducibility were obtained. The analyses demonstrated that the relative ratio of AA and DHAA in post-mortem fish muscle differs among fish species and seems to be related with the susceptibility to oxidation.