Chromatographic determination of riboflavin and its derivatives in food

Abstract

Three elution methods on two different reversed-phase C 18 columns were developed to determine flavin 1 1 Derivatives of the 7,8-dimethyl isoalloxazine nucleus are defined also as “flavins”. derivatives in raw egg white, raw egg yolk, egg powder, pasteurised milk, fermented milk products and liver (chicken, calf and pig). Additionally, 11 thin-layer chromatography solvent systems were used to confirm presence of flavins detected in assessed products. It was found that an Alphabond C 18 column was not as effective as a Symmetry C 18 column. Method A (mobile phase gradient of methanol–0.05 M ammonium acetate, pH 6.0 applied on an Alphabond C 18 column) can be used for determination of flavin adenine dinucleotide, flavin mononucleotide, riboflavin 4′,5′-cyclic phosphate, riboflavin, 10-formylmethylflavin and 10-hydroxyethylflavin in products that do not contain 7α-hydroxyriboflavin. Method B (mobile phase gradient of methanol–demineralized water, on an Alphabond C 18 column) can be useful to separate flavin coenzymes from other flavin compounds or to confirm the presence of 7α-hydroxyriboflavin and 10-hydroxyethylflavin in analysed samples. Method C (mobile phase gradient of methanol–0.05 M ammonium acetate, pH 6.0, on a Symmetry C 18 column) allows separation of all flavins detected in tested products: flavin adenine dinucleotide, flavin mononucleotide, riboflavin 4′,5′-cyclic phosphate, riboflavin, 10-formylmethylflavin, 10-hydroxyethylflavin, 7α-hydroxyriboflavin, riboflavin-β- d-galactoside and riboflavin-α- d-glucoside.