Chromatographic determination of riboflavin and its derivatives in food

Three elution methods on two different reversed-phase C18 columns were developed to determine flavin11Derivatives of the 7,8-dimethyl isoalloxazine nucleus are defined also as “flavins”. atives 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 C18 column was not as effective as a Symmetry C18 column. Method A (mobile phase gradient of methanol–0.05 M ammonium acetate, pH 6.0 applied on an Alphabond C18 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 C18 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 C18 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.