Development of a HPLC method for the simultaneous analysis of riboflavin and other flavin compounds in liquid milk and milk products

A high-performance liquid chromatography method with fluorescence detection was developed for the quantification of riboflavin (RF), flavin mononucleotide (FMN), flavin adenine dinucleotide (FAD) and their photodegradation products, lumichrome (LC) and lumiflavin (LF), in liquid milk and milk products. Both sample preparation and chromatographic separation were studied to avoid acidic conditions that proved to affect flavin stability and degrade FAD into FMN. The sample preparation includes centrifugal skimming and ultrafiltration steps and is suitable for routine application. Linear response was obtained for individual flavins in the respective concentration ranges of interest and relative standard deviation (RSD) was lower than 5%, except for FAD (RSD 11%). The recovery ranged between 80–100%. The proposed method proved to be suitable for assessing flavins in commercial liquid milk and fermented milk products, and for monitoring the degradation of FAD, FMN and RF and the formation of LF and LC in bottled milk exposed to light during shelf storage.