LC-MS/MS Determination of Mono-Glutamate Folates and Folic Acid in Beer

A reliable LC-MS/MS method was developed for determination of folates in beer. Clean-up steps and chromatographic conditions were optimized to remove interferences and decrease the matrix effect. The work showed that an immuno-affinity column designed for folic acid (FA) was also suitable for the oxidized forms 10-formylfolic acid (10-HCO-FA) and 7,8-dihydrofolic acid (DHF), obtaining clean extract and improving the limit of detection. For the other vitamer forms, an Oasis MAX column was used. For chromatographic conditions, a RP-18 column with low ligand density showed satisfactory retention times; moreover, the use of methanol instead of acetonitrile lowered the limit of detection up to 0.1 μg l−1. The most popular beers in Italy were collected and analyzed; mainly oxidized mono-glutamate forms, in particular 10-HCO-FA and FA, were found, at levels from 12.4 to 98.6 μg l−1 and 1.4 and 11.9 μg l−1, respectively. The vitamers 5-formyl-tetrahydrofolate and 5-methyl-tetrahydrofolate were also detected.