Applicability of microbiological assay and affinity chromatography purification followed by high‐performance liquid chromatography (HPLC) in studying folate contents in rye

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Abstract
In this study the folate contents in several rye varieties and their fractions were determined. Total folate content was analysed microbiologically using chloramphenicol‐resistant Lactobacillus rhamnosus as the growth organism. The amount of folate in rye grains was high, 630–780 µg kg−1, and variation between varieties was 8% (% CV). Affinity chromatography purification (with folate‐binding protein from bovine milk) followed by high‐performance liquid chromatography (HPLC) analysis proved to be an effective method for determining folate vitamers. 5‐Formyltetrahydrofolate showed a lower affinity to the folate‐binding protein than other vitamers; in order to obtain recoveries over 90%, the total folate load should not exceed 25% of the column capacity. Owing to intefering compounds, fluorescence detection was not always as specific as UV detection and thus not adequate for peak identification and quantitation alone. Formylated folates and 5‐methyltetrahydrofolate were the most abundant vitamers in rye, with tetrahydrofolate and folic acid representing only a minor proportion. However, different milling fractions had different folate vitamer distribution patterns. Both microbiological and HPLC methods were useful and reliable in determining folates in rye.© 2001 Society of Chemical Industry