A general method for characterizing naturally occurring folate compounds, illustrated by characterizing Torula yeast (Candida utilis) folates

A method previously found suitable for the chromatographic separation and identification of simpler folates has been extended and found suitable for separating and characterizing all the complex polyglutamyl folyl derivatives occurring naturally. Folates were characterized employing the combined use of analytical DEAE-cellulose chromatography, folylpoly-γ-glutamyl carboxypeptidase digestion, and differential microbiological growth response studies. An observed relation between the log phosphate concentration of the eluting buffer and the number of γ-glutamyl residues in successive pteroylpoly-γ-glutamyl derivatives provides a simple tool for a rapid and accurate identification of folate compounds from their elution profile. Twelve folate compunds present in Torula yeast (Candida utilis) were characterized employing this method; 80% of the total folates appeared to be pteroylpolyglutamates. The method characterizes not only the number of γ-glutamyl residues but also the state of reduction of the pteridine ring and the nature of the 1-C substituents attached.