Folic Acid Derivatives in Yeast.

Dried yeast has found wide acceptance as a rich source of protein and vitamin B, and has been successfully incorporated into many foods. Up to 2.5% dried yeast is frequently used in cereals and canned vegetables for infants and children. Lesser amounts are added to breads, cereals, soups, and other enriched products for adult consumption for both flavoring and nutritional purposes. The National Formulary (1) sets minimum specifications on both brewers’ dried yeast and primary dried yeast for protein at 45%, niacin at 300 pg per g, riboflavin at 40 fig per g, and thiamine at 120 pg per g. Nutritionally significant quantities of pantothenic acid, pyridoxine, biotin, and folic acid may also be present in 10 g, a portion of reasonable size. The average total folic acid content of brewers’ dried yeast is reported to be about 20 pg per g (2). However, not all of the chemical forms of folic acid (folates) that comprise the total content (determined microbiologically) are equally potent for animals. A proper evaluation of the various derivatives of folic acid and their conjugated forms, each contributing some biological activity, and their added or cumulative potency for humans is of continuing importance. No single chemical or microbiological method for the assay of total natural folates exists. Such a method, to be useful for evaluation of folic acid activity in human nutrition, should provide for the quantitative determination of each of the biologically active compounds. Chemical methods have not yet been applied. Microbiological methods, which are inherently dependent on the variable growth-promoting activity of the many derivatives for assay organisms, give results that are difficult to assess quantitatively. Various substituted forms of naturally occurring folates have been characterized by applying special procedures during extraction, chromatography, and assay to protect the labile forms from oxidation and heat destruction. For example, appreciable quantities of the labile WO-formyltetrahydrofolate and N5-methyltetrahydrofolate have been isolated and identified in chicken liver extract (3). In earlier studies in which protective techniques were not used, it is possible that only the relatively stable iV5-formyltetrahydrofolate conjugates had been properly identified (4). Naturally occurring folates from various other sources such as bacterial cells, human blood, horse liver, mouse liver, plant materials, yeast, and gas glands of Physalia physalis L. have been reported (4-10). The majority of the naturally occurring folates, previously identified by procedures designed to provide complete protection from oxidative degradation, have been the tetrahydro forms. The oxidized forms (dihydrofolates and folates) have been referred to as artifacts resulting from nonenzymatic oxidation of the natural tetrahydro forms. The purpose of the present investigation is to characterize the pattern of folic acid derivatives in primary dried yeast.1 A chromatographic procedure similar to that employed by Noronha and Silverman (3) was used to fractionate a yeast extract. The fractions were subsequently assayed by differential microbiological techniques before and after digestion with a y-glutamyl conjugase preparation from chicken pancreas. Antioxidants were used throughout the procedure, and autoclaving temperatures above 100” were avoided to protect the labile yeast folates from oxidative degradation. A characterization of the folates in the separate fractions is presented, and their approximate degree of conjugation and type of substitution are given. A tabulation of total activities of all fractions is included.