The Synthesis of Biologically Active Pteroyloligo-γ-l-Glutamates (Folic Acid Conjugates)

Homogeneous pteroyltri-, pteroylpenta-, and pteroylhepta-γ- l -glutamates (folic acid conjugates) have been synthesized and, when desired, specifically labeled with tritium. The synthetic procedures consist of: ( a ) production of pteroic acid from folic acid by microbiological techniques; ( b ) preparation of [3',5',9- 3 H]pteroic acid; ( c ) synthesis of crystalline homogeneous N 10 -trifluoroacetylpteroic acid; ( d ) mixed anhydride condensation of N 10 -trifluoroacetylpteroic acid with tri-, penta-, and hepta-γ- l -glutamic acid tert -butyl esters prepared by peptide synthesis in solution; ( e ) purification of protected products by LH-20 column chromatography; and ( f ) removal of protecting groups and final purification of products by gel filtration. The compounds have been obtained in good yields and have been characterized by correct elemental analyses, amino acid analyses, ultraviolet spectral data, ascending paper chromatography, and high voltage electrophoresis. These data compare favorably with analytical information reported for natural products. Behavior of the synthetic materials in microbiological assays and enzyme studies of intestinal mucosal homogenates confirms their biological identity with natural conjugates.