The biosynthesis of three natural products, L-2-amino-4-pentynoic acid (propargylglycine) (1), 3-(3-isocyanocyclopent-2-enylidene) propionic acid (2), and (+)-(alpha)-(S)-amino-(2,5-dihydro-5(S)-methyl)-furan-2(R)-acetic acid (3) (furanomycin), produced by an unidentified Streptomyces, Trichoderma hamatum, and Streptomyces threomyceticus respectively, have been investigated. Although two prospective labeled precursors of the amino acid (1), {1-('14)C}-norvaline and {1-('14)C}-allylglycine were synthesized, and procedures for the isolation, detection and degradation of (1) devised, the elucidation of the biogenesis of (1) was hampered due to lack of production of (1) by the microorganism. The isonitrile acid (2) has been shown by using ('14)C- and ('3)H-labeled precursor incorporation experiments to be derived from the amino acid tyrosine which undergoes an oxidative cleavage of the aromatic ring followed by recyclization. In addition to specific incorporation studies with {('14)C}- and {('3)H}-labeled precursors, experiments using {('2)H}, {('18)O}, and {('13)C}-labeled precursors in conjunction with ('2)H-NMR and ('13)C-NMR have been undertaken to investigate the biosynthetic origin of the amino acid (3). The evidence indicates that (3) arises from one unit of propionic acid and two units of acetic acid.