The biosynthesis of the spermidine and guanidino groups has been studied with carbon-14 and carbon-13 labeled intermediates. Arginine, citrulline and ornithine are incorporated in good efficiency. The guanidino group of arginine and the ureido group of citrulline both label the guanidino group on the hexose sugar. None of the ureido groups in the antibiotic was enriched. It is likely that citrulline is converted to arginine before use in the biosynthesis. Arginine, citrulline and ornithine are incorporated as the four carbon unit of spermidine. All of the labeled 5 carbon from ornithine or from citrulline appears adjacent to the secondary amine in the four carbon unit of spermidine. This would indicate that unbound putrescine is not an immediate precursor of spermidine.