The present note reports the preparation of a novel A23187 analog, demethylamino A23187 (named cezomycin), via controlled biosynthesis. A23187 is an ionophore antibiotic that transports alkaline-earth cations across biological membranes, and its analogs help elucidate action mechanisms. For preparation, L-tryptophan (1 g/liter at 24 hours) was added to the culture medium of Streptomyces chartreusis NRRL 3882 (comprising glucose, Casamino acids, yeast extract, and meat extract, pH 7.2), which inhibited A23187 production (40 mg/liter) and favored cezomycin formation (100–150 mg/liter). Spectroscopic data confirmed cezomycin lacks the 3-methylamino group: mass spectrum showed a major peak at m/z 494 (corresponding to A23187 minus NHCH3); 1H NMR lacked the methyl signal (δ 2.95 ppm) and nitrogen-bound hydrogen signal (δ 8.1 ppm) of A23187's methylamine group; 13C NMR lacked the 30 ppm signal from A23187's nitrogen-bound methyl. The biosynthesis mechanism involves tryptophan metabolism producing 3-hydroxyanthranilic acid (HAA) and anthranilic acid: excess anthranilic acid competitively inhibits HAA hydroxylation, leading the carboxylic acid synthon (with spiroketal and pyrrole functions) to couple with HAA instead of 6-methylamino 3-hydroxyanthranilic acid (MAHAA), forming cezomycin. This is supported by adding anthranilic acid (0.5 g/liter) inhibiting A23187 and favoring cezomycin, while adding anthranilic acid plus HAA restored normal A23187 production. The cezomycin-Ca complex in methanol was observed via UV spectroscopy, with equilibrium constant determination ongoing. A preliminary study indicated cezomycin has calcium transport activity similar to A23187.