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The effects of certain nutrients on natamycin production by Streptomyces natalensis in submerged batch culture were studied. The production of this antibiotic required glucose in the cultivation medium with a concentration of 20 g/l. On the other hand, the highest antibiotic production was obtained in a cultivation medium containing 0.05 g/l of potassium dihydrogen phosphate. Further increase in phosphate concentration resulted in a significant increase in biomass concomitant with lower antibiotic production. Among different N‐sources tested, only ammonium sulphate, sodium nitrate and beef extract were the suitable nitrogen sources in supporting the antibiotic production. Furthermore, a mixture of beef extract and yeast extract (8 g/l and 2 g/l, respectively) exhibited a synergistic effect in enhancing the natamycin production reaching about 1.5 g/l.