A synthetic medium, containing L-proline, glycine, L-arginine, L-cystine, L-glutamine, L-histidine, L-isoleucine, L-leucine, L-lysine, L-methionine, L-phenylalanine, nicotinamide, thiamine, glucose, sodium citrate, Na2HPO4, KH2PO4, (NH4)2SO4, MgSO4∙7H2O, FeSO4∙7H2O, and CaCl2∙2H2O, was developed. It supported growth and beta hemolysin production by the 681C strain of Staphylococcus aureus when the culture was incubated under carbon dioxide. L-Threonine, L-tyrosine, i-inositol, folic acid, riboflavin, pyridoxal HCl, choline Cl, and D-Ca pantothenate were added for obtaining good growth and toxin formation under air. L-Proline, L-glutamine, and L-cystine were the absolute requirements for the production of beta hemolysin under carbon dioxide. Carbon dioxide stimulated the production of beta hemolysin, but invariably resulted in lower cell yield.

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