The aim of this study was to optimize the culture conditions for the production of biological cyclic hexadepsipeptides (enniatins H, land MK1688) from Fusarium oxysporum KFCC 11363P. Tests of 10 complete or chemically defined liquid culture media revealed that Fusarium defined medium was the best for the production of enniatins (produced amounts: enniatin H, 185.4 mg/L; enniatin I, 349.1 mg/L; enniatin MK1688, 541.1 mg/L; and total enniatins, 1075.6 mg/L). On the eighth day after inoculation, the maximal production of enniatins was observed at 25 degrees C in Fusarium defined medium. The optimal carbon and nitrogen sources for producing biological cyclic hexadepsipeptides (enniatins H, I, and MK1688) were sucrose and NaNO(3), respectively, and their optimal concentrations were determined by the principle of response surface methodology. It was confirmed that using the optimized growth medium compositions increased the amounts of enniatins H. I. and MK1688, and total enniatins produced to 695.2, 882.4, 824.8, and 2398.5 mg/L, respectively. These findings will assist in formulating microbiological media useful for enniatin research. (C) 2010, The Society for Biotechnology, Japan. All rights reserved.