T HE CURRENT research deals with optimization of the factors affecting lipase production under submerged culture system. The most efficient isolate R1 was identified depending on cultural and morphological characteristics together with 18S rRNA sequence as Rhizopus oryzae. Using one variable at a time, the maximum lipase activity (171.8 U/mL) was recorded in the presence of 1% fish-frying oil, mixture of peptone and yeast extract at pH 5 with 8% v/v of fungal inoculum after 4 days at 30°C. The screening of the most significant factors using Plackett-Burman design revealed that among ten variables, four, i.e. incubation temperature, inoculum size, incubation period and agitation speed, affected significantly (p-values ranged from 0.003 to 0.049) on the lipase activity. Optimization by using response surface methodology (RSM) through central composite design (CCD) resulted in the highest predicted lipase activity (216.2 U/mL) in which fermentation medium was inoculated with 8% inoculum size and incubated at 28°C under agitation speed of 150 rpm for 4 days.