BACKGROUND: The membrane filtration process enables the treatment of wastewater, producing permeate which is less polluted. However, disposal is usually required for the retentate, which is produced as a concentrated constituent along with the permeate. In this study, the authors explored the possibility of reusing, rather than disposing of, the retentate of pre-filtered palm oil mill effluent (POME) as a fermentation substrate in protease production by a wild type strain of Aspergillus terreus IMI 282743. In addition, the quantitative and interactive effects of the concentration factor for retentate, temperature, inoculum concentration, and fermentation time on the optimization of protease production were investigated using response surface methodology (RSM). RESULTS: Using RSM, the optimum conditions were found to be a concentration factor of 7.27, temperature of 37.95 degrees C, inoculum concentration of 1.30% (v/v) and fermentation time of 3.83 days. The protease production was increased 4.37-fold in comparison with the results obtained under non-optimized conditions. CONCLUSION: To a certain extent, protease production could be enhanced with an increase in concentration factor and temperature, and a decrease of inoculum concentration and fermentation time. Also, POME retentate was found to be a good substrate for protease production with high product activity and without nutrient supplementation. (c) 2009 Society of Chemical Industry