A solid-state fermentation system, using polyurethane foam as an inert carrier, was used for the production of nuclease P1 by Penicillium citrinum. Optimization of nuclease P1 production was carried out using a synthetic liquid medium. After a hue-step medium optimization using a fractional factorial experimental design, nuclease P1 productivity and specific activity are greatly increased. Nuclease P1 productivity and specific activity are ninefold and fourfold higher, respectively, than in solid-state fermentation of wheat bran and 500-fold and 20-fold higher, respectively, than in submerged fermentation. Additional advantages of the solid-state fermentation process using inert carrier are discussed.