AbstractThe present study was carried out to optimize the cultural conditions of phytase production by Aspergillus niger USM AI1 using palm kernel cake (PKC) as the substrate in a solid state fermentation (SSF) system. The optimized cultural conditions of 10 g of PKC with a particle size of ≤0.5 mm, an inoculum size of 1x105 spores/ mL, moisture content of 60% (v/w), and a mixing frequency of once every 48 hours produced 4.91±0.17 U/g dried substrate of phytase and 1.14±0.04 mg glucosamine/ g dried substrate of fungal growth. Maximum growth as well as enzyme production was recorded on the 4th day of cultivation at room temperature (30±2 ºC). The findings indicated that phytase production increased 171.3% after optimization (4.91±0.17 U/g dried substrate) compared to before optimization (1.81±0.16 U/g dried substrate). This study showed that PKC is a potential medium for phytase production in SSF.