The main objective of this research project was the production of laccase under optimum conditions, its partial purification and the effect of UV mutagenesis in Neurospora sitophila for hyper production of laccase. The temperature and pH of the incubation were 28 oC and 5.5 respectively. Growth medium employed to culture the Neurospora sitophila for the production of laccase was optimized through various experiments. The fermentation conditions like incubation period, moisture level, inoculum size, yeast extract and tween-20 were optimized. It was observed that the addition of yeast extract and tween-20 enhanced laccase production by Neurospora sitophila in corn cob. Neurospora sitophila produced maximum laccase 3.995U/mL/min in the growth medium of 70% moisture level, 5mL of the inoculum, 0.4% yeast extract and 0.2% of tween-20. The laccase produced by Neurosopora sitophila under optimized conditions was partially purified using ammonium sulfate precipitation.