Efforts were made to utilize the waste/by-product of two agro-process industries namely pressmud from sugar processing industries and spent yeast from distilleries manufacturing ethanol from cane molasses, for the production of microbial invertase. Our experimentation indicated that these two sources could be ideally utilized for the production of invertase through solid substrate fermentation (SSF). SSF with spent yeast had given highest specific activity of 430 U/mg in 72 h of fermentation. Inoculum percentage of yeast cells on pressmud was optimized as 50% (w/w) with a combination inoculum of spent yeast and fresh cultured yeast at a ratio of 7:3. Crude enzyme was characterized for optimum pH and temperature and maximum activity was recorded at pH 5.0 and at a temperature of 40°C. Impacts of metal ions and detergents on invertase action were studied in which Mn2+, Fe3+, Al3+ and detergents had enhanced the activity of the enzyme whereas Cu2+ and Zn2+ inhibited the enzyme activity. Purification of 9.8 folds was obtained by using three phase partition method.