AbstractFactors affecting the β‐galactosidase production byPenicillium notatum1 were studied using fermentation media of different chemical composition. The medium containing lactose, salts, peptone and yeast extract with intitial pH 2.5 was selected as the best for enzyme production. Monobasic ammonium phosphate (0.9%) was found to be the best inorganic nitrogen source for lactase production. Various extraction media and metabolic inhibitors were examined for effective releasing of β‐galactosidase from the fungal cells. Using a simple method of mycelium extraction with 0.1 Triton X‐100, it was possible to obtain about 4‐fold higher amounts of enzyme in the cell free extracts, than those excreted into the post‐cul