The physiology of a colony of Pleurotus pulmonarius grown on potato dextrose agar overlaid with a Cellophane membrane (PDA-WC) was studied. On PDA-WC, the colony presented higher biomass density and productivity of fruit body formation (4.84+/-0.23 mg mycelial biomass/cm(2) and 23.7+/-3.12 mug ml(-1) h(-1), respectively) than that which developed on potato dextrose agar without Cellophane (PDA-OC; 0.26+/-0.01 mg mycelial biomass/cm(2) and 10.8+/-1.57 mug ml(-1) h(-1), respectively). In cultures developed on PDA-WC, intracellular laccases and beta-1,3-glucanases activities were lower [12+/-0.9 arbitrary units (AU)/g mycelial biomass and 1.33+/-0.1 international units (IU)/g mycelial biomass, respectively] than those observed on PDA-OC (20.65+/-1.0 AU/g mycelial biomass and 3.67+/-0.2 IU/g mycelial biomass, respectively). In cultures developed on PDA-WC, intracellular protein and glycogen concentrations were lower (1.9+/-0.9 and 117+/-3.5 mg/g mycelial biomass, respectively) than those observed on PDA-OC (14.3+/-1.1 and 347+/-2.9 mg/g mycelial biomass, respectively). The radial growth rate and the content of glucans in the cell wall were not significantly different between cultures developed on PDA-WC and PDA-OC. These results show that the use of the Cellophane as a tool to study in vitro fungal physiology might affect the interpretation of experimental results, since the physiology under otherwise similar conditions was different on medium with and without Cellophane.