Fructose, glucose, and an equimolar mixture of both sugars affected differently hyphae thickness, biomass production and secretion of β-fructofuranosidase in Penicillium janczewskii. Reduced growth, thinner hyphae and visible injuries were early observed during fungal cultivation in fructose-containing medium, reaching the maximum between 12 and 15 days of culture. Total sugar content from the cell wall was lower when fructose was supplied and polysaccharides lower than 10 kDa predominated, regardless the culture age. Maximal inulinase and invertase activities were detected in culture filtrates after 12 days, excepting in the glucose-containing medium. Structural changes in cell walls coincided with the increase of extracellular enzyme activity in the fructose-containing medium. The fragility of the hyphae might be related with both low carbohydrate content and predominance of low molecular weight glucans in the walls. Data presented here suggest changes in carbohydrate component of the cell walls are induced by the carbon source.