The plants have developed several defense mechanisms to counteract pathogens. Among others, it includes activation of antioxidant enzymes like β-glucosidase and guaiacol peroxidase (GPX). These proteins participate in the oxidation of phenolic compounds, contributing to their increased fungitoxicity. The study aimed to analyze changes in the activity of β-glucosidase and GPX in four genotypes of winter oilseed rape (Mendel, Monolit, Polka, line L1425) inoculated with seven isolates: Alternaria brassicicola, Alternaria brassicae, Alternaria alternata (3 strains), Ulocladium chartarum (syn. A. chartarum), and Cladosporium cladosporioides. We noted that the varieties of oilseed rape, tested fungal species, and time of the plant material collection had significant (P < 0.001) effect on the activity of β-glucosidase and GPX per protein and fresh matter content comparing to the control group. A. brassicicola caused the highest mean increase in β-glucosidase and GPX activity in all examined genotypes, while other pathogens had a lower impact. Significantly lower β-glucosidase activity inoculated by various pathogens was noted between the L 1425 line and other varieties. GPX activity was in the opposite, the lowest activity was recorded in the Mendel variety, and the highest in the L 1425 line. Keywords Oilseed rape • β-glucosidase • Guaiacol peroxidase • Alternaria black spot Communicated by M. Horbowicz.