Plant pathogenic fungi cause important yield losses in crops. A proteomic approach was used to study the changes in the leaf proteome profile of the plant Mentha arvensis infected with a necrotrophic fungus, Alternaria alternata. High-resolution two-dimensional gel electrophoresis (2-DE) followed by colloidal Coomassie staining and mass spectrometric analysis was used to identify highly abundant proteins differentially expressed in response to fungal infection. From a total of 210 reproducibly detected and analyzed spots, the intensity of sixty-seven spots was altered, and forty-five of them were successfully identified by matrix assisted laser desorption/ionization time of flight-mass spectrometry (MALDI TOF/TOF MS/MS). Fifty-six percent of the identified proteins belonged to energy and metabolism whereas 29% were stress and defense related. Taken together, the results allow to assess changes at the proteomic level in the host due to the defense response. Results show an initial defense response, not strong enough to overcome the pathogenesis, which may be similar to other susceptible plant-pathogen interactions; however, cross-talks between various defense pathways, regulatory networks and physiological conditions are other important aspects to be considered.