Virgin olive oil samples with similar oxidative stabilities and fatty acid compositions were stored for 24 months. Changes in the lipid substrate were followed by peroxide value and K(232) measurements. HPLC was used to evaluate changes in the alpha-tocopherol, pigment, and squalene contents. Total polar phenol content was measured colorimetrically. The loss of alpha-tocopherol and carotenoids was comparable with that of polar phenol content, suggesting an active participation in autoxidation. The limited role of squalene in autoxidation was further confirmed using an olive oil model and in the presence of alpha-tocopherol. Pheophytin a degradation was high, although spectrometric estimation of chlorophyll content did not indicate so. Evaluation of pheophytin a activity at three different levels of addition on the oil model indicated a concentration-dependent antioxidant role more pronounced at elevated temperatures, which could be partially due to the activity of certain degradation products.