This study investigates the significance of various parameters, including water content, total acid content, sugar content, polyphenol content, antioxidant activity, anthocyanin content, and flavonoid content, on the quality of apples and their resistance to Penicillium expansum infection and subsequent patulin production. The research was conducted on four apple cultivars, namely, the traditional cultivars 'Wagener' and 'Ilzer Rosenapfel', and the commercial cultivars 'Jonagold' and 'Idared'. Among the cultivars, 'Wagener' demonstrated the highest values for total soluble matter (15.9%), sugar content (15.22 ± 0.03%), total polyphenol content (707.63 ± 22.81 mg/kg), and antioxidant activity (0.29 ± 0.00 mmol TE/kg dw). 'Jonagold' exhibited the highest anthocyanin content (5.76 mg/kg) and total flavonoid content (104.74 ± 3.11 g CE/kg), while 'Idared' displayed the highest acid content (0.13 g/100 g). Furthermore, the presence of patulin, a mycotoxin produced by Penicillium expansum in infected apple fruits, was analyzed using liquid chromatography coupled to tandem mass spectrometry. The levels of patulin varied among the cultivars, with 'Wagener' exhibiting the highest concentration (18592 ± 101.82 µg/kg), followed by 'Idared' (4732.4 ± 57.10 µg/kg), 'Jonagold' (292.56 ± 20.93 µg/kg), and 'Ilzer Rosenapfel' (130.92 ± 0.06 µg/kg). In conclusion, the comparative analysis of the data provides valuable insights into the composition, quality attributes, and potential resistance to Penicillium expansum infection among different apple cultivars. These findings have implications for the selection and cultivation of apple varieties with desirable characteristics, such as taste, antioxidant potential, and reduced mycotoxin contamination. This research was funded by the Croatian Science Foundation (UIP-2020-02-8461).