Identification of patulin-producing Penicillium isolates from decayed apples, their patulin production profiles, and the feasibility of high-purity patulin preparation.

This study presents a combined evaluation of patulin contamination by integrating fungal identification, assessment of toxin production, and the feasibility of obtaining high-purity patulin from biological sources. Fungal isolates recovered from naturally decayed apples collected from different agricultural environments in Türkiye were identified using MALDI-TOF MS and morphological characteristics, and were assigned to Penicillium expansum at a probable species level. The patulin production potential of the isolates was evaluated by inoculating them onto red, yellow, and green apple matrices under controlled laboratory conditions over an extended incubation period. Quantitative analysis using HPLC-UV, supported by LC-MS/MS confirmation, revealed substantial variability in patulin levels (7.58-49.80 mg/kg) depending on both isolate and apple matrix. Higher patulin levels were generally observed in yellow apple tissues, whereas lower levels were detected in red apples, suggesting a possible influence of matrix-related compositional factors. In addition, patulin was isolated and enriched using column chromatography with TLC-guided fractionation, yielding a high-purity compound (98.8%) with low moisture content (0.21%). Structural identity was confirmed by LC-MS/MS analysis. Overall, the findings indicate that patulin production is influenced by strain-level variability and matrix-related factors. Furthermore, the study demonstrates the feasibility of producing high-purity patulin suitable for in-house analytical applications. However, the produced material should not be considered a certified reference material, and further validation would be required for broader analytical use.