Citrus limon of Rocca Imperiale IGP: “Proteomic fingerprint” and traceability of derived products

Trace proteins in fruit act as a distinctive molecular “fingerprint” that remains detectable in derived products, offering a reliable means of confirming product authenticity. In this study, we employed Combinatorial Peptide Ligand Libraries (CPLLs) to investigate both the Citrus limon peel of Rocca Imperiale IGP (Italy) and the corresponding Limoncello, a popular Italian liqueur produced in Calabria. By compressing the dynamic range of protein concentrations, CPLLs simultaneously reduce the abundance of highly expressed proteins and enrich rare ones, allowing a more comprehensive proteomic analysis. We tested different extraction protocols on C. limon peel and Limoncello prior to CPLL treatment at two pH values (2.2 and 7.2). Mass spectrometry revealed a broad panel of proteins in C. limon extracts, many of which were also identified in the commercial liqueur, underscoring their potential as molecular markers for traceability. Our findings show that CPLLs increase proteome coverage by over eightfold compared to controls, uncovering additional proteins related to stress response, primary metabolism, and secondary metabolite biosynthesis. The presence of overlapping proteins in both C. limon peel and Limoncello highlights the feasibility of CPLL-based proteomics for authenticating high-quality citrus products in terms of both geographical and varietal origin.