Roast-Driven Coffee Proteome Changes Characterized by Bradford Assay, SDS-PAGE, and LC-MS

Coffee proteins are key precursors of roasting flavor. However, heat-driven changes in the bean proteome remain underexplored. This work aimed to investigate these changes and study proteomic markers of the coffee bean. The green and roasted coffee beans were quantified for their total soluble protein and compared by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and liquid chromatography-mass spectrometry (LC-MS) proteomics. The protein profiles identified by LC-MS were processed using principal component analysis (PCA) and partial least-squares discriminant analysis (PLS-DA) modeling to identify possible roast-sensitive protein markers. The alkaline-aided aqueous extract protein concentration was reduced from 14-23 g to 3-10 g/100 g dry weight (DW). SDS-PAGE showed dominant 17-26, 34-43, and 55-72 kDa bands weakened after roasting, while high molecular peaks (>180 kDa) were present only in roasted samples. In-solution tryptic digestion yielded nine protein groups. PCA scores revealed partial separation of green and roasted groups, while PLS-DA delivered unambiguous classification (Q² > 0.90 by cross-validation). The variable importance in projection scores highlighted that structural proteins in common plant beans are markedly down-regulated after roasting, indicating heat-induced structural disruption. The identified protein groups represent candidate markers associated with severe thermal treatment and provide possible molecular targets for investigating flavor precursor development.