Comparative proteomic profiling of faba bean tissues using an optimized extraction method and data-independent acquisition mass spectrometry

Faba beans ( Vicia faba ) have demonstrated potential to replace traditional animal-based protein sources. The quantity and quality of protein found in faba beans makes them a promising candidate for a sustainable protein alternative. However, the understanding of their proteomic profile remains limited. In this study, we present an optimized proteomic workflow for faba beans, comparing the extraction efficiency of detergent-based (T-PER) vs chaotrope-based (Urea) extraction methods. We conducted liquid chromatography-mass spectrometry (LC-MS) using data-independent acquisition (DIA) to improve coverage and analysed the protein profiles of different faba bean tissues: dried mature whole beans, freeze-dried fresh whole beans, and freeze-dried seed coats and cores (cotyledons). Of the two methods, urea extraction yielded higher protein amounts, identified more protein groups and exhibited lower coefficient of variation and higher replicate correlations. This study represents the first proteomic analysis to employ a DIA-MS approach in conjunction with an annotated proteome database for faba beans. Future studies can utilize this optimized workflow for the functional characterization of faba bean proteins, advancing the exploration of sustainable plant-based protein sources. This study provides a valuable resource for advancing proteomic research of legume species, contributing to more efficient proteomic profiling in plant research. Optimized chaotrope-based method improved faba bean protein extraction efficiency DIA-MS enabled high-resolution proteomic profiling of faba bean tissues First comprehensive proteomic comparison among faba bean tissues Tissue-specific proteomics revealed distinct functional specialization in faba beans