Methodological Modifications of the INFOGEST 2.0 Protocol in Plant-Based Food Studies: A Systematic Review

The INFOGEST 2.0 protocol is widely applied as a standardized static in vitro digestion model, and its use in studies involving plant-based foods has increased substantially in recent years. This systematic review aimed to identify, classify, and critically evaluate methodological modifications applied to the INFOGEST 2.0 protocol in studies investigating plant-based foods and supplements. A systematic search was conducted in the ScienceDirect, Scopus, and PubMed database, identifying 119 original research articles published between 2019 and 2024. Methodological modifications across the oral, gastric, and intestinal phases were extracted and analyzed according to the parameters altered and their underlying rationale. Approximately, half of studies (50.4%) deviated from standardized INFOGEST conditions, particularly regarding enzyme activity, pH, bile concentration, and digestion time. Importantly, these adaptations were not exclusively driven by the plant-based nature of the food matrices, but also reflected methodological factors related to the study population or objectives, including aging-related physiological conditions, specific experimental needs, the tracing of particular compounds (e.g., minerals and phenolic compounds), and blank subtraction. While such modifications may enhance physiological relevance or analytical accuracy, they introduce a trade-off with interstudy comparability, highlighting the need for clearer reporting practices and consensus-based extensions of the INFOGEST framework to balance physiological relevance with methodological consistency.