Green extraction of RuBisCO-rich leaf protein for sustainable food systems: process efficiency, functional quality, and sustainability assessment

Leaf biomass, particularly ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO)-enriched fractions, represents a viable secondary source of protein for sustainable food systems; this review presents a focused, comparing green extraction approaches deep eutectic solvents, enzymatic pretreatments, ultrasound- and microwave-assisted methods, and hybrid combinations with respect to protein yield, purity and functional properties. Enzymatic pretreatment most consistently enhances protein recovery while preserving solubility and emulsification capacity; ultrasound and microwave assistance reduce processing time but require careful parameter control to avoid denaturation. Key barriers to scale-up include inconsistent reporting of performance metrics, solvent-recovery inefficiencies, limited pilot-scale validation, and gaps in techno-economic and safety data. Finally, a forward-looking roadmap proposed to overcome persistent challenges such as solvent recovery and the absence of standardized protocols, highlighting the promise of artificial intelligence (AI)-guided process optimization and tailored solvent design. Collectively, this review provides a distinct framework for advancing leaf protein valorization toward sustainable food system transformation.