Dynamic Nutrient Consumption of Cultured Meat in Bioreactors: Metabolomics-Guided Optimization for Enhanced Cell Proliferation and Stemness.

Cultured meat offers substantial advantages over conventional livestock production, including slaughter-free production and enhanced food safety. However, current understanding of dynamic nutrient consumption and metabolite accumulation in large-scale bioreactors remains inadequate. To address this gap, nontargeted metabolomics was employed to characterize the dynamic alterations in conditioned medium from an 80 L stirred-tank bioreactor (STR) and to identify key metabolites regulating muscle stem cell proliferation. A total of 1761 metabolites were detected; leucine (Leu) and arginine (Arg) were identified as having high variable importance in projection (VIP) scores and pathway enrichment analysis. Functional validation in both 2D monolayer and 3D suspension shake flask systems confirmed that targeted supplementation with these two amino acids significantly enhanced muscle stem cell proliferation and dose-dependently upregulated the stemness marker PAX7. This study elucidates dynamic metabolic shifts in a scaled cultured meat bioreactor, providing a crucial theoretical and data foundation for medium optimization and dynamic bioprocess control in industrialized cultured meat production.