Abstract

Cultivated meat has the potential to reduce the environmental footprint of conventional meat production; however, the high cost of culture media remains a major obstacle to large-scale implementation, motivating efforts to better understand and optimize cellular metabolism. Here, we present iSSC2487, a curated, highquality genome-scale metabolic model of Sus scrofa comprising 2487 genes, 9124 reactions, and 6790 metabolites distributed across nine compartments, which correctly reproduces the essentiality of the nine classical essential amino acids. Measured uptake and secretion rates from 40 L fed-batch cultures of porcine embryonic stem cells were used to constrain dynamic flux balance (dFBA) simulations, which reproduced the observed bioreactor trajectories of biomass and extracellular metabolites. Using this data-constrained framework, we mapped the intracellular routing of carbon and nitrogen, resolved how it differs across the early, exponential, and late growth phases, and identified medium modifications that reduce cost. The flux distributions revealed a progressive rerouting of carbon and an overflow metabolism centered on glutamine, in which transamination generated the secreted alanine, glutamate, and aspartate, with byproduct secretion declining as growth rates decreased. iSSC2487 identified a minimal medium containing 27 essential nutrients. We provide a feed formulation aligned with metabolic requirements, which would cost 10 USD L-1, whereas mixed-integer linear programming (MILP)-based optimization further lowered the predicted cost to 6.5 USD L-1 (35% reduction). Although these model-guided predictions remain to be tested, the reconstruction provides a general scaffold for future work, including omics integration and bioprocess optimization in porcine cell culture.