Thermo-Dynamic Flux Balance Analysis, a novel approach to simulate circadian metabolism

Circadian metabolism arises from complex, time-dependent interactions across organs, yet experimental characterization in humans remains limited. Current whole-body metabolic models are either too large for dynamic simulation or insufficiently detailed to capture temporal physiology. We developed a multi-tissue human metabolic reconstruction, HGEM1.19+, an extensively curated model for dynamic simulation. We introduced Thermo-Dynamic Flux Balance Analysis (tdFBA), a novel formulation that integrates thermodynamic constraints, organ-specific enzyme capacities, solubility limits, osmotic balance and pH buffering. Murine circadian transcriptomics and metabolomics were integrated with human datasets to parameterize temporal dynamics. A genetic algorithm optimized internal parameters under CHOW diet conditions and model performance was evaluated using circadian metabolomics and gene-sage patterns. The framework recapitulated key metabolic oscillations under CHOW diet and partially reconstructed perturbed states including HFD, BMAL1 knockout, and tissue-specific re-entrainment. tdFBA enables physiologically grounded, multi-organ dynamic simulations and provides a foundation for exploring human circadian metabolism and perturbation responses.