Metabolomic insights into functional protein mediated salt stress adaptation in the osmophilic yeast Zygosaccharomyces rouxii

High salinity inhibits microbial growth by disturbing osmotic balance, redox equilibrium, and membrane integrity. Zygosaccharomyces rouxii , an osmophilic yeast widely used in high-salt fermented foods, has evolved adaptive responses to such stress, but the global metabolic mechanisms underlying these responses, especially under functional protein supplementation, remain poorly understood. In this study, an untargeted UPLC-HRMS-based metabolomics approach was used to investigate the metabolic responses of Z. rouxii to high-salt stress and functional protein supplementation. Multivariate analysis revealed significant metabolic reprogramming at 120 g/L NaCl, while functional protein supplementation induced a distinct metabolic state beyond that caused by salt stress alone. Pathway enrichment analysis showed that protein supplementation markedly enhanced amino acid metabolism, antioxidant-related pathways, and nutrient transport, while preferentially modulating lipid metabolism associated with membrane remodeling. In particular, pathways related to unsaturated fatty acids and glycerolipids were finely regulated, suggesting improved membrane flexibility under hyperosmotic conditions. Collectively, these findings indicate that functional protein supplementation enhances coordinated metabolic regulation in Z. rouxii , thereby improving metabolic flexibility and salt stress tolerance. This study provides mechanistic insights into osmotic stress adaptation and proposes a feasible strategy to enhance yeast robustness under high-salt conditions.