Crocin enhances the quality of frozen-thawed boar sperm by modulating oxidative stress and glycerophospholipid metabolism

BACKGROUND: Cryopreservation is a fundamental tool for conserving endangered species and enhancing reproductive management in livestock; however, post-thaw sperm performance is easily influenced by several stressors. Crocin, a natural carotenoid, has antioxidant properties. However, its effects on frozen-thawed boar sperm remain insufficiently studied. OBJECTIVE: This study aimed to assess the role of crocin in improving the quality of frozen boar sperm by focusing on its mechanisms of regulating oxidative stress, mitochondrial membrane potential, and glycerophospholipid metabolism. MATERIALS AND METHODS: Boar sperm samples were supplemented with varying crocin concentrations (0, 0.5, 1, and 2 mM). Sperm kinematic parameters and motility were assessed using computer-assisted semen analysis, whereas the DNA fragmentation index and mitochondrial membrane potential were assessed using a fluorescence microscope. RESULTS: Treatment with 1 mM crocin significantly enhanced the kinematic parameters ( P < 0.05) and frozen sperm viability ( P < 0.05) while enhancing the integrity of the plasma- and acrosomal membranes ( P < 0.05). The oxidative stress indices, including reactive oxygen species, 8-hydroxy-2-deoxyguanosine, and malondialdehyde content ( P < 0.01), were effectively lowered by crocin. Furthermore, crocin enhanced the mitochondrial membrane potential and sperm DNA integrity ( P < 0.01). Metabolomic analysis showed that crocin affected several differential metabolites, including phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, and 2-lysophosphatidylcholine ( P < 0.05), highlighting its role in glycerophospholipid metabolism. CONCLUSION: Crocin markedly improved the quality of frozen boar sperm by regulating oxidative stress and lipid metabolism, offering new insights for optimising sperm cryopreservation. This study provides practical insights into the optimisation of boar semen cryopreservation in livestock breeding.