5-Aminolevulinate acid improves boar semen quality by enhancing the sperm mitochondrial function

5-Aminolevulinic acid (5-ALA) is a crucial metabolic intermediate that affect mitochondrial function in somatic cells. However, the mechanisms by which 5-ALA regulates boar reproductive performance remain unclear. The effect of dietary supplementation of 5-ALA on boar semen quality and reproductive performance were investigated in this study. Forty-five boars were randomly assigned to a control group and four 5-ALA-treatment groups (125, 250, 500, and 1000 mg/kg/d). After nine weeks of treatment, serum and semen samples were collected from the boars for analysis. Results showed that the supplementation of 5-ALA to boar diet significantly (p < 0.05) increased semen volume, total sperm count, and sperm motility while reducing the proportion of abnormality (p < 0.05). Specifically, 250 mg/kg/d 5-ALA treatment significantly (p < 0.05) reduced sperm DNA oxidative damage and elevated serum testosterone levels. Metabolomic analysis revealed that 250 mg/kg/d 5-ALA supplementation significantly increased (p < 0.05) the levels of tricarboxylic acid (TCA) cycle intermediates such as malate and isocitrate, while significantly (p < 0.05) reducing harmful metabolites such as N-acetyl phenylalanine that negatively impact sperm quality and fertility. Notably, 250 mg/kg/d 5-ALA supplementation upregulated the protein levels of mitochondrial oxidative phosphorylation (OXPHOS) complex subunits (NDUFB8, SDHB, UQCRC2, MTCO2, and ATP5A1) and improved sperm mitochondrial membrane potential, adenosine triphosphate (ATP) levels, and complex IV activity (p < 0.05). Regarding reproductive performance, compared to the control, the percentage of piglets born alive increased by 2.97 %, the farrowing rate improved by 10.59 %, and the occurrence of mummified fetuses decreased by 1.07 % in the 5-ALA-treated groups (p < 0.05). Additionally, 250 mg/kg/d 5-ALA had a long-term beneficial advantage on boar semen quality parameters. Accordingly, dietary supplementation of 5-ALA, specifically, 250 mg/kg/d 5-ALA improved boar semen quality by enhancing seminal plasma metabolome, sperm mitochondrial function and overall reproductive performance.