The wide use of glyphosate-based herbicides (GBHs) has become a matter of concern due to its potential harmful effects on human health, including men fertility. This study sought to investigate, using the pig as a model, the impact of pure glyphosate and its most known commercial formulation, Roundup, on sperm function and survival. With this purpose, fresh commercial semen doses were incubated with different concentrations (0-360 µg/mL) of glyphosate (GLY; exp. 1) or Roundup, at the equivalent GLY concentration (exp. 2), at 38 °C for 3 h. Glyphosate at 360 µg/mL significantly (P < 0.05) decreased sperm motility, viability, mitochondrial activity and acrosome integrity but had no detrimental effect at lower doses. On the other hand, Roundup did significantly (P < 0.05) reduce sperm motility at ≥ 5 µg/mL GLY-equivalent concentration; mitochondrial activity at ≥ 25 µg/mL GLY-equivalent concentration; and sperm viability and acrosome integrity at ≥ 100 µg/mL GLY-equivalent concentration as early as 1 h of incubation. In a similar fashion, GLY and Roundup did not inflict any detrimental effect on sperm DNA integrity. Taken together, these data indicate that, while both glyphosate and Roundup exert a negative impact on male gametes, Roundup is more toxic than its main component, glyphosate.