Recently, male impact has been recognized as a momentous cause of infertility. Male infertility isn’t an entity but it evokes a variety of mechanisms and explanations to understand it. In between, a biochemical semen additive that improves or overcome certain semen situations so can enhance or treat semen fertility and infertility, respectively. L-carnitine is one of these biochemical semen additives. Till now, the exact effects of L-carnitine on buffalo semen processing outcomes haven’t been discovered. The current study aimed to clarify L-carnitine roles during buffalo semen cryopreservation. Semen samples were obtained from six fertile buffalo bulls (aged 3 to 5 years). Weekly, two consecutive ejaculates were collected from each bull for successive six weeks using artificial vagina. The ejaculates were pooled to eliminate samples variability. Semen samples were extended with Tris-based extender supplemented with different concentrations of L-carnitine (0.01, 0.05 and 0.1 mg/ml) Vs. Tris-based extender only (control). Then they were processed to cryopreservation and thawing to assess different semen characteristics. It had been found that L-carnitine (0.05mg/ml) significantly (p