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The use of artificial insemination with cryopreserved semen in cattle greatly advances animal breeding and production, reducing its costs and facilitating the distribution of semen (3). A major cause of the poor quality of buffalo bull semen and the low fertility of frozenthawed semen is cryocapacitation-inflicted damage to sperm during freezing and thawing (2). Spermatozoa are significantly affected by interactions of extracellular and intracellular fluid, sugars, proteins, salts of individual microelements, and other substances, whose concentration increases simultaneously with the osmotic pressure after ice crystal formation (33). Cold shock caused by the action of low temperatures on spermatozoa during the freezing process may damage mitochondria (28), plasma, and the acrosomal membrane of spermatozoa (24), inducing changes in the lipid composition or in the integrity and permeability of plasma and membranes (20). Vitamin B12 (cobalamin) is a large, complex molecule responsible for a range of functions within the body. In biochemical reactions, vitamin B12 acts as a coenzyme (22). Moreover, it is a form usually used in vitamin supplements because of its stability. It has been demonstrated in experiments with rats that atrophic changes and the arrest of spermatogenesis resulted from a vitamin B12 deficiency. Watanabe et al. (32) observed that a deficiency of vitamin B12 resulted in a decreased motility and velocity of sperm in male rats, as well as in an increased incidence of abnormal sperm. Research by Boxmeer et al. (5) showed that there was a positive correlation between the total cyanocobalamin concentration in seminal plasma and the spermatozoa concentration in semen. Cyanocobalamin is also known for its antioxidative potential, because, as a supplement, it decreases the amount of reactive oxygen species (ROS) produced during oxidative stress in human semen (8, 9). It has been suggested that vitamins B1, B6, and B12 in rams play a key role in the thermoregulation of scrotal skin and rectal temperature and in maintaining libido, semen quality, and fertility during heat stress (10). DNA sequences are involved in regulating the expression of genetic information. Sperm DNA integrity is important for the success of natural and assisted Influence of addition of cobalamin to the extender on the post-thaw motility, viability, and DNA integrity of bovine ejaculate