Improving the quality and in vitro fertilization rate of frozen-thawed semen of buffalo (Bubalus bubalis) bulls with the inclusion of vitamin B12 in the cryopreservation medium

This study was designed to evaluate the effects of vitamin B12 in cryopreservation medium on frozen-thawed semen of buffalo (Bubalus Bubalis) bulls. Semen from five bulls (fertility-proven) were diluted in five aliquots not supplemented (control), or supplemented with 1, 2, 4, or 5 mg/mL of vitamin B12 and evaluated using the Computer Assisted Sperm motion Analysis, antioxidant enzymes, lipid peroxidation (LPO), reactive oxygen species (ROS), ATP concentrations, and in vitro fertilization rate (%). Sperm progressive motility, rapid velocity (%), mitochondrial potential, and acrosome integrity were greater (P < 0.05) with supplementation of 4, and 5 mg/mL vitamin B12 than the control sample. Similarly, compared with the control, samples with 5 mg/mL vitamin B12 supplementation had markedly greater average-path, straight-line, and curved-line velocities (μm/sec). Semen samples supplemented with 2, 4 and 5 mg/mL vitamin B12 had greater concentrations of GPx (U/mL) and SOD (U/mL), whereas LPO (μM/mL) was less (P < 0.05) compared with the control sample. Seminal plasma ROS concentrations (104/25 × 106) were less in the 5 mg/mL vitamin B12 supplemented than control sample. Semen samples supplemented with 5 mg/mL of vitamin B12 had greater concentrations of ATP than control and the 1 mg/mL vitamin B12 supplemented sample. Semen samples supplemented with 5 mg/mL of vitamin B12 had greater plasmalemma and DNA integrities (%) than the control sample. In summary, vitamin B12 supplementation augments semen quality, as evidenced by values for CASA variables, antioxidant enzymes, and ATP concentrations, which may occur as a consequence of inhibition in LPO and ROS production by buffalo spermatozoa.