Arachidic Acid in Extender Improves Post‐thaw Parameters of Cryopreserved Nili‐Ravi Buffalo Bull Semen

Cryopreservation process reduces lipids and phospholipids from buffalo bull spermatozoa. It was therefore hypothesized that supplementation of fatty acid to extender may improve the post‐thaw quality of buffalo semen. The objective was to evaluate the effect of arachidic acid supplementation in extender on post‐thaw quality of buffalo bull (Bubalus bubalis) spermatozoa. Semen was collected from three adult Nili‐Ravi buffalo bulls of similar age group with artificial vagina (42°C) for 3 weeks (replicate). Qualified semen ejaculates (n = 18) were split into four aliquots and diluted in tris–citric acid extender containing 0.0 (control), 5.0, 10.0 and 20.0 ng/ml at 37°C having approximately 50 × 10⁶spermatozoa/ml. Diluted semen was cooled to 4°C in 2 h and equilibrated for 4 h at 4°C. Cooled semen was filled in 0.5‐ml straws at 4°C, kept on liquid nitrogen vapours for 10 min and plunged in liquid nitrogen for storage. Thawing of frozen semen was performed after 24 h at 37°C for 30 s. Sperm progressive motility (%) was improved in a dose‐dependent manner by supplementing arachidic acid at 5.0, 10.0 and 20.0 ng/ml compared with control. Structural and functional integrity of sperm plasma membrane (%), number of acrosome‐intact live sperm (%) and sperm chromatin integrity (%) were better (p < 0.05) in extender having 5.0 ng/ml of arachidic acid compared with control. At 10.0 ng/ml, these values did not vary (p > 0.05) from those at 5.0 ng/ml. Further improvement in structural and functional integrity of sperm plasma membrane, number of acrosome‐intact live sperm and chromatin integrity was observed at 20.0 ng/ml of arachidic acid in extender. In conclusion, arachidic acid supplementation in extender improved the post‐thaw quality parameters of cryopreserved Nili‐Ravi buffalo bull spermatozoa. Among the arachidic acid concentrations studied, maximum improvement in post‐thaw semen quality parameters was observed at 20.0 ng/ml.