Sodium dodecyl sulphate, N-octyl β-D glucopyranoside and 4-methoxy phenyl β-D glucopyranoside effect on post-thaw sperm motion and viability traits of Murrah buffalo (Bubalus bubalis) bulls

Abstract

Sodium Dodecyl Sulphate (SDS), N-Octyl β-D Glucopyranoside (NOG), 4-Methoxy Phenyl β-D Glucopyranoside (4-MPG) as ice recrystallization inhibitors were added to Tris Egg Yolk Glycerol (TEYG) semen extender for cryopreservation of semen of buffalo bulls. Post-thaw sperm motion and viability traits were evaluated. Pilot study involved six semen ejaculates (2 ejaculates/bull, from three bulls); second experiment was conducted using twenty seven semen ejaculates (9 ejaculates/bull, from 3 bulls) and in third experiment three semen ejaculates (one bull) were used. Eight concentrations of SDS (2, 1, 0.5, 0.25, 0.15, 0.125, 0.0625 and 0.0312%), twelve concentrations of NOG (33, 22, 11, 5.5, 2.5, 0.75, 0.5, 0.25, 0.125, 0.0625, 0.03125 and 0.0156 mM), and, eleven concentrations of 4-MPG (220, 165, 110, 55, 50, 25, 12.5, 6.25, 3.125, 1.56 and 0.78 mM) were supplemented in TEYG semen extender to evaluate the post-thaw sperm motility and viability traits. Computer Assisted Sperm Analysis (CASA) was used to measure the kinetic and functional parameters for sperm motion traits, Hypo Osmotic Swelling Test (HOST) for sperm plasma membrane integrity, Eosin Nigrosin staining for viability and Rose Bengal staining for sperm abnormalities for all the experiments except for pilot study where only Total Motility (TM) and Rapid Progressive Motility (RP) were evaluated. Three freezing protocols; i) Normal P24 (freezing rate of -30 °C min^-1 from 4 °C to -15 °C; -40 °C min^-1 from -15 °C to -60 °C; and -50 °C min^-1 from -60 °C to -140 °C; and then plunged in liquid Nitrogen at -196 °C); ii) Moderate P25 (freezing rate of -30 °C min^-1 from 4 °C to -15 °C; -50 °C min^-1 from -15 °C to -60 °C; and -50 °C min^-1 from -60 °C to -140 °C; and then plunged in liquid Nitrogen at -196 °C); and iii) Rapid P26 (freezing rate of -30 °C min^-1 from 4 °C to -15 °C; -60 °C min^-1 from -15 °C to -60 °C; and -50 °C min^-1 from -60 °C to -140 °C; and then plunged in liquid Nitrogen at -196 °C) were evaluated using SDS 0.125% in TEYG semen extender. SDS ≤0.125%, NOG ≤0.0625 mM and 4-MPG ≤ 3.125 mM in TEYG buffalo semen extender improved significantly (p < .05) the kinetic and functional parameters as compared to the other Ice Recrystallization Inhibitors (IRIs) concentrations used for cryopreservation of buffalo bull semen in the pilot study. SDS 0.125% supplementation was the best IRI among all which resulted in improved kinetic and functional parameters of bull semen in second experiment. Conclusion was drawn that buffalo bull semen cryopreservation using sodium dodecyl sulphate, 0.125% as IRI in TEYG semen extender along with freezing protocol P 25 revealed optimum kinetic and functional parameters for post-thaw spermatozoa.

Keywords: Cryopreservation; Ice re-crystallization; Murrah buffalo; Plasma membrane integrity; Sodium dodecyl sulphate.