Cryopreservation of Swamp Buffalo Semen in Skim Milk Yolk-based Diluent with Two Different Cryoprotectants

Abstract

The successful of artificial insemination (AI) in buffalo is still low. The quality of frozen semen determines the successful of AI program. Glycerol is widely used for cryopreservation of buffalo semen with unsatisfactory results due to its toxicity and antifertility properties. This study aimed to investigate the effect of glycerol and dimethylformamide (DMF) concentrations in skim milk egg yolk (SMEY)-based diluent on the quality of frozen-thawed swamp buffalo bull semen. Fresh semen was divided into eight aliquots. Each aliquot was diluted in SMEY containing 4%, 5%, 6%, and 7% glycerol and 4%, 5%, 6%, and 7% DMF. Diluted semen was packed into mini straw (0.25 mL), equilibrated at 4 °C for 4 hours, frozen in liquid nitrogen vapor for 10 minutes, and stored at liquid nitrogen container. The straws were thawed after 24 hours at 37 °C for 30 seconds for evaluation. The results showed that the post thawed sperm motility and recovery rate in SMEY extender containing 7% glycerol and 5% DMF were higher than the other glycerol or DMF concentrations. Post thawed sperm viability and plasma membrane integrity in SMEY containing 4%-6% DMF did not differ significantly, however both values were higher than those in 7% DMF. Post thawed sperm viability and plasma membrane integrity did not differ between SMEY containing 6% and 7% glycerol. However, both values were higher than those in 4% and 5% glycerol. It is concluded that 7% glycerol or 5% DMF is considered to be the most suitable cryoprotective agent for swamp buffalo semen cryopreservation.

The Quality of Buffalo Sperm Following Preservation Using Different Diluents and Sperm Concentrations

“…The viability of buffalo sperm in this study (35.33 -66.67%) was different from reports Herbowo et al (2019) which used tris egg-yolk diluent (49.08-61.52%) and reports Ansari et al (2017) which used andromed® and tris egg-yolk diluents (61.5-67.5%). The difference in viability was caused by different types of buffalo, type of diluent and feed (Manjunath, 2012).…”

“…However, the abnormality value in this study was almost the same as the abnormality of buffalo sperm in tris-egg yolk diluent at different glycerol concentrations (7.80-16.30%) (El-Sisy et al, 2016). The percentage of PMI of buffalo sperm in this study was higher than PMI of buffalo sperm in Pakistan (44.4-46.8%) that used andromed® and tris egg-yolk diluent (Ansari et al, 2017) but not different from PMI of buffalo sperm in Ciawi that used tris egg-yolk diluent (51.38-62.41%) (Herbowo et al, 2019).…”

“…The tris diluent were homogenized for 15 minutes, boiled and cooled to a temperature of 37 0 C before mixed with 20mL of egg yolks. This tris-egg yolk diluent were then homogenized for 60 minutes, added by 6% glycerol, 1000IU/mL benzylpenicillin and 1000mg/mL streptomycin sulfate, homogenized and cooled at a temperature of 37 0 C (Herbowo et al, 2019). All chemicals purchased from Sigma-Aldrich Co, USA.…”

Supplementation of Pandanus conoideus Oil in Cryopreservation Diluents for Maintaining the Semen Quality of Ongole Grade Bull

“…Before the freezing process, the semen should be equilibrated to adapt the sperm to the freezing conditions. Equilibration of sperm has been carried out with various times and temperatures, for example, with temperatures of 4 o C for 6 hours (Herbowo et al, 2019) and 5 o C for 22 hours (Dwinofanto et al, 2018). The metabo-lism of sperm can be suppressed if the semen is cooled at 0 o C-5 o C. Stress conditions of sperm can interfere with the process of inhibition of phosphorylation and phospholipid oxidation, which results in the increased production of reactive oxygen species (ROS) in the semen, and excessive levels of ROS in the cells can increase the oxidation of lipids, proteins, and DNA.…”

Storage Period of Liquid Semen Eligible for Insemination in Thin Tail Sheep Semen Diluted with Egg Yolk Citrate with the Addition of Noni (Morinda citrifolia Linn) Fruit Extract

“…This demonstrated that the addition of Noni fruit extract in egg yolk citrate diluent might significantly preserve the standards of thin tail ram sperm during storage at 5°C for 6 days, even though the best results were obtained at D-5. The result also showed that the infused of noni extract in P1 (4%) and P2(6%) to the sperm diluent could maintain the sperm post thawing motility (PTM) in 40% which still suitable for insemination [11]. The high percentage of sperm motility in P1 and P2 due to the high vitamin C content of noni fruit [12] which was 29.29 mg/ml [13].…”