• Asian-Australasian Journal of Animal Sciences
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• v.24 no.10
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• pp.1348-1357
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• 2011

The objective of this study was to compare the effect of Butylated Hydroxy Toluene (BHT), Pentoxifylline (PTX) and ${\alpha}$-tocopherol (Vit E) on semen quality parameters of Karan Fries bulls. The fortification of extender by various semen additives improves motility as well as fertility of spermatozoa. Split samples of 24 ejaculates of four Karan Fries bulls were extended in extender with or without various additives such as BHT, PTX and Vit E, and performance was evaluated at an interval of 0, 24, 48 and 72 h at refrigerated temperature (4-$7^{\circ}C$). Results of the present study revealed that addition of BHT, PTX and Vit E in extender improved sperm cell function, such as motility, viability, HOST, and acrosome integrity, as compared to the control during liquid storage up to 48 h of preservation at refrigerated temperature. There was no significant (p<0.05) difference between any of the additives up to 48 h of preservation. Overall, the results showed a significant (p<0.05) deterioration in motility after each storage interval. The results showed a significant deterioration in the acrosome integrity and plasma membrane integrity up to 48 h; subsequently, there was not much degradation of both the semen quality parameters. There was a significant increase in spermatozoal tail and total abnormality after each storage interval at refrigerator temperature (4 to $7^{\circ}C$); however, the head and mid-piece abnormalities were almost unaffected. Tail and total abnormality were least in extender fortified with BHT, PTX and Vit E at different hours of incubation as compared to the control. The addition of 1.5 mM BHT, 3.6 mM PTX and 1 mg/ml Vit E in the semen extender has more beneficial effect in terms of semen quality and preservability of spermatozoa.

• Journal of Embryo Transfer
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• v.28 no.4
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• pp.349-353
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• 2013

The objective of this study was to compare the effect of semen extenders on the sperm motility, viability, acrosome integrity and functional integrity of plasma membrane (HOST: hypo-osmotic swelling test) during liquid preservation of Korean Native boar semen. In this experiment, semen was diluted in Androhep plus, Beltsville Thawing Solution (BTS), ModenaTM, Seminark and Vitasem LD. Sperm-rich fractions were collected from three Korean Native boars and sub-samples were diluted ($30{\times}10^6$ spermatozoa/ml) in different semen extenders. Semen samples were stored at $17^{\circ}C$ for 96 hours. On everyday (0, 24, 48, 72, 96 h) after storage, the sperm characteristics relevant for fertility, such as sperm motility, viability, acrosome integrity and HOST positive were evaluated. The motility of spermatozoa stored in different extenders was no significantly different among other extenders (P>0.05). Also, no difference was observed among samples processed with different extenders in the percentage of sperm viability, acrosome integrity and HOST positive. All extenders maintained a high percentage (70%) of sperm motility, viability and acrosome integrity through 96 h of storage. The result of this study show that there was no significant differences among extenders in their capacity to preserve motility, viability and membrane integrity of spermatozoa from normal, fertile Korean Native boars for 96 h of liquid preservation at $17^{\circ}C$.

• Animal Bioscience
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• v.34 no.2
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• pp.192-197
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• 2021

Objective: The present study evaluated the preservation of ram semen at 0℃ using soybean lecithin with a Tris-fructose extender. Methods: Semen was collected by artificial vagina ejaculation from six rams with proven fertility. High quality ejaculates were diluted by soybean lecithin (0.25%, 0.5%, 0.75%, 1.0%, 1.25%) using Tris-fructose extender and control (Tris-fructose egg yolk extender), respectively. The ejaculates were diluted to a concentration of 5×108 sperm/mL, followed by cooling to 0℃ in 90 min and maintaining the temperature for 12 days. The diluted semen samples were examined and recorded for sperm progressive motility, acrosome integrity at 0, 24, 72, 144, 216, 288 h, respectively. Two hundred and twenty-three ewes were inseminated for 216 h with optimal soybean lecithin concentrated semen or control via trans-cervical insemination. Results: The results showed that there were no differences in sperm progressive motility at 0, 24, 72, and 144 h (p>0.05). After 216 h, the sperm progressive motility in the control group and 0.5% concentration groups was significantly higher when compared to 0.25% concentration (p<0.05). The 0.5% concentration group demonstrated the highest survival rate and had no difference with the control group (p>0.05). At 216 h, the sperm progressive motility of all groups was still above 50%. The acrosome integrity of all groups was decreased with prolongation of storage time, but there was no difference at each time point (p>0.05). There was no significant difference in the lambing rate and pregnancy rate between the 0.5% concentration group and the control group (p>0.05). Conclusion: These results suggest that ram sperm is capable of fertilization after preservation at 0℃ with 0.5% of soybean lecithin in Tris-based extender substituted for egg yolk and produce normal offspring after insemination.

• Animal Bioscience
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• v.37 no.6
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• pp.1001-1006
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• 2024

Objective: This study aimed to investigate the effect of Codonopsis pilosula polysaccharide (CPP) on the motility, mitochondrial integrity, acrosome integrity rate, and antioxidant ability of sheep sperm after preservation at 4℃. Methods: Semen from healthy adult rams were collected and divided into four groups with separate addition of 0, 200, 400, and 1,000 mg/L CPP. Sperm motility was analyzed using the Computer-Assisted Semen Analysis software after preservation at 4℃ for 24, 72, 120, and 168 h. Sperm acrosome integrity rate was analyzed by Giemsa staining at 24, 72, and 120 h, and mitochondrial membrane integrity was analyzed by Mito-Tracker Red CMXRos. The total antioxidant capacity (T-AOC) and malondialdehyde (MDA) content of spermatozoa were measured after 120 h of preservation. Results: The sperm viability and forward-moving sperm under 200 mg/L CPP were significantly higher than that in the control group at 72 h (61.28%±3.89% vs 52.83%±0.70%, 51.53%±4.06% vs 42.84%±1.14%), and 168 h (47.21%±0.85% vs 41.43%±0.37%, 38.68%±0.87% vs 31.68%±0.89%). The percentage of fast-moving sperm (15.03%±1.10% vs 11.39%±1.03%) and slow-moving sperm (23.63%±0.76% vs 20.29%±1.11%) in the 200 mg/L group was significantly higher than control group at 168 h. The mitochondrial membrane integrity of the sperm in the group with 200 mg/L CPP was significantly higher than those in the control group after storage at 4℃ for 120 h (74.76%±2.54% vs 65.67%±4.51%, p<0.05). The acrosome integrity rate in the group with 200 mg/L (87.66%±1.26%) and 400 mg/L (84.00%±2.95%) was significantly higher than those in the control group (80.65%±0.16%) after storage for 24 h (p<0.05). CPP also increased T-AOC and decreased the MDA concentration after preservation at 4℃ (p<0.05). Conclusion: Adding CPP could improve the T-AOC of sperm, inhibit lipid peroxidation, and facilitate semen preservation.

• Reproductive and Developmental Biology
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• v.29 no.2
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• pp.133-139
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• 2005

Oxidative stress is one of the major causes of failure in in vitro storage of boar semen. Reactive oxygen species (ROS) are known to be important mediators of such stress. The present study examined the effects of pyruvate and taurine on sperm motility and expression of BAD, Cytochrome c, Caspase-3 and Cox-2 protein in in vitro storage of boar semen, and tested the effect of semen treated with antioxidant with or without hydrogen peroxide on the development of IVM/IVF porcine embryos. Semen samples were transported to the laboratory at $17^{\circ}C$ within 2 hr after collection and were treated with different concentration of pyruvate $(1\~10mM)$ and taurine $(25\~100mM)$ with or without 250uM $H_2O_2$ respectively. The supplementation of pyruvate and taurine increased sperm motility in boar semen during in vitro incubation at $37^{\circ}C$. Expression of apoptosis protein (BAD, cytochrome c, caspase-3 and cox-2) were reduced in the group of boar semen treated with pyruvate and taurine when compared to the other groups. The developmental rates of IVM/IVF porcine embryos fertilized by semen treated with pyruvate and taurine were significantly increased when compared to control (P<0.005). These results indicate that supplementation of pyruvate and taurine as antioxidants in boar semen extender can improve the semen quality and increase in vitro development of porcine IVM/IVF embryos when boar semen treated with antioxidants was used for in vitro fertilization.

• Journal of Embryo Transfer
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• v.33 no.1
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• pp.23-30
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• 2018

The purpose of this study was to investigate the effects of the concentration of seminal plasma in aerobic and anaerobic conditions on the total motility(TM) and the progressive motility(PM) of spermatozoa in long term preservation of cooled equine semen. We also examine the pregnancy rates after artificial insemination using fresh, cooled or frozen semen, and different durations of cooled-preserved equine semen. In the aerobic state of cooled-preserved semen, As the increase of preserved duration to 24h, 48h, 72h, and 96h, TM tended to decrease in each of different concentrations of formalin-containing experimental group, TM tended to decrease regardless of the concentrations of SP. In different concentrations of SP, TM of without seminal plasma(SP W/O) group tended to be higher than that of SP 20%, SP 33% and SP 50%, especially TM of SP W/O group was significantly higher than other groups at 96 h (p<0.05). PM was higher in the groups of SP W/O and SP 20% than in the groups of SP 33% and SP 50% from 24 h to 72 h in cooled-preservation, especially PM of SP W/O group was significantly higher than other groups at 96 h (p<0.05). In the anaerobic condition of cooled-preserved semen, the results of TM and PM at different concentrations of SP were similar to the results in the aerobic condition although there was a difference in the ratio. The pregnancy rates of fresh-cooled, cooled-preserved and frozen semen were 66.3%, 60.7% and 34.5%, respectively, and the pregnancy rate of frozen semen was the lowest. We also found that it is possible to pregnancy after artificial insemination using 72 h cooled-preserved equine semen. There was similar of the pregnancy rates in the different month from April to August.

• Clinical and Experimental Reproductive Medicine
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• v.49 no.3
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• pp.196-201
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• 2022

Objective: This prospective consecutive study investigated the variation in sperm DNA fragmentation (SDF) in multiple semen samples from patients with cancer. Methods: Eighty-one patients with various cancers underwent multiple semen collections on 3 consecutive days for sperm cryopreservation prior to cancer treatment. A commercial Halosperm kit was used to measure SDF. Within- and between-subject coefficients of variation were estimated via random-effects analysis of variance to assess the consistency of semen parameters and SDF. Intraclass correlation coefficients (ICCs) were calculated to assess the magnitude of the between-subject component of variance relative to the total variance. Results: The volume of semen in the day-2 and day-3 samples was significantly lower compared with the day-1 sample. Most parameters showed high ICC values, suggesting that within-subject fluctuations were small relative to the between-subject variability. The highest ICC values were identified for the SDF (ICC, 0.68; 95% confidence interval [CI], 0.45-0.84) and semen volume (ICC, 0.67; 95% CI, 0.45-0.84). Conclusion: Our findings showed that repeated ejaculates from patients with cancer had stable SDF levels.

• Journal of Embryo Transfer
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• v.28 no.1
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• pp.31-39
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• 2013

Alpha-tocopherol as an antioxidant acts in preservation of chilled semen by preserving cell membrane damage from lipid peroxidation. Optimum concentrations of ${\alpha}$-tocopherol in egg yolk-citrate (EYC) extender need to be studied in crossbred bull's semen. Different concentrations of ${\alpha}$-tocopherol viz. 0, 1, 2, 4 and 6mg per ml of extender were used. Semen was collected once a week from four bulls used to regular collection, aged 4 to 7 years, weighing 320 to 450 kg, and with body condition score 4 to 4.5 and scrotal circumference 23 to 32 cm. Semen was evaluated routinely and sperm morphology was viewed under light microscope at ${\times}1,000$ magnification after fixing with buffered formal saline. Over 90% had normal head, acrosome, mid-piece and tail. Semen was diluted with egg-yolk-citrate extender to produce $15{\times}10^6$ spermatozoa/ml and 0, 1, 2, 4 and 6 mg/ml ${\alpha}$-tocopherol were added. The semens amples were kept at $8^{\circ}C$. Sperm motility and viability were examined daily up to 5 days under light microscopy at ${\times}200$ magnification. Sperm viability was acceptable (${\geq}40%$) up to the $4^{th}$ day with all concentrations of ${\alpha}$-tocopherol and up to the $5^{th}$ day with 2 mg/ml ${\alpha}$-tocopherol. Sperm motility was acceptable (${\geq}40%$) up to the $3^{rd}$ day irrespective of ${\alpha}$-tocopherol concentration, and up to the $4^{th}$ day with 2 mg/ml ${\alpha}$-tocopherol. It is suggested that the lifespan of chilled semen may be extended up to 4 days by adding 2mg/ml ${\alpha}$-tocopherol.

• Journal of Embryo Transfer
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• v.27 no.1
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• pp.29-35
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• 2012

The purpose of this study was to examine the effect of growing stages of the Korean Native Striped Bull (KNSB) on the freezability and fertility of frozen-thawed semen. First, we investigated the total motility (TM) and progressive motility (PM) according to the diluent used for semen freezing. Second, we examined the effect of the age of KNSB on semen volume, TM and PM of fresh and frozen-thawed semen. Third, we examined the effect of frozen semen from the different age of KNSB on the $in-vitro$ fertilization rate, and the artificial insemination pregnancy rate. The diluents used in this experiment were Triladyl$^{(R)}$ and Tris-egg yolk extender (EYE). Semen was collected from 5 KNSB in the growing stage (15 months) and 5 adult KNSB (36 months). When Triladyl or Tris-EYE extender was used for semen freezing, there was no difference of the mean TM and the mean PM. However, the mean TM was significantly higher in Bull No. 1885 than Bull No. 4283 ($p$ <0.05). The mean volume of semen collected from the 15-month-old bulls (2.3 ml) was significantly lower ($p$ <0.05) than that from the 36-month-old bulls (5.0 ml). The mean semen concentration was similar for the 15-month-old ($2.1{\times}10^9$ spermatozoa/ml) and 36-month-old ($1.8{\times}10^9$ spermatozoa/ml) bulls. For the 15-month-old and 36-month-old bulls, the mean TM of fresh semen were 93.7% and 88.3%, respectively, and the mean PM were 97.0% and 88.3%, respectively; the 15-month-old bulls showed a particularly high PM ($p$ <0.05). For the 15-month-old and 36-month-old bulls, the mean TM (56.0% and 58.0%, respectively) and the mean PM (64.0% and 70.7%, respectively) of frozen-thawed semen did not differ. The development rates of embryos after $in-vitro$ fertilization and the pregnancy rate after artificial insemination using frozen-thawed semen did not differ according to the bull's age. In summary, semen volume differed according to the bull's age, but semen concentration and survival rate, the $in-vitro$ fertilization rate, and the pregnancy rate did not differ according to the stripe bull's age. Accordingly, semen from bulls in the growing stage can be collected and frozen for the preservation and multiplication of rare livestock.

• Korean Journal of Animal Reproduction
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• v.20 no.2
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• pp.101-109
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• 1996

This study was done to find out the methods of long-term use of liquid boar semen in 100 ml plastic bottle for artificial insemination and to investigate differences between Lactose-Egg yolk and Biitschwiler diluents according to storage temperature, and effect of final glycerol concent ration in Lactose-Egg yolk diluent. Liquid boar semen diluted with Lactose-Egg yolk diluent showed significantly higher sperm motility (p<0.05) after 0.5 and 2h incubation at 37$^{\circ}C$,than Butschwiler diluent at all storage length when it was preserved in the 5$^{\circ}C$ refrigerator. The NAR acrosome in Lactose-Egg yolk diluent a after 0.5 and 2h incubation at 37$^{\circ}C$, respectively, during preservation periods was similar to that in Biitschwiler diluent. When liquid boar semen was preserved at 15$^{\circ}C$, liquid boar semen in the Butschwiler diluent showed significantly higher percentages of sperm motility and NAR acrosome from third day to seventh than that in Lactose-Egg yolk diluent. In the effect of final glycerol concentration of liquid boar semen in the Lactose-Egg yolk diluent, the final glycerol concentration of 2% showed higer percentages of sperm motility and NAR acrosome than that of 0, 1, 3, and 5%. Farrowing rate, litter size and average pig weight at birth did not differ significantly between Lactose-Egg yolk and But schwiler diluents. As a result of this study, we found out that liquid boar semen can be stored for 6-7 days at 5$^{\circ}C$ in Lactose-Egg yolk diluent and at 15$^{\circ}C$ in Butschwiler diluent.