70 REFREEZING POST-THAWED GOAT SEMEN

2011 ◽  
Vol 23 (1) ◽  
pp. 140
Author(s):  
D. B. Carwell ◽  
B. R. Scott ◽  
G. T. Gentry ◽  
K. R. Bondioli ◽  
R. A. Godke
Keyword(s):  
Liquid Nitrogen ◽  
Sperm Morphology ◽  
Membrane Integrity ◽  
Egg Yolk ◽  
Slow Cool ◽  
Semen Parameters ◽  
Slow Cooling ◽  
Freeze Thaw ◽  
Thaw Cycle ◽  
Frozen Semen

The ability to successfully refreeze caprine sperm could provide a means of salvaging semen that was mistakenly thawed. The objective of this study was to compare treatment post-thaw semen parameters of twice-frozen caprine semen. Frozen semen from six mature Boer bucks (range in age from 2 to 6 years) was utilised for this experiment. Semen from each buck was extended in an egg yolk-based extender and packaged in 0.5-mL plastic straws before freezing and stored in liquid nitrogen. Three units of frozen semen from each buck was randomly allotted to each of four treatments as follows: (A) thaw and evaluate (control), (B) thaw, then plunge into liquid nitrogen, thaw, and evaluate, (C) thaw, incubate for 3 min at 37°C, slow cool and freeze, thaw, and evaluate, and (D) thaw, incubate for 5 min at 37°C, slow cool and freeze, thaw, and evaluate. Post-thaw parameters included total motility (TM), progressive motility (PM), membrane integrity (MI), and sperm abnormalities (AB). To obtain MI and AB, samples were stained with an eosin-nigrosin stain. A computerized programmable freezer was used to refreeze semen samples in treatment (Trt) C and Trt D. During the slow cooling portion of the protocol, samples were allowed to equilibrate at 38°C, then cooled to 4°C at a rate of 0.30°C min–1, and then held for 5 min. Samples were then cooled to –8°C at a rate of 15°C min–1, seeded, and cooled to –10°C at 15°C min–1, samples were then ramped to –80°C at 30°C min–1 before plunging into liquid nitrogen. Results indicate that post-thaw TM was significantly greater for Trt A (60%) when compared with Trt B, C, and D (0.05, 35, and 39%, respectively). Mean TM were not different between Trt C (35%) and Trt D (39%) but were greater than that for Trt B (0.05%). The PM for post-thaw semen in Trt A was also significantly greater (P < 0.05) when compared with that for Trt B and C (0.05 and 25%); however, no difference was found for mean PM for Trt A (47%) and Trt D (30%), nor were differences found between Trt C (25%) or Trt D (30%). Membrane integrity was higher in Trt A (27%) when compared to Trt B (2.2%). No differences in membrane integrity where found between Trt A, C, and D (27, 13, and 14%, respectively). Additionally, no differences were found between Trt B, C, and D for membrane integrity. Sperm morphology were not different were found with across all treatment groups. These results (i.e. Trt C and D) indicate that semen from mature Boer bucks can undergo a second freeze thaw cycle and still retain motility without dramatically affecting sperm morphology and membrane integrity. These findings indicate that directly plunging recently thawed semen back into liquid nitrogen should not be used for artificial insemination.

Preservation of Black Bengal buck semen in soybean lecithin based chemically defined extender

2018 ◽  
Author(s):  
Pangdun Konyak ◽  
Ajoy Mandal ◽  
Mohan . ◽  
C. Bhakat ◽  
S. K. Das ◽  
...  
Keyword(s):  
Liquid Nitrogen ◽  
Soybean Lecithin ◽  
Membrane Integrity ◽  
Egg Yolk ◽  
Semen Parameters ◽  
Control Group ◽  
Initial Dilution ◽  
Nitrogen Vapor ◽  
Sperm Characters

This experiment was conducted with the aim to study the effect of replacing egg yolk with soybean lecithin (SL) for cryopreservation of Black Bengal buck semen. Sexually matured Black Bengal buck (n = 5) were used and the ejaculates were obtained using an artificial vagina method. The semen samples were pooled and diluted in Tris extender with 5% Glycerol containing either 15% egg yolk (control group) or SL at different concentrations (1% SL, 1.5% SL and 2% SL). The semen samples were filled in straws and cooled gradually to 5 °C. Semen straws were equilibrated for 3 hours at 5°C and were frozen in static liquid nitrogen vapor and stored in liquid nitrogen. Semen samples were evaluated after initial dilution, after completion of equilibration and after freeze thawing for in vitro sperm characters such as sperm motility, functional membrane integrity and malondioldehyde (MDA) concentration. Semen samples preserved in extender containing 1% SL was able to maintain in vitro sperm characters similar to the extender containing egg yolk. However, significant (P less than 0.05) reduction in all semen parameters was observed as the concentration of soybean lecithin increased above 1% level. It is concluded that an extender containing soybean lecithin @ 1% with 5% Glycerol can be used for replacing egg yolk for cryopreservation of Black Bengal buck semen.

scholarly journals Fertilitas Semen Beku dalam Tris Kuning Telur dan Skim yang Diberi Omega-3 pada Sapi Simmental dengan Ransum Berimbuhan Seng dan Selenium Minimal

2018 ◽  
Vol 19 (2) ◽  
pp. 251 ◽  
Author(s):  
Asep Kurnia ◽  
Soeparna Soeparna ◽  
Raden Iis Arifiantini ◽  
Rahmat Hidayat
Keyword(s):  
Plasma Membrane ◽  
Liquid Nitrogen ◽  
Membrane Integrity ◽  
Egg Yolk ◽  
The Other ◽  
Omega 3 ◽  
Nitrogen Vapor ◽  
Frozen Semen ◽  
Sperm Plasma Membrane ◽  
The Individual

This study aims to examine the quality of frozen semen in Tris egg yolk (TEY) extender and skimmed milk extender with or witout omega-3. A total of 18 Simmental bulls belong to Lembang Artificial Insemination Centre were divided into three groups. Each was fed with standard feed (R1), standard feeds supplemented with minimal Se and Zn (R2) and standard feed supplemented with maximal of Se and Zn concentration. Semen were collected using an artificial vagina and were evaluated macro- and microscopically. The semen then were divided into four tubes and each diluted with Skimmed, SkimmedOmega 3, TEY or TEY-O. The semen was then packed into a 0.25 ml straw and equilibrated at 5 oC for 4 hours, then frozen above liquid nitrogen vapor, and stored in liquid nitrogen container (-196 oC). The qualities of frozen semen were evaluated on the motility, individual score, viability and integrity of the plasma membrane of sperms. Sperm motility of bulls fed with standard feed (R1) in TEY extender and R3 in TEY and TEY-Omega-3 extender were higher (p <0.05) compared to the other combinations. No difference was found on the individual score. The viability of sperms in bulls fed with standard feed in SkimmedOmega-3 extender was higher than the other treatments and the highest sperm plasma membrane integrity was demonstrated by sperm in bull feeding with R2 in TEY extender.

scholarly journals Effect of the addition of sodium caseinate on the viability of cryopreserved buffalo semen

2020 ◽  
pp. 2209-2218
Author(s):  
Fernando Evaristo da Silva ◽  
Jaqueline Candido Carvalho ◽  
Camila de Paula Freitas Dell'Aqua ◽  
Frederico Ozanam Papa ◽  
Marc Roger Jean Marie Henry ◽  
...  
Keyword(s):  
Cell Damage ◽  
Membrane Integrity ◽  
Sodium Caseinate ◽  
Egg Yolk ◽  
Freezing Process ◽  
Computer Assisted ◽  
Semen Cryopreservation ◽  
Sperm Analysis ◽  
Frozen Semen ◽  
Buffalo Semen

The use of cooled semen in artificial insemination operations results in higher pregnancy rates than the use of frozen semen. This result seems to be related to the more severe damage triggered by the freezing process than that observed during refrigeration. Due to its ability to bind to sperm-binding proteins and calcium ions, sodium caseinate has been studied as a substance capable of preventing early sperm capacitation, a significant cause of the decreased pregnancy rate resulting from the use of frozen semen. The first objective of this study was to evaluate whether a commercial egg yolk diluent developed for frozen bovine semen could be used for buffalo semen cryopreservation; the second objective was to investigate the effect of this diluent in combination with sodium caseinate during the procedures of buffalo sperm cryopreservation using flow cytometry and computer-assisted sperm analysis. In the first part of the study, comparing the results of spermatic kinetics and plasma and acrosomal membrane integrity, it was observed that the freezing process resulted in more cell damage than the cooling process. In the second part of the study, no effects of the addition of sodium caseinate to the egg yolk diluent were observed. From the results of the present study, it was possible to conclude that the egg yolk-based diluent was suitable for buffalo semen cryopreservation and that the addition of sodium caseinate did not decrease the harmful effects related to seminal cryopreservation.

scholarly journals Effect of sunflower lecithin on Kalahari Red goat semen during cryopreservation

2018 ◽  
Vol 51 (1) ◽  
pp. 21-28 ◽  
Author(s):  
Sakirat Opeyemi Adeyanju ◽  
James Olatinbo Daramola ◽  
Jimoh Alao Olanite ◽  
Olufiropo Samson Awokola
Keyword(s):  
Soybean Lecithin ◽  
Membrane Integrity ◽  
Egg Yolk ◽  
Domestic Animal ◽  
Arginase Activity ◽  
Semen Parameters ◽  
Phospholipid Content ◽  
Acrosome Integrity ◽  
Semen Extender ◽  
Different Levels

Abstract Soybean lecithin had been used as an alternative to egg yolk in domestic animal semen extender during cryopreservation due to its characteristic phospholipid content which played a major cryoprotective role. This composition of soybean lecithin informed the replacement of soybean with sunflower lecithin (SL) in the extender for the Kalahari Red (KR) buck semen cryopreservation in this study. Effect of different levels of SL on the quality of the KR buck semen during cryopreservation using slow freezing method was evaluated. Semen samples were collected from four KR bucks of between two and two and half of age using artificial vagina, evaluated for motility and then diluted in extenders containing different levels of SL (1.5%, 3.0% and 4.5%) as experimental group and 0% SL or 20% egg yolk as control. Semen parameters including motility, acrosome integrity (AcI), membrane integrity (MI), malondialdehyde (MDA) concentration, cholesterol level and seminal arginase activity were evaluated for. The results showed that motility, acrosome integrity (AI) and membrane integrity were comparable at 0%, (22.00 ± 4.58, 82.00 ± 3.51 and 96.00 ± 2.03); 1.5%, (23.00 ± 2.08, 87.00 ± 3.79 and 89.00 ± 2.08); 3.0%, (13.00 ± 2.52, 81.33 ± 0.41 and 76.67 ± 1.20) and 4.5% (11.00 ± 4.51, 85.33 ± 9.88 and 84.00 ± 8.50), respectively, after thawing. SL at 0% had the highest (P < 0.05) values for MDA, cholesterol and seminal arginase activity (1.10 ± 0.008 nmol/ml, 236.35 ± 4.08 mg/dl and 0.54 ± 3.3 E-3 units/mg protein, respectively). Our data suggest that 1.5% sunflower lecithin can be used in place of soy lecithin as a substitute for egg yolk during the cryopreservation of caprine semen.

46 Effect of Bioxcell® and Triladyl® Extenders on Washed Semen of South African Indigenous Bucks

2018 ◽  
Vol 30 (1) ◽  
pp. 162
Author(s):  
L. P. Nethenzheni ◽  
M. L. Mphaphathi ◽  
N. C. Negota ◽  
T. L. Nedambale
Keyword(s):  
Treatment Group ◽  
South African ◽  
Liquid Nitrogen ◽  
Seminal Plasma ◽  
Semen Parameters ◽  
Mean Values ◽  
Treatment Groups ◽  
Frozen Semen ◽  
The Mean

Semen extenders and seminal plasma are vital for cryopreservation of buck semen. The objectives of the study were to evaluate the effect of 2 extenders: Triladyl® (Minitube, Tiefenbach, Bavaria) and Bioxcell® (IMV, L’Aigle, France) and the removal of seminal plasma on buck semen. Six indigenous bucks were used in this study and 6 ejaculates were collected from individual bucks. The semen was pooled and then randomly allocated into 6 groups: (1) raw-washed, (2) raw-non-washed, (3) Triladyl®-washed, 4) Triladyl®-non-washed, (5) Bioxcell®-washed, and (6) Bioxcell®-non-washed. Spermatozoa viability was assessed using Eosin-Nigrosin and morphology using Spermac® (Vitrolife, Göteborg, Sweden) stains. The washed semen samples were all diluted into (1:4 v/v) with PBS and centrifuged at 1500 × g for 10 min. Semen samples were then extended with Triladyl® or Bioxcell® per treatment groups and equilibrated for 2 h at 5°C. The semen samples were loaded into straws per treatment groups and placed 5 cm above a liquid nitrogen vapour for 10 min and then stored at –196°C until use. After 1 month of storage, frozen semen straws per treatment group were thawed at 37°C for 30 s, and spermatozoa parameters were analysed post-thaw. Significant differences among the mean values of semen parameters were determined by Tukey’s test using ANOVA, GLM procedure of SAS version 12.1 of 2010 (SAS Institute Inc., Cary, NC, USA). There was a higher (P < 0.05) live and normal spermatozoa percentage in non-washed semen extended with Bioxcell® (45.7 ± 21.2) than the semen extended with Triladyl® (24.5 ± 22.2%). Live and normal spermatozoa percentages were drastically reduced in the Bioxcell® (5.2 ± 4.9) and Triladyl® (6.9 ± 8.6%) washed semen groups. There was a higher (P < 0.05) percentage of spermatozoa with head abnormalities in non-washed semen extended with Triladyl® (20.4 ± 10.2), compared with the semen extended with Bioxcell® (18.3 ± 12.4%) following freeze-thawing. There was a higher (P < 0.05) percentage of spermatozoa with head abnormalities in washed semen samples extended with Triladyl® (34.0 ± 16.0) compared with the semen extended with Bioxcell® (10.1 ± 7.0%). There were higher (P < 0.05) percentages of spermatozoa with coiled tail abnormalities in washed semen extended with Bioxcell® (65.4 ± 25.0) compared with Triladyl® (35.9 ± 21.6%). In conclusion, the liveability of spermatozoa was negatively affected by washing of semen extended with Bioxcell® and Triladyl® extender. Bioxcell® significantly increased tail abnormalities and Triladyl® gave less protection against head abnormalities following cryopreservation of South African unimproved indigenous bucks’ semen.

21 Effect of egg yolk extracted low-density lipoprotein on cryopreserved Nguni bull semen

2019 ◽  
Vol 31 (1) ◽  
pp. 136
Author(s):  
M. M. Tshabalala ◽  
K. A. Nephawe ◽  
M. L. Mphaphathi ◽  
C. M. Pilane ◽  
T. L. Nedambale
Keyword(s):  
Plasma Membrane ◽  
Sperm Motility ◽  
Sperm Quality ◽  
Egg Yolk ◽  
Semen Parameters ◽  
Dna Integrity ◽  
Low Density ◽  
Bull Semen ◽  
Frozen Semen ◽  
Live Sperm

Egg yolk has been reported to have a beneficial effect on sperm quality following cryopreservation, and this led to its widespread use in semen extenders. However, egg yolk contains substances that inhibit respiration of sperm cells and diminish their motility rate. Moreover, it also contains low-density lipoproteins (LDL) that have a protective effect on sperm during the cryopreservation process. The objective of this study was improve cryopreservation of Nguni bull semen using egg yolk low-density protein. A total of 25 ejaculates were collected from 5 Nguni bulls aged 4 to 5 years using an electroejaculator during the natural breeding season. Collected raw semen samples were transported to the laboratory and evaluated for sperm motility, viability, plasma membrane, acrosome, and DNA integrity before dilution. Semen was randomly diluted with a sodium citrate extender supplemented with 20% egg yolk (control) and with 6, 8, 10, and 12% LDL concentrations. The diluted semen sample groups were equilibrated for 4h at 5°C. Following equilibration, semen was loaded into 0.25-mL straws and frozen in a controlled-rate programmable freezer. The groups of semen straws were then plunged into LN and transferred into LN tanks (−196°C) for storage. The frozen semen straws per treatment group were thawed at 37°C and evaluated for sperm motility, viability, plasma membrane, acrosome, and DNA integrity. Data were analysed with ANOVA using Stata V12 statistical software (StataCorp., College Station, TX), and treatment means were separated using Fisher’s protected t-test at the significant level of P&lt;0.05. Sperm motility and membrane integrity were significantly higher (P&lt;0.05) in frozen-thawed semen diluted with 8% LDL compared with the other concentrations. However, 6 and 8% LDL resulted in a significantly higher (P&lt;0.05) live sperm, DNA, and acrosome integrity. Frozen-thawed semen diluted with 10 and 12% LDL resulted in the lowest percentages of sperm motility, live sperm, plasma membrane, acrosome, and DNA integrity following cryopreservation. In conclusion, extender containing 8% LDL resulted in improved Nguni bull semen parameters such as sperm motility, viability, plasma membrane, acrosome, and DNA integrity following cryopreservation. Further studies are required to determine the fertilizing capacity of semen diluted and frozen with LDL in vitro and in vivo.

Effect of Bioxcell® and Triladyl® Extenders and Removal of Seminal Plasma on Equilibrated and Cryopreserved Semen from South African Unimproved Indigenous Bucks

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Nethenzheni LP ◽  
◽  
Mphaphathi ML ◽  
Madzhie LR ◽  
Negota NC ◽  
...  
Keyword(s):  
Seminal Plasma ◽  
Membrane Integrity ◽  
The Other ◽  
Semen Parameters ◽  
Lower Percentage ◽  
Mean Values ◽  
Sperm Analysis ◽  
Progressive Motility ◽  
Frozen Semen ◽  
Motility Rate

The objectives of the study were to evaluate the effect of two extenders (Triladyl® and Bioxcell®) and the removal of seminal plasma on indigenous buck’s semen. Semen was collected from six indigenous bucks using an electro-ejaculator. Raw semen was pooled and randomly allocated into six groups as follows: (i) Raw non-washed, (ii) Raw washed, (iii) Triladyl®-washed, (iv) Triladyl®-non-washed, (v) Bioxcell®-washed and (vi) Bioxcell®-non-washed. Both the Triladyl® and Bioxcell® washed semen samples groups were diluted (1:4 v/v) with Phosphate Buffered Saline (PBS) then centrifuged at 1500x g for ten min and seminal plasma was removed. The groups were analysed for spermatozoa motility rates using Computer-Aided Sperm Analysis (CASA). The spermatozoa viability was assessed using Eosin-Nigrosin, acrosome integrity using Spermac, chromatin structure using Acridine Orange, and mitochondria using JC-1 staining solutions. Semen samples were diluted (1:4 v/v) as follows: Triladyl® (washed and non-washed) or Bioxcell® (washed and non-washed) and then equilibrated at 5°C for 2 hours. Equilibrated semen samples in 0.25 mL French straws were placed 5 cm above a Liquid Nitrogen (LN2) vapour for 10 min, and stored for one month. Frozen semen straws per treatment group were thawed at 37°C for 30 seconds. Significant differences among the mean values of semen parameters were determined by Tukey’s test using ANOVA, GLM procedure of SAS version 12.1 of 2010. The spermatozoa progressive motility rate in non-washed semen extended with Bioxcell® was significantly higher (89.6±7.5a) compared with that of non-washed Triladyl®, washed Bioxcell® and Triladyl® (P<0.05). Live spermatozoa percentage in washed semen extended with Triladyl® extender was reduced (27.7±17.1) significantly compared with the other groups (P<0.05). There was a lower percentage of spermatozoa with high mitochondrial membrane potential in non-washed and washed semen extended with Bioxcell® (39.5±23.2 and 37.9±28.6, respectively) compared with that of non-washed and washed semen extended with Triladyl® (P>0.05). The spermatozoa progressive motility rate in non-washed semen extended with Bioxcell® (58.5±10.0) extender was significantly higher compared with that of the other groups (P<0.05). There was a higher live and normal spermatozoa percentage in non-washed semen extended with Bioxcell® (45.7±21.2) compared with that of the other groups (P<0.05). In conclusion, Washing of seminal plasma in semen extended with Triladyl® was not essential, as it lowered viability, progressive motility and chromatin membrane integrity prior and post-cryopreservation. However, Bioxcell® extender was found to be more suitable for preserving spermatozoa during equilibration and freezing/thawing process of non-washed and washed buck semen.

scholarly journals The effect of freezing on cryosurvival of yak sperm

2018 ◽  
Vol 15 (2) ◽  
pp. 215-218
Author(s):  
S Deori
Keyword(s):  
Liquid Nitrogen ◽  
Sperm Motility ◽  
Artificial Insemination ◽  
Sperm Concentration ◽  
Egg Yolk ◽  
Mean Values ◽  
Frozen Semen ◽  
Sperm Abnormality ◽  
Semen Characteristics ◽  
Live Sperm

A study was carried out to study the effect of freezing on cryosurvival of yak semen. Artificial insemination in yak is still in infancy. Semen cryopreservation and use of artificial insemination can be applied in yak husbandry for conservation and rapid multiplication of superior germplasm. Semen was collected from four adult yak bulls using artificial vagina method managed under uniform conditions. A total of 40 ejaculates comprising of 10 ejaculates each bull were collected following twice a week schedule and evaluated for fresh semen characteristics. The fresh yak semen characteristics viz. ejaculate volume (ml), mass activity (0-4), initial sperm motility (%), sperm concentration (x 106/ml), live sperm (%), sperm abnormality (%) and intact acrosome (%) were 3.10 ± 0.18, 3.53 ± 0.96, 83.89 ± 2.87, 1180.22 ± 42.32, 77.63 ± 4.23, 8.45 ± 3.33 and 93.61 ± 3.78 respectively. The ejaculates were diluted (1:10) with Tris extender consisting of 6.4 ml glycerol and 20 ml of fresh egg yolk. Straws were equilibrated at 5°C for 4 hours followed by exposure to liquid nitrogen vapour for 10 minutes and finally transferred to liquid nitrogen container for storage. The cryosurvival rate was studied after 7 days of storage in liquid nitrogen. The frozen semen was thawed in warm water (37°C) for 30 seconds for evaluation. Mean values of postthaw sperm motility (%), live sperm (%) and intact acrosome (%) in yaks were 55.67 ± 4.67, 65.62 ± 3.23 and 89.26 ± 3.67 respectively. In conclusion, yak semen has a better cryosurvival while freezing in tris extender with 6.4 per cent glycerol and 20 per cent egg yolk following an equilibration period of 4h.SAARC J. Agri., 15(2): 215-218 (2017)

scholarly journals Pore Damage Properties and Permeability Change of Coal Caused by Freeze-Thaw Action of Liquid Nitrogen

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Bo Li ◽  
Lulu Zhang ◽  
Jianping Wei ◽  
Yongjie Ren
Keyword(s):  
Pore Structure ◽  
Liquid Nitrogen ◽  
Cold Shock ◽  
Water Saturation ◽  
Temperature Drop ◽  
Freeze Thaw ◽  
Thaw Cycle ◽  
Before And After ◽  
Porosity And Permeability ◽  
Coal Rock

A laboratory test was conducted to investigate the effect of the freeze-thaw action of liquid nitrogen on the pore structure and permeability of coal rock. First, coal rock samples with similar sound velocities and permeabilities were selected. These samples were prepared in different water saturation levels and subjected to nuclear magnetic resonance (NMR) test before and after the freeze-thaw action. Furthermore, the freeze-thaw cycle of liquid nitrogen, freezing time, and water saturation of coal rocks were controlled in permeability test. Results showed that the pore diameter, porosity, and permeability of the coal rocks increase after the freeze-thaw action of liquid nitrogen. These characteristics increase further with the increase of water saturation. The fracturing mechanisms of the freeze-thaw action of liquid nitrogen were summarized in two aspects, phase change of pore water and cold shock, and cold shock was mainly discussed. The results indicate that the effect of cold shock is still crucial at low water saturation, but it is limited by the degree of temperature drop. In general, freeze-thaw action of liquid nitrogen can cause damage to pore structure, promote the formation of fracture networks, and consequently improve the permeability of coal rock.

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