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Bovine serum albumin: survival and osmolarity effect in bovine spermatozoa stored above freezing point

Nang CF, Osman K, Budin SB, Ismail MI, Jaffar FH, Mohamad SF, et al.

Andrologia, 2012 May;44 Suppl 1:447-53.
PMID: 21806660 DOI: 10.1111/j.1439-0272.2011.01203.x

Liquid nitrogen preservation in remote farms is a limitation. The goal of this study was to determine optimum temperature above freezing point for bovine spermatozoa preservation using bovine serum albumin (BSA) as a supplementation. Pooled semen sample from three ejaculates was subjected to various BSA concentration (1, 4, 8 and 12 mg ml(-1)), before incubation in different above freezing point temperatures (4, 25 and 37 °C). Viability assessment was carried out against time from day 0 (fresh sample) until all spermatozoa become nonviable. Optimal condition for bovine spermatozoa storage was at 4 °C with 1 mg ml(-1) BSA for almost 7 days. BSA improved bovine spermatozoa viability declining rate to 44.28% at day 4 and 57.59% at day 7 compared to control, with 80.54% and 98.57% at day 4 and 7 respectively. Increase in BSA concentration did not improve sperm viability. Our results also confirmed that there was a strong negative correlation between media osmolarity and bovine spermatozoa survival rate with r = 0.885, P < 0.0001. Bovine serum albumin helps to improve survival rate of bovine spermatozoa stored above freezing point.

Matched MeSH terms: Semen Preservation*
The effects of magnetic separation on cryopreserved bovine spermatozoa motility, viability and cryo-capacitation status

Faezah SS, Zuraina FM, Farah JH, Khairul O, Hilwani NI, Iswadi MI, et al.

Zygote, 2014 Aug;22(3):378-86.
PMID: 23237064 DOI: 10.1017/S0967199412000597

Cryopreservation is a technique used to preserve cells for long-time storage. It is widely used in agriculture to store male gametes in liquid nitrogen. The aim of this study was to determine the optimum thawing temperature and time for samples subjected to annexin V magnetic-activated cell sorting (AnMACS) as the sperm preparation technique. Pooled semen samples from three ejaculates were divided into two groups. The treatment group was subjected both to AnMACS and to being cryopreserved, whilst the control group was cryopreserved directly without MACS. Post-thaw analysis was carried out for samples thawed at either 20°C for 13 s, 37°C for 30 s, 40°C for 7 s, 60°C for 6 s or 80°C for 5 s. Sperm kinematics, viability and capacitation status were determined for samples subjected to all thawing temperatures described. Results showed that thawing at 37°C for 13 s for MACS-processed samples was a superior option compared with other thawing procedures; there was a significant difference in P < 0.05 values for curvilinear velocity (VCL μm/s) and sperm straightness (STR %) when samples were thawed at 40°C for 7 s, with fewer capacitated spermatozoa (P < 0.05) when samples were thawed at 37°C for 30 s, 40°C for 7 s or 60°C for 6 s. Hence, we can speculate that the use of AnMACS as the sperm preparation technique can somehow enhance sperm cryosurvival rate after cryopreservation, however the fertilization potential of these cells has yet to be determined.

Matched MeSH terms: Semen Preservation/methods
Effect of ascorbic acid concentrations, methods of cooling and freezing on Boer goat semen cryopreservation

Memon AA, Wahid H, Rosnina Y, Goh YM, Ebrahimi M, Nadia FM

Reprod. Domest. Anim., 2013 Apr;48(2):325-30.
PMID: 22909427 DOI: 10.1111/j.1439-0531.2012.02155.x

To improve the Boer goat semen quality during cryopreservation process, three experiments were carried out to investigate the effect of (i) different concentration of ascorbic acid supplementation (ii) rate of cooling with chilled semen characteristics and (iii) method of freezing on post-thaw Boer goat sperm using Tris-based extender. Ascorbic acid at 8.5 mg/ml improved the sperm parameters (motility, integrity of membrane and acrosome, morphology and viability), compared to control in cooled samples (p < 0.05). With regard to other concentrations and post-thawed parameters, ascorbic acid at 2.5-8.5 mg/ml led to higher percentages of sperm motility and integrities of membrane and acrosome when compared to control (p < 0.05). Slow cooling rises to higher percentages of sperm motility, acrosome integrity and viability, in comparison with fast cooling, in terms of cooled and frozen samples (p < 0.05). Programmable freezing method produced the higher percentages of sperm motility, integrities of membrane and acrosome and viability when compared to the freezing method of polystyrene box during goat sperm freezing (p < 0.05). In conclusion, chilled and post-thawed sperm quality of Boer goat was improved when a Tris-based extender supplemented with ascorbic acid was used at stages of different cooling rates and freezing methods.

Matched MeSH terms: Semen Preservation/methods; Semen Preservation/veterinary*
Effect of butylated hydroxytoluene on cryopreservation of Boer goat semen in Tris egg yolk extender

Memon AA, Wahid H, Rosnina Y, Goh YM, Ebrahimi M, Nadia FM, et al.

Anim. Reprod. Sci., 2011 Nov;129(1-2):44-9.
PMID: 22024366 DOI: 10.1016/j.anireprosci.2011.10.004

The aim of this study was to determine the effect of butylated hydroxytoluene (BHT), a lipid-soluble anti-oxidant added in different concentrations to the Tris egg yolk extenders on semen cytological parameters pre freezing and post thawing (motility, morphology, viability, acrosome integrity and membrane integrity) of Boer goat spermatozoa. A total of 40 ejaculates from four Boer goat bucks were collected using an artificial vagina. Ten replicates of the ejaculates were diluted with a Tris egg yolk based extender which contained various concentrations (0.5mM, 1.0mM, 2.0mM and 3.0mM) of butylated hydroxytoluene while one sample was processed without supplementation of antioxidant and served as control. The diluted semen was cooled at 4°C and loaded into the straw and then stored in liquid nitrogen. It was evident that supplementation of BHT produces positive effect in terms of motility, membrane integrity and acrosome integrity in comparison with the control group in cooled and frozen Boer goat semen. Results showed significant differences in motility, membrane integrity, acrosome integrity and viability of cooled and frozen Boer goat spermatozoa at different concentrations. Motility, membrane integrity, acrosome integrity and viability was significantly higher in all treated groups than the control group (P<0.05) while there was no significant differences (P>0.05) in morphology trait between all group in cooled semen. However, improvement (P<0.05) was observed only in terms of the membrane integrity and acrosome integrity compared to the control and other treated groups in frozen semen. In conclusion, BHT can be used in cryopreservation of Boer goat semen in order to reduce the oxidative stress on spermatozoa.

Matched MeSH terms: Semen Preservation/methods; Semen Preservation/veterinary*
Effect of different concentrations of egg yolk and virgin coconut oil in Tris-based extenders on chilled and frozen-thawed bull semen

Tarig AA, Wahid H, Rosnina Y, Yimer N, Goh YM, Baiee FH, et al.

Anim. Reprod. Sci., 2017 Jul;182:21-27.
PMID: 28511862 DOI: 10.1016/j.anireprosci.2017.03.024

The aim of this study was to evaluate the effects of 8% virgin coconut oil (VCO) combined with different percentages of egg yolk in Tris extender on the quality of chilled and frozen-thawed bull semen. A total of 24 ejaculates from four bulls were collected using an electroejaculator. Semen samples were diluted with 8% VCO in Tris extender which contained different concentrations 0% (control), 4%, 8%, 12%, 16% and 20% egg yolk. The diluted semen samples were divided into two fractions: one was chilled and stored at 4°C until evaluation after 24, 72, and 144h; the second fraction was processed by chilling for 3h at 4°C to equilibrate, then packaged in 0.25ml straws and frozen and stored in liquid nitrogen at -196°C until evaluation after 7 and 14 days. Both chilled and frozen semen samples were then thawed at 37°C and assessed for general motility using computer-assisted semen analysis (CASA), viability, acrosome integrity, and morphology (eosin-nigrosin), membrane integrity (hypo-osmotic swelling test) and lipid peroxidation (thiobarbituric acid-reactive substances (TBARS)). The results indicate treatments with 8%, 12%, 16% and 20% egg yolk with 8% VCO had greater sperm quality (P<0.05) as compared with the control. The treatment with 20% egg yolk had the greatest sperm quality (P<0.05) among the treated groups for both chilled and frozen-thawed semen. In conclusion, the use of 8% VCO combined with 20% egg yolk in a Tris-based extender enhanced the values for chilled and frozen-thawed quality variables of bull sperm.

Matched MeSH terms: Semen Preservation/methods; Semen Preservation/veterinary*
Impact of Eurycoma longifolia extract on DNA integrity, lipid peroxidation, and functional parameters in chilled and cryopreserved bull sperm

Baiee FH, Wahid H, Rosnina Y, Ariff O, Yimer N, Jeber Z, et al.

Cryobiology, 2018 02;80:43-50.
PMID: 29269043 DOI: 10.1016/j.cryobiol.2017.12.006

This study aims to assess the effect of Eurycoma longifolia aqueous extract on chilled and cryopreserved quality of bull sperm. Semen samples were obtained from four Simmental-Brangus. Each sample was divided into two fractions: the first fraction was used for chilling the semen, and the second fraction was used for the freezing process. Both fractions were extended with Tris-egg yolk extender supplemented with 0.0, 0.25, 0.5, 1.0, 2.5, 5.0, and 7.5 mg/ml Eurycoma longifolia aqueous extract. The diluted chilled fraction was chilled at 5 °C for 6 days, whereas the frozen-thawed fraction was frozen in liquid nitrogen. Data revealed that 1 mg/ml E. longifolia aqueous extract yielded significantly (p semen evaluation. For cryopreserved sperm, a significant difference (p semen and 5 mg/ml E. longifolia aqueous extract to cryopreserved sperm into Tris-egg yolk extender helps in maintaining superior quality of bull spermatozoa during chilling and freezing.

Matched MeSH terms: Semen Preservation/methods*; Semen Preservation/veterinary
Fulltext Effects of four extenders on the quality of frozen semen in Arabian stallions

Alamaary MS, Haron AW, Ali M, Hiew MWH, Adamu L, Peter ID

Vet World, 2019 Jan;12(1):34-40.
PMID: 30936651 DOI: 10.14202/vetworld.2019.34-40

Aim: Different types of extenders have a variety of components which show the tolerance effect on sperm protection during freezing procedures. In the present study, we have examined the impact of the extenders HF-20 and Tris, which were locally manufactured, and they are competing with commercial extenders INRA Freeze® (IMV Technologies, France) and EquiPlus Freeze® (Minitube, Germany) on the quality of horses frozen semen.
Materials and Methods: A total of 15 ejaculates from three healthy stallions were collected and cryopreserved in the same environment. Each semen sample collected was divided into four equal parts and processed. All samples were analyzed before and after freezing for motility, viability, plasma membrane integrity, and morphology. Furthermore, twenty mares were inseminated using post-thawed semen.
Results: There were no differences observed among all extenders in all the parameters before freezing. Sperm cryopreserved using HF-20 showed better motility, viability, and plasma membrane integrity than Tris extender. The Tris extender showed the most inferior quality of post-thawed semen between all the extenders. HF-20, INRA Freeze®, and EquiPlus Freeze® extenders revealed the same capacity of semen preservation in vitro and in vivo.
Conclusion: HF-20 extender has the same quality as INRA Freeze® and EquiPlus Freeze® that can be considered as one of the best extenders for the semen cryopreservation in horses. In contrast, Tris extender needs some degree of improvement.

Matched MeSH terms: Semen Preservation
Effect of docosahexanoic acid on quality of frozen-thawed bull semen in BioXcell extender

Kaka A, Haron W, Yusoff R, Yimer N, Khumran AM, Sarsaifi K, et al.

Reprod Fertil Dev, 2017 Mar;29(3):490-495.
PMID: 28442061 DOI: 10.1071/RD15089

This study was conducted to investigate the effect of docosahexanoic acid (DHA) supplementation in BioXcell extender on the quality of frozen-thawed bull semen. Twenty-four ejaculates were collected from three bulls (eight from each bull). Ejaculates with motility ≥70% and normal morphology ≥80% were extended into BioXcell extender to which 0 (control), 3, 5, 10 or 15ngmL(-1) DHA was added. The supplemented semen samples were incubated at 37°C for 15min for DHA uptake by spermatozoa. Later, samples were cooled for 2h at 5°C and packaged into 0.25-mL straws, frozen in liquid nitrogen for 24h and subsequently thawed for evaluation. Results are presented as percentages ± s.e.m. Supplementation with DHA at 3ngmL(-1) significantly improved sperm functional parameters including sperm motility, normal morphology, viability, acrosome integrity and membrane integrity when compared with other supplemented groups and the control. Lipid peroxidation increased as the incorporation of DHA supplementation increased. In conclusion, 3ngmL(-1) concentration of DHA resulted in superior quality of frozen-thawed bull spermatozoa and is suggested as the optimum level of DHA to be added into BioXcell extender.

Matched MeSH terms: Semen Preservation
Effect of semen collection methods on the quality of pre- and post-thawed Bali cattle (Bos javanicus) spermatozoa

Sarsaifi K, Rosnina Y, Ariff MO, Wahid H, Hani H, Yimer N, et al.

Reprod. Domest. Anim., 2013 Dec;48(6):1006-12.
PMID: 23808560 DOI: 10.1111/rda.12206

This study was conducted to evaluate the response of Bali bulls (Bos javanicus) to different semen collection methods and their effects on fresh and post-thawed semen quality. The collection methods employed were electro-ejaculation (EE), transrectal massage (RM) and RM followed by EE (RM + EE). A total of 25 untrained Bali bulls (age between 2 and 4 years old) were subjected to the different semen collection methods. Fresh semen samples from all the 25 bulls were evaluated for volume, pH, general motility, live/dead ratio and abnormality using the conventional method. For fresh and frozen samples collected by EE and RM from 10 bulls, computer-assisted semen analysis system was used for precise quantitative measurement of motility, velocity and forward progression. Accucell photometer was used to measure sperm concentration in all samples, regardless fresh and frozen. Semen samples were obtained 100% of the attempts using EE, 84% using RM and 96% using RM + EE. There were no differences among the collection methods for fresh semen quality characteristics, including motility, morphology and viability, but pH and volume were higher for EE than RM and RM + EE. Higher sperm concentration was observed in semen collected by RM than the other two methods. Different age groups (2-3 and >3-4 years old) of the bulls did not show significant differences in volume, pH, sperm concentration, percentages in motility, live/dead ratio and normal sperm morphology. The quality of semen for general and progressive motility, VAP, VSL and VCL and acrosomal integrity after thawing was higher for RM than EE. In conclusion, Bali bulls appeared to respond best to EE and the combination of RM + EE than RM, as a method of semen collection, with a shorter time of stimulation required. Differences in age of the Bali bulls did not affect the semen quality.

Matched MeSH terms: Semen Preservation/veterinary*
Fulltext Effects of cysteine and ascorbic acid in freezing extender on sperm characteristics and level of enzymes in post-thawed stallion semen

Alamaary MS, Haron AW, Hiew MWH, Ali M

Vet Med Sci, 2020 11;6(4):666-672.
PMID: 32602662 DOI: 10.1002/vms3.315

Present study aimed to investigate the effect of adding antioxidants, cysteine and ascorbic acid on the levels of glutamic oxaloacetic transaminase (GOT), glutamic-pyruvate (GPT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and γ-glutamyl transpeptidase (GGT) enzymes of post-thawed stallion sperm. Ten ejaculates were collected each from four healthy stallions and cryopreserved using HF-20 freezing extender containing either 0 mg/ml cysteine or ascorbic acid, 0.5 mg/ml cysteine and 0.5 mg/ml ascorbic acid. All samples in freezing extender containing cysteine or ascorbic acid or none of them were assessed for sperm motility, viability, plasma membrane integrity, morphology and enzymes concentration. The ALP, LDH and GGT were significantly higher in 0-group compared with cysteine and ascorbic acid groups. The sperm motility of frozen-thawed semen with 0-group was significantly better compared with cysteine and ascorbic acid groups. The variation on viability, sperm membrane integrity and morphology were insignificant between all treated groups. Therefore, these enzymes were reduced when using antioxidants in the freezing extender. Results of the present study suggest that concentration of ALP, LDH and GGT enzymes could be used as parameters for prediction of frozen-thawed stallion semen.

Matched MeSH terms: Semen Preservation/veterinary*
Comparative characteristics of spermatozoa harvested and cryopreserved in culture and cryoprotectant media with or without donor serum proteins

Ata'Allah GA, Adenan NAM, Razali N, Palaniappan K, Saad R, Idris SK, et al.

Reprod Biol, 2017 Jun;17(2):172-179.
PMID: 28511996 DOI: 10.1016/j.repbio.2017.04.004

The objectives of this study is to evaluate the efficacy of protein-free media in the preparation, holding and crypreservation of spermatazoa for use in ART. Normozoospermic semen samples (N=71) were used to compare the effects of media on the survival and quality of spermatozoa when washed and cultured with different media with and without added proteins at 4°C, 15°C, 22°C and 37°C for 0, 4-7 and 24h. Survival and quality of spermatozoa were assessed after freeze-thaw with synthetic cryoprotectant with and without proteins. Ethics/IRB approval was obtained (Ref. 1073.52). Spermatozoa parameters were similar in all media after washing and culture for 24h. Post-thaw survival and quality of spermatozoa was not significantly different 24h after thawing of samples frozen in all cryoprotectant medium. In conclusion synthetic protein-free culture and cryoprotectant media are equal in efficacy to protein-containing media in culture and cryopreservation of spermatozoa . Use of these synthetic media are anticipated to significantly reduce the risk, potentially associated with conventional protein-containing media, of transmission of disease and possibly harmful undeclared proteins to the patient, baby and the healtcare worker. Synthetic media also ensure consistency of quality between batches of media.

Matched MeSH terms: Semen Preservation*
Successful sperm cryopreservation of the brown-marbled grouper, Epinephelus fuscoguttatus using propylene glycol as cryoprotectant

Yusoff M, Hassan BN, Ikhwanuddin M, Sheriff SM, Hashim F, Mustafa S, et al.

Cryobiology, 2018 04;81:168-173.
PMID: 29355519 DOI: 10.1016/j.cryobiol.2018.01.005

This study developed the cryopreservation of brown-marbled grouper spermatozoa for practical application. We examined 32 cryodiluents, developed from four types of cryoprotectants [propylene glycol (PG), dimethyl-sulphoxide (Me2SO), dimethyl-acetamide (DMA) and ethylene glycol (EG)] at four concentrations of 5, 10, 15 and 20% in combination with two extenders [Fetal bovine serum (FBS) and artificial seminal plasma (ASP). Cooling rates were examined by adjusting the height of straws (2.5-12.5 cm) from the liquid nitrogen (LN) vapor and cooled for 5 min before immersion into LN. DNA laddering was used to detect DNA damage in cryopreserved sperm. In fertilization trials, 0.5 g of eggs was mixed with cryopreserved sperm stored for 30 days in LN. The best motility of post-thaw sperm was achieved using 15% PG + 85% FBS (76.7 ± 8.8%); 10% PG + 90% FBS was also effective as cryodiluent. Generally, FBS gave better post-thaw motility compared to ASP. The optimum cooling rate was at 17.6 °C min-1 obtained by freezing at the height of 7.5 cm surface of LN. The results obtained showed that cryopreserved sperm of brown-marbled grouper suffered slight DNA fragmentation, which resulted in significantly lower motility. However, the fertilization (90.9 ± 0.5%), hatching (64.5 ± 4.1%) and deformity rates (3.8 ± 0.2%) obtained from cryopreserved sperm showed no significant difference with fresh sperm. These findings show that the developed protocol for cryopreservation of brown-marbled grouper sperm was viable and will be useful for successful breeding and seed production of brown-marbled grouper.

Matched MeSH terms: Semen Preservation/veterinary*
Fulltext Comparison between mechanical freezer and conventional freezing using liquid nitrogen in normozoospermia

Rahana AR, Ng SP, Leong CF, Rahimah MD

Singapore Med J, 2011 Oct;52(10):734-7.
PMID: 22009393

INTRODUCTION: This study evaluated the effect of human semen cryopreservation using an ultra-low temperature technique with a mechanical freezer at -85°C as an alternative method to the conventional liquid nitrogen technique at -196°C.
METHODS: This was a prospective experimental study conducted in the Medically Assisted Conception unit, Department of Obstetrics and Gynaecology, National University Hospital, Malaysia from January 1, 2006 to April 30, 2007. All normozoospermic semen samples were included in the study. The concentration, motility and percentage of intact DNA of each semen sample were assessed before and after freezing and thawing on Days 7 and 30 post freezing.
RESULTS: Sperm cryopreservation at -85°C was comparable to the conventional liquid nitrogen technique for a period of up to 30 days in a normozoospermic sample. There was no statistical difference in concentration (Day 7 p-value is 0.1, Day 30 p-value is 0.2), motility (Day 7 p-value is 0.9, Day 30 p-value is 0.5) and proportion of intact DNA (Day 7 p-value is 0.1, Day 30 p-value is 0.2) between the ultra-low temperature technique and conventional liquid nitrogen cryopreservation at Days 7 and 30 post thawing.
CONCLUSION: This study clearly demonstrates that short-term storage of sperm at -85°C could be a viable alternative to conventional liquid nitrogen cryopreservation at -196°C due to their comparable post-thaw results.

Matched MeSH terms: Semen Preservation/methods*
Fulltext α-Linolenic acid supplementation in BioXcell® extender can improve the quality of post-cooling and frozen-thawed bovine sperm

Kaka A, Wahid H, Rosnina Y, Yimer N, Khumran AM, Sarsaifi K, et al.

Anim. Reprod. Sci., 2015 Feb;153:1-7.
PMID: 25544152 DOI: 10.1016/j.anireprosci.2014.12.001

The present study was conducted to determine the effects of supplementing α-linolenic acid (ALA) into BioXcell(®) extender on post-cooling, post-thawed bovine spermatozoa and post thawed fatty acid composition. Twenty-four semen samples were collected from three bulls using an electro-ejaculator. Fresh semen samples were evaluated for general motility using computer assisted semen analyzer (CASA) whereas morphology and viability with eosin-nigrosin stain. Semen samples extended into BioXcell(®) were divided into five groups to which 0, 3, 5, 10 and 15 ng/ml of ALA were added, respectively. The treated samples were incubated at 37°C for 15 min for ALA uptake by sperm cells before being cooled for 2 h at 5°C. After evaluation, the cooled samples were packed into 0.25 ml straws and frozen in liquid nitrogen for 24 h before thawing and evaluation for semen quality. Evaluation of cooled and frozen-thawed semen showed that the percentages of all the sperm parameters improved with 5 ng/ml ALA supplement. ALA was higher in all treated groups than control groups than control group. In conclusion, 5 ng/ml ALA supplemented into BioXcell(®) extender improved the cooled and frozen-thawed quality of bull spermatozoa.

Matched MeSH terms: Semen Preservation/methods*; Semen Preservation/veterinary
Alpha-linolenic acid supplementation in tris extender can improve frozen-thawed bull semen quality

Kaka A, Wahid H, Rosnina Y, Yimer N, Khumran AM, Behan AA, et al.

Reprod. Domest. Anim., 2015 Feb;50(1):29-33.
PMID: 25366298 DOI: 10.1111/rda.12445

The study was conducted to evaluate the effects of α-linolenic acid (ALA) on frozen-thawed quality and fatty acid composition of bull sperm. For that, twenty-four ejaculates obtained from three bulls were diluted in a Tris extender containing 0 (control), 3, 5, 10 and 15 ng/ml of ALA. Extended semen was incubated at 37°C for 15 min, to allow absorption of ALA by sperm cell membrane. The sample was chilled for 2 h, packed into 0.25-ml straws and frozen in liquid nitrogen for 24 h. Subsequently, straws were thawed and evaluated for total sperm motility (computer-assisted semen analysis), membrane functional integrity (hypo-osmotic swelling test), viability (eosin-nigrosin), fatty acid composition (gas chromatography) and lipid peroxidation (thiobarbituric acid-reactive substances (TBARS)). A higher (p < 0.05) percentage of total sperm motility was observed in ALA groups 5 ng/ml (47.74 ± 07) and 10 ng/ml (44.90 ± 0.7) in comparison with control (34.53 ± 3.0), 3 ng/ml (34.40 ± 2.6) and 15 ng/ml (34.60 ± 2.9). Still, the 5 ng/ml ALA group presented a higher (p < 0.05) percentage of viable sperms (74.13 ± 0.8) and sperms with intact membrane (74.46 ± 09) than all other experimental groups. ALA concentration and lipid peroxidation in post-thawed sperm was higher in all treated groups when compared to the control group. As such, the addition of 5 ng/ml of ALA to Tris extender improved quality of frozen-thawed bull spermatozoa.

Matched MeSH terms: Semen Preservation/methods; Semen Preservation/veterinary*
Fulltext Effect of different concentrations of soybean lecithin and virgin coconut oil in Tris-based extender on the quality of chilled and frozen-thawed bull semen

Tarig AA, Wahid H, Rosnina Y, Yimer N, Goh YM, Baiee FH, et al.

Vet World, 2017 Jun;10(6):672-678.
PMID: 28717321 DOI: 10.14202/vetworld.2017.672-678

AIM: The objective of this study was to evaluate the effects of different concentrations of soybean lecithin (SL) and virgin coconut oil (VCO) in Tris-based extender on chilled and frozen-thawed bull semen quality parameters.
MATERIALS AND METHODS: A total of 24 ejaculates were collected from four bulls via an electroejaculator. Semen samples were diluted with 2% VCO in Tris-based extender which consists of various concentrations of SL (1, 1.25, 1.5, and 1.75%). A 20% egg yolk in Tris used as a positive control (C+). The diluted semen samples were divided into two fractions; one for chilling which were stored at 4°C for 24, 72, and 144 h before evaluated for semen quality parameters. The second fraction used for freezing was chilled for 3 h at 4°C, packed into 0.25 mL straws and then cryopreserved in liquid nitrogen. The samples were then evaluated after 7 and 14 days. Chilled and frozen semen samples were thawed at 37°C and assessed for general motility using computer-assisted semen analysis, viability, acrosome integrity and morphology (eosin-nigrosin stain), membrane integrity, and lipid peroxidation using thiobarbituric acid reaction test.
RESULTS: The results showed that all the quality parameters assessed were significantly (p<0.05) improved at 1.5% SL concentration in chilled semen. Treatment groups of 1, 1.25, 1.5, and 1.75% SL were higher in quality parameters than the control group (C+) in chilled semen. However, all the quality parameters in frozen-thawed semen were significantly higher in the C+ than the treated groups.
CONCLUSION: In conclusion, supplementation of 1.5% SL in 2% VCO Tris-based extender enhanced the chilled bull semen. However, there was no marked improvement in the frozen-thawed quality parameters after treatment.

Matched MeSH terms: Semen Preservation
Effect of sugars on characteristics of Boer goat semen after cryopreservation

Naing SW, Wahid H, Mohd Azam K, Rosnina Y, Zuki AB, Kazhal S, et al.

Anim. Reprod. Sci., 2010 Oct;122(1-2):23-8.
PMID: 20637550 DOI: 10.1016/j.anireprosci.2010.06.006

In order to improve Boer goat semen quality during cryopreservation process, the influence of sugar supplementation on semen characteristics of sperm were investigated. Three experiments were carried out to investigate the effect of (a) addition of two monosaccharides (fructose and glucose) and two disaccharides sugars (trehalose and sucrose) (b) sugar combination (fructose and trehalose, sucrose and trehalose, glucose and trehalose), and control (glucose without trehalose) (c) different concentrations of trehalose on cryopreservation using Tris based extender. The total motility, forward motility, viability, normal spermatozoa, acrosome integrity and membrane integrity were assessed subjectively. Differences were not detected among monosaccharides, but glucose increased (P<0.05) sperm forward motility in post-thaw goat semen compared to trehalose or sucrose supplementation. Semen quality did not differ (P>0.05) among disaccharide sugar supplementation. Combination of glucose and trehalose significantly improved the characteristics of Boer spermatozoa after cryopreservation (P<0.05). Supplementation of trehalose (198.24mM) into the glucose extender significantly increased total motility, forward motility, live spermatozoa, acrosome integrity and membrane integrity following cryopreservation (P<0.05). In conclusion, glucose had the better ability to support Boer sperm motility and movement patterns. Combination of monosaccharide (glucose) and disaccharide (trehalose) improved semen quality following cryopreservation. Trehalose supplementation at the concentration of 198.24mM to the glucose extender conferred the greater improvement of semen quality for Boer semen cryopreservation.

Matched MeSH terms: Semen Preservation*
Fulltext First Evaluations and Cryopreservation of Semen Samples from Sunda Clouded Leopards (Neofelis diardi)

Zainuddin ZZ, Mohamed Tarmizi MR, Yap KC, Comizzoli P, Sipangkui S

Animals (Basel), 2020 06 22;10(6).
PMID: 32580372 DOI: 10.3390/ani10061072

A better understanding of semen characteristics and resilience to freezing temperatures is necessary before developing assisted reproductive techniques and systematic biobanking for the Sunda clouded leopard. The objective of this study was to evaluate for the first time the semen and sperm quality (in fresh and frozen samples) of two captive Sunda clouded leopards in Malaysia. A total of 17 examinations of the reproductive tract (using ultrasonography) and electro-ejaculations were performed on the two leopards over a 2-year period. Samples obtained from Leopard 1 (8 years old) varied in terms of volume (402 ± 92 µL), pH (7.9 ± 0.9), sperm motility (54.5 ± 24.2%), sperm concentration (122.4 ± 84.7 × 106 sperm/mL), normal morphology (23.9 ± 12.3%), and viability (55.2 ± 18.2%). Midpiece defects represented the most common structural abnormality followed by abnormal tail and head defects. Samples from Leopard 2 (11 year old with abnormal testicular tissue) were of lesser quality. Two frozen semen samples from Leopard 1 were thawed and examined for acrosome integrity. Post-thawed samples contained <10% of motile spermatozoa but almost 50% of abnormal acrosomes. The present results emphasized the high incidence of structurally-abnormal spermatozoa, similar to the mainland clouded leopard. Post-thaw evaluations showed that the few surviving spermatozoa could potentially be used for in vitro fertilization or sperm injection. However, more individuals must be studied to validate those first findings that are exciting but still preliminary.

Matched MeSH terms: Semen Preservation
Two-dimensional polyacrylamide gel electrophoresis of Bali bull (Bos javanicus) seminal plasma proteins and their relationship with semen quality

Sarsaifi K, Haron AW, Vejayan J, Yusoff R, Hani H, Omar MA, et al.

Theriogenology, 2015 Oct 1;84(6):956-68.
PMID: 26119476 DOI: 10.1016/j.theriogenology.2015.05.035

The present study evaluated the relationship between Bali bull (Bos javanicus) seminal plasma proteins and different semen quality parameters. Semen samples from 10 mature Bali bulls were evaluated for conventional semen parameters (general motility, viability, and normal morphology), sperm functionality (acrosome reaction, sperm penetration rate, sperm penetration index), sperm kinetics (computer-assisted semen analysis parameters such as sperm velocity), and sperm morphology (acrosome and membrane integrity). Frozen-thawed semen with higher sperm motility, viability, acrosome integrity, and membrane integrity (P < 0.05) are consistently higher in acrosome reaction and sperm penetration assay. Three bulls showed the highest, four bulls displayed the medium, and the remaining three bulls showed the lowest for all sperm parameters and SPA. The proteome maps of seminal plasma from high-quality and low-quality Bali bulls were also established. Seminal plasma of both high-quality and low-quality Bali bulls was subjected to two-dimensional SDS-PAGE with isoelectric point ranged from 3 to 10 and molecular weight from 10 to 250 kDa. Approximately 116 spots were detected with Blue Silver stain, and of these spots, 29 were selected and identified by MALDI-TOF/TOF-MS/MS. A majority of the proteins visualized in the seminal plasma two-dimensional maps was successfully identified. An essential group of the identified spots represented binder of sperm 1 (BSP1), clusterin, spermadhesin, tissue inhibitor of metalloproteinases 2 (TIMP-2), and phospholipase A2 (PLA2). Other proteins found in high abundance included seminal ribonuclease, serum albumin, cationic trypsin, and peptide similar to β2 microglobulin. Thus, a reference map of Bali bull seminal plasma proteins has been generated for the very first time and can be used to relate protein pattern changes to physiopathologic events that may influence Bali bull reproductive performance.

Matched MeSH terms: Semen Preservation

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