Goat Spermatozoa Research Articles

Procyanidins supplemented soybean lecithin-based extenders improve post-thaw quality of goat spermatozoa

Procyanidins supplemented soybean lecithin-based extenders improve post-thaw quality of goat spermatozoa

Kualitas Spermatozoa Kambing Peranakan Boer yang Dipreservasi dengan Pengencer Air Kelapa Muda dan Ekstrak Daun Kelor

ABSTRACT
 Moringa leaves contain various nutrients that can protect spermatozoa during semen preservation. The purpose of this study was to examine the ability of moringa leaf extract with different concentrations of coconut water extender on the quality of Boer crossbreed goat spermatozoa preserved at 3-5°C. The treatment of this study was the concentration level of moringa leaf extract in coconut water extender. Fresh semen was divided into five tubes and diluted with 80% coconut water + 20% egg yolk (P0, control), 79% coconut water + 20% egg yolk + 1% moringa leaf extract (P1), 77% coconut water + 20% egg yolk + 3% moringa leaves ectract (P2), 75% coconut water + 20% egg yolk + 5% moringa leaf extract (P3), and 73% coconut water + 20% egg yolk + 7% moringa leaf extract (P4), respectively. The variables observed were motility and spermatozoa viability. Results of this study showed that supplementation of different concentrations of moringa leaf extracts in coconut water extender was significantly (P<0.05) on motility and viability of Boer crossbreed goat spermatozoa during preservation. Supplementation of 3% moringa leaf extract in coconut water extender was better than 1, 5, and 7% in maintain the quality of Boer crossbreed goat spermatozoa during preservation.
 Keywords: Moringa Leaf Extract, Semen Preservation, Boer Crossbreed Goat

Effect of Vitamin C and E Supplementation and Combination of Both in Egg Yolk Tris Diluter on the Quality of Sapera Goat Spermatozoa in the 5 °C Cooling Process

This study was aimed to determine the effect of vitamin C, E supplementation and both combination in the egg yolk tris diluent on the quality of Sapera goat spermatozoa in 5˚C cooling process. The samples were fresh semen collected from Sapera goat. It was divided into four different treatment groups. The control treatment or P0 contained semen sample + egg yolk tris diluent, P1 contained semen sample + egg yolk tris diluent + vitamin C 0,9 mg/ml, P2 contained semen sample + egg yolk tris diluent + vitamin E 1 mg/ml, P3 contained semen sample + egg yolk tris diluent + vitamin C 0,9 mg/ml + vitamin E 1 mg/ ml. The least decrease of spermatozoa motility, viability and membrane integrity was found in P3, which was significantly different to P0 (p<0,05). However, P1 and P2 did not show significant difference in the results of motility and viability evaluation (p>0,05). It can be concluded that the combination of vitamin C and E supplementation can be an optimal antioxidants supplement in egg yolk tris diluent on the quality of Sapera goat spermatozoa in 5˚C cooling process.

Evaluation of lipidomic change in goat sperm after cryopreservation.

The current study aimed to detect the relationship between the spermatozoa cryotolerance and the post-thawed sperm lipidome. Ejaculates from 20 goats, and performed a uniform frozen-thawed procedure in this study. According to the total motility of thawed sperm of goats, semen samples were classified into HF group (High Freezers, n = 8) with >60% total motility and LF group (Low Freezers, n = 8) with < 45% total motility. The lipidomic analysis based on UHPLC-MS/MS was utilized to investigate the relationship between sperm cryotolerance and their lipid metabolites expression. The results showed that the cryotolerance of sperm from different individual goats were in great variation. The total motility of post-thawed sperm in HF group (60.93 ± 2.43%) is significantly higher than that in LF group (34.04 ± 3.41%, P < 0.01). And the post-thawed sperm in HF group exhibited significantly higher plasma membrane (59.06 ± 2.34%) and acrosome integrity (62.93 ± 1.15%) than that in LF group (34.06 ± 4.85%, 44.92 ± 2.19% respectively, P < 0.01). The total of 29 lipid subclasses and 1,133 lipid molecules in the post-thawed goat sperm were identified by lipidomics analysis. The lipid content of thawed sperm in HF group was higher than that in LF group, the lipid profile in HF group was significantly separated from LF group, which indicated that the difference in lipid composition and lipid metabolism mode of sperm between the two groups was existed, especially the expression of phosphatidylcholine and triglyceride molecules. In conclusion, the cryotolerance of sperm from different individual goats were in great variation. Sperm with high cryotolerance may be able to uptake more lipids during cryopreservation. The increase in phosphatidylcholine and triglyceride content of thawed. Sperm may relate to more active lipid anabolic processes.

Effects of freeze-drying on the quality and fertilising ability of goat sperm recovered from different parts of the epididymis

Lyophilisation is an alternative method for sperm preservation. The aim of this study was to evaluate the effects of freeze-thawing (F/T) and freeze-drying (F/D) on the quality of epididymal goat sperm. Sperm from each region of the epididymis (caput, corpus and cauda) were collected and evaluated for the expression of phospholipase C zeta (PLC-ζ), protamine 1 (PRM1), transition protein 1 (TNP1) and 2 (TNP2). The effects of F/T and F/D on sperm quality in terms of PLC-ζ expression, chromatin stability (Chromomycin A3; CMA3) and DNA integrity were examined. The fertilising ability after intracytoplasmic sperm injection (ICSI) was also tested. Fresh sperm existed PLC-ζ, PRM1, TNP1 and TNP2, irrespective of the regions of the epididymis. However, different patterns of PLC-ζ expression were found. Although PRM1, TNP1, TNP2 were still expressed after F/T or F/D, only F/T could preserve the presence of PLC-ζ. For fresh sperm, caput epididymal sperm had the lowest evidence of chromatin stability when compared to sperm harvested from other regions of the epididymis. The F/T and F/D further increased the numbers of CMA3-positive sperm (P < 0.001). In all cases, no CMA3 staining was observed in caudal epididymal sperm. The caudal epididymal sperm had significantly greater proportions of sperm with intact DNA compared with caput and corpus epididymal sperm, especially when F/T and F/D were performed. The fertilisation rates of F/D sperm tended to decrease when compared with F/T sperm (4.2 ± 3.2 vs. 13.6 ± 9.0, P = 0.08). It is concluded that the sperm recovered from the caudal epididymis is suitable for freezing and lyophilisation. However, poor fertilisation rates of F/D sperm were coincidently observed, with a deficit demonstration of PLC-ζ.

Beneficial Effect of Proline Supplementation on Goat Spermatozoa Quality during Cryopreservation.

Simple SummarySperm suffers damage from reactive oxygen species stress (ROS) during cryopreservation. Supplementation of antioxidants is suggested to reduce sperm cryodamage induced by ROS. Proline has been regarded as a powerful antioxidant. However, there is no information on the role proline plays during goat sperm cryopreservation. Thus, this study aims to investigate the effects of proline during goat sperm cryopreservation procedures. In the present study, supplementation of 2 mM proline significantly enhanced post-thaw goat sperm quality by regulating sperm redox homeostasis. Proline supplementation protects goat sperm against ROS stress through PRODH-mediated metabolism.Sperm cryopreservation contributes to the extensive utilization of artificial insemination (AI) in the daily livestock industry. However, due to the presence of few sperm with good biological function in post-thaw goat sperm, its use has been limited for AI purposes. Hence, its improvement has been the focus of many research studies. This study aimed to investigate the effects of proline supplementation of the freezing medium on goat sperm. The goat semen was cryopreserved with freezing medium supplementation of different concentrations of proline (0, 0.5, 1, 2 and 4 mM). The post-thaw sperm motility patterns, membrane integrity, acrosome integrity, lipid peroxidation (LPO) levels, malondialdehyde (MDA) levels, total antioxidant capacity (T-AOC), proline dehydrogenase (PRODH) activity, superoxide dis-mutase (SOD) activity, glutathione (GSH) levels and GSH/GSSG were evaluated. Likewise, the expression and immunofluorescent localization of PRODH in post-thaw goat sperm was also detected. It was observed that addition of 2 mM proline to the freezing medium significantly enhanced post-thaw goat sperm total motility, progressive motility, straight-linear velocity (VSL), curvilinear velocity (VCL), average path velocity (VAP), straightness (STR), linearity (LIN), membrane integrity and acrosome integrity. Interestingly, PRODH was expressed in post-thaw goat sperm, especially in the post-acrosome and sperm tail. Addition of 2 mM proline also significantly increased the post-thaw sperm PRODH activity compared to the control. Moreover, post-thaw goat sperm LPO levels and MDA levels were reduced by supplementation of 2 mM proline. Furthermore, compared to the control, the values of post-thaw goat sperm T-AOC, SOD activity, GSH level and GSH/GSSG were also significantly increased in 2 mM proline treatment. Reduction of post-thaw goat sperm apoptosis in 2 mM proline treatment was also observed as the levels of Caspase3 and Caspase9 were decreased by the supplementation with 2 mM proline. These observations suggest that the addition of 2 mM proline to the freezing medium increased post-thaw goat sperm quality by reducing oxidative stress during cryopreservation. These findings also provide novel insights into the use of proline as an efficient additive to enhance post-thaw goat sperm quality during cryopreservation.

Effects of water-soluble curcuminoid-rich extract in a solid dispersion form (CRE-SD) on the sperm characteristics, longevity and casein kinase II catalytic subunit alpha protein stability in chilled goat semen

The present study evaluated the effects of water-soluble curcuminoid-rich extract in a solid dispersion form (CRE-SD) on goat sperm qualities and sperm protein CSNK2A2 expression during liquid storage. Semen was collected from five fertile goats, using an artificial vagina. Ejaculates with a motility above 70% were cooled to 4 °C using TRIS-citric acid-fructose diluent with 10% egg yolk containing various concentrations of CRE-SD (0, 0.1, 1, 10 and 100 μg/mL). Chilled sperm were evaluated for sperm characteristics, casein kinase II catalytic subunit alpha (CSNK2A2) protein level and oxidative status up to 15 days. After 12 days of preservation, sperm motility, viability, acrosomal integrity and mitochondrial activity were significantly higher in the group preserved with 10 μg/mL CRE-SD as compared with the control group. Supplementation of CRE-SD at this concentration was also able to conserve the CSNK2A2 a significantly higher than that in control group until 9 days of cold storage, possibly by reducing oxidative stress. The molecular mass of the sperm CSNK2A2 protein detected in this study was 37 kDa; it was mostly located in the post-acrosomal region, midpiece and flagellum. These results demonstrate the possibility to use the CRE-SD as a natural antioxidant during liquid semen storage in goats.

62 Impact of Feeding Male Goats with L-Carnitine on Viability of Their Semen Stored at 4 °C for 72 Hours

Abstract Goats have the ability for genetic improvement. Artificial insemination (AI) is a significant technique used to increase genetic gain. Low fertility in goats with AI was attributed to semen quality, which can be affected by cooling/freezing procedures. Cooling of spermatozoa causes reduction in viability, and loss of fertility, mainly because of oxidative stress. L-carnitine (LC) is important for fatty acid metabolism and acts as antioxidant. Many studies have reported positive impact of LC on quality of cooled spermatozoa in different species; however, no research has been done on goats. Here, we investigated the impacts of dietary supplementation of LC on the quality of cooled goat semen. Six mature Alpine bucks were allocated into two groups; control (three bucks were fed a regular diet without LC supplementation), and treatment (three bucks were fed the normal diet plus 0.022 g/kg of BW, LC). Bucks were fed their respective diet for sixty days from January to March (non-breeding season). Semen was collected once/week and evaluated for color, consistency, volume and sperm concentrations. Diluted semen was then stored at 4 °C for 72 h. Sperm motility, viability, and membrane integrity were assessed at 0, 24, 48, and 72 h post-cooling. Results showed that colors of samples collected from the two groups were white to yellow, consistency ranged from watery to creamy, and sperm concentrations ranged from 8.1 to 12.2 x 109 sperm/ml. Sperm motility, viability and membrane integrity reduced (P≤ 0.05) in a time dependent manner in both groups. At 72 h, motility decreased by one-half as compared with 0 h (48.9% vs. 91.4% and 32.5% vs. 80.8%, in LC and control groups, respectively). In conclusion, under our experimental conditions, goat spermatozoa can be stored at 4°C for 72 h without severe loss in their quality. LC treatment has less impact on the quality of cooled goat semen.

Effect of Astragalus polysaccharides on the cryopreservation of goat semen

During cryopreservation, sperm encounters oxidative stress induced by excessive reactive oxygen species (ROS), destroying the sperm plasma membrane structure and reducing its physiological functions. The present study aimed to evaluate the effect of Astragalus polysaccharides (APS) on the cryopreservation of dairy goat semen. Semen was collected from six goats, and then qualified semen with movement >80% was selected after preliminary evaluation. The semen was divided into six aliquots, diluted with dairy goat semen extender (1:10) at 37 °C, containing 0 g/L (control), 0.1 g/L, 0.2 g/L, 0.3 g/L, 0.4 g/L and 0.5 g/L APS, cryopreserved, and stored in liquid nitrogen (−196 °C). Sperm quality was assessed after freeze-thawing. The highest sperm motility, motion performance, plasma membrane integrity, acrosome integrity, and antioxidant properties (total antioxidant capacity and levels of antioxidant enzymes) were recorded (P < 0.05) in the 0.2 g/L APS group after the semen was freeze-thawed. The control and the optimal group (0.2 g/L) were then selected to analyze the effects of APS on sperm energy metabolism (mitochondrial membrane potential [MMP] and adenosine triphosphate [ATP]), sperm apoptosis, and the expression of the AMPK signaling pathway. The results showed that treatment with 0.2 g/L APS increased sperm MMP and ATP content after freeze-thawing, reduced sperm apoptosis by regulating apoptosis-related proteins, and promoted AMPK phosphorylation by activating the AMPK signaling pathway. The cleavage rate of frozen goat sperm during in vitro fertilization (IVF) was also observed to increase. These findings suggest meaningful ways to improve cryopreservation of dairy goat semen and provide new insights into the mechanism by which APS protects sperm from oxidative damage via AMPK activation.

The effects of green synthesized anionic cupric oxide nanoparticles on Zaraibi goat spermatozoa during cryopreservation with and without removal of seminal plasma

Sperm motility, normal morphology, viability, spermatozoa DNA damage, and lipid peroxidation are all affected by semen cryopreservation. The goal of this study was to see how effective cupric oxide nanoparticles (CuONPs) are as a cryo-extender additive on post-thawed sperm parameters. An artificial vagina was used to collect semen samples from five mature Zaraibi bucks (2–3 years). Ejaculates were pooled and separated into two fractions (A&B), a fraction (A) was left without being centrifuged and a fraction (B) was centrifuged to remove seminal plasma. Both fractions were diluted with tris egg yolk citrate extender (TECE) and then divided into five equal aliquots, each supplemented with (0, 10, 20, 40, and 60 ppm/ml) CuONPs. The findings revealed that removing seminal plasma before cryopreservation harms sperm parameters. Sperm motility, viability index, membrane integrity, biochemical antioxidant marker, DNA integrity, and MDA level improved after supplementation with CuONPs up to 60 ppm/ml, the most prominent significant positive effect was obtained with the highest dose (60 ppm/ml) without removal of the seminal plasm compared to control group. In conclusion: The presence of seminal plasma with a high concentration of CuONPs (up to 60 ppm/ml) may help to mitigate the negative effects of cryo-preservation.

Green Tea Extract in the Extender Improved the Post-Thawed Semen Quality and Decreased Amino Acid Mutation of Kacang Buck Sperm.

This study was the first to combine the addition of antioxidants to a skim milk-egg yolk (SM-EY) extender and different equilibration periods to obtain higher quality post-thawed Kacang buck semen. This study aimed to determine the effects of green tea extract (GTE) on the quality of frozen Kacang goat sperm equilibrated for one and two hours. The pool of Kacang buck ejaculate was equally divided into four portions and was diluted in an SM-EY extender that contained four doses of 0, 0.05, 0.10, and 0.15 mg of GTE/100 mL for T0, T1, T2, and T3 groups, respectively. The aliquots were treated for an equilibration period of 1-2 h before further processing as frozen semen. Post-thawed semen quality was evaluated for sperm quality. The Sanger method was used for DNA sequencing, and the amino acid sequence was read using MEGA v.7.0. The post-thawed semen of the T2 group that was equilibrated for one hour had the highest semen quality. Pre-freezing motility had the highest determination coefficient compared to post-thawed sperm motility. This study is the first to report amino acid mutation due to freeze-thawing. The frequency of amino acid mutations revealed that T2 was the least mutated amino acid. Glycine, valine, leucine, serine, and asparagine strongly correlated to post-thawed sperm motility. It can be concluded that a combination of 0.1 mg GTE/100 mL extender as an antioxidant and one-hour equilibration period resulted in the best post-thawed Kacang buck semen quality.

Use of Purslane (Portulaca oleracea) Extracts and Cysteine in Diluents to Improve the Cryopreserved Quality of Goat Sperm.

Highlights Using cysteine and purslane extracts in extenders improved significantly the post-thaw sperm characteristics. Sperm viability, DNA integrity, and mitochondrial activity demonstrate an improvement in post-thaw sperm. Malondialdehyde production was decreased based on the positive effects of treated extenders. The obtained results demonstrate that supplementation of 50 μg/mL of purslane methanolic extract with cysteine to freezing extenders was significantly superior compared with other treatments.

The Proteomic Modification of Buck Ejaculated Sperm Induced by the Cryopreservation Process.

Using two-dimensional electrophoresis along with mass spectroscopy, we have investigated how the cryopreservation process affected the protein profile of goat ejaculated sperm. In this study, five bucks were used for semen collection. After removal of seminal plasma, the Tris-based extender containing glycerol and egg yolk was used to freeze semen. The results indicated that the post-thaw sperm quality showed a significant reduction compared with fresh sperm. The numbers of protein spots acquired in fresh and post-thaw sperm were 2926 ± 57 and 3061 ± 81, respectively. Twenty-two different abundant proteins (DAPs) were identified between fresh sperm and frozen-thawed sperm (≥3.0-folds, p < 0.05). The abundances of 19 proteins were significantly higher in the fresh sperm than the post-thaw sperm. The results of the gene ontology annotation showed the primary location of the DAPs on sperm cytoskeleton, protein complex, cytoplasm, and mitochondria. In addition, these proteins were mainly involved in ion binding, small molecular metabolic processes, structure molecule activity, guanosine triphosphatase activity, oxidoreductase activity, and protein complex assembly. The interaction networks among these DAPs demonstrated that they may play roles in oxidoreductase activity, structure, acrosomal function, and motility of sperm. Collectively, the proteome of goat sperm was altered during the cryopreservation process, demonstrating that protein modification induced by cryopreservation may be associated with the reduced quality of goat sperm after thawing.

Novel Cold-Inducible RNA-Binding Protein (CIRBP) in goat spermatozoa: A possible cryo-stress reliever during the process of Cryopreservation

The great expectation of the dairy owners from the frozen semen industry is quality semen with high fertility. Many researchers concentrate in this area to produce frozen semen of high quality and fertility. However, the problem still exists to minimize or reduce the cryoinjury related damages that occur during the process of cryopreservation with the use of all kinds of semen additives. The success rate of the additives added semen is variable at a larger scale. Therefore, any new molecules which can effectively prevent or minimize the cryoinjury are warranted and it could be low cost substances. Our incidental finding and protein identification with molecular weight ≈ 19 kDa (cold-inducible RNA-binding protein CIRBP) in goat spermatozoa seems to be effective in minimizing cryoinjury. The identified CIRBP was confirmed via Western blotting. In future, further studies are needed to confirm the molecular mechanism of the CIRBP on the cryoinjury as semen additives, and application of this protein added semen in the field to improve the level of conception rate in cattle.

Study on cryopreservation of Guanzhong dairy goat semen with bovine semen seminal plasma

There are phospholipase and yolk agglutinase in the seminal plasma of dairy goat. The existence of these enzymes will hydrolyze lecithin in yolk into lysophosphatides and cause the change of pH value of diluent, resulting in toxic and side effects on sperm. In order to explore the effect of bovine seminal plasma replacing different doses of dairy goat seminal plasma (0%, 25%, 50%, 100%) on semen freezing quality of Guanzhong dairy goat, we used computer-aided sperm analysis (CASA) system, specific fluorescent probe detection technology, hypo osmotic swelling test (HOST), oxidation index determination and in vivo fertilization verification test to study the semen cryopreservation of Guanzhong dairy goat. The results showed that if we used bovine seminal plasma instead of 100% dairy goat seminal plasma, the sperm motility, plasma membrane functionality and acrosome integrity rate were the highest after thawing, reaching 67.04%, 38.12% and 68.83% respectively, which was significantly higher than that of other groups (P < 0.05); In terms of oxidation indexes, the values of SOD and GSH-PX in sperm of dairy goats were 199.78 U/mL and 133.55 U/L, respectively, which were significantly higher than those in other groups (P < 0.05), and the values of ROS and MDA were 397.58 U/mL and 5.16 nmol/L, respectively, which were significantly lower than those in other groups (P < 0.05); The results of artificial insemination also showed that using bovine seminal plasma instead of 100% of dairy goat seminal plasma could significantly improve the fertilization ability of sperm. The quality of frozen Guanzhong dairy goat sperm can be improved by using bovine seminal plasma instead of dairy goat seminal plasma and using bovine diluent.

The replacement of fresh egg yolk by lyophilized egg yolk in Tris-base extender in cryopreserved Boer and Saanen semen.

Egg yolk is a common cryoprotectant that can be used as a semen extender to protect the spermatozoa from damage during cryopreservation. Therefore, the aim of this study was to investigate the efficacy of fresh and lyophilized egg yolk, as a Tris-base extender, on the quality of cryopreserved goat semen. Semen from 10 rams of two different breeds (Boer and Saanen) was collected using an artificial vagina. Each ejaculate sample was divided into four equal aliquots, which contained 20% of the fresh egg yolk (a control group), and then 10%, 15%, and 20% of the lyophilized egg yolk as a Tris-base extender. Sperm motility and kinetic parameters were determined using a computer-assisted semen analyser. The results showed that the addition of 20% of the fresh egg yolk in Tris-base extender exhibited significantly higher progressive motility, progressive fast motility, distance curve line, and beat-cross frequency parameters in the post-thaw Boer and Saanen goat sperm when compared with the addition of 10%, 15%, and 20% of the lyophilized egg yolk. The percentage of total motility and immotile parameters in the post-thaw Boer and Saanen goat sperm were not significantly different between the control and 10%, 15% as well as 20% of the lyophilized egg yolk groups. Moreover, the percentage of viability parameter in the Boer and Saanen goat sperm was not significantly different between the control and 10% of the lyophilized egg yolk group but showed significant difference between the control group and 15% and 20% of the lyophilized egg yolk groups. Furthermore, the interaction between the two breeds was significantly different in terms of head activity and straightness parameter. In conclusion, the treatment with 20% of fresh egg yolk in Tris-base extender is superior to the lyophilized egg yolk. However, an addition of 10% of the lyophilized egg yolk in Tris-base extender presented the percentage of total motility and viability parameters showing no difference with 20% of fresh egg yolk. Therefore, 10% of the lyophilized egg yolk in Tris-base extender provided detail of the lyophilized egg yolk protocol in cryopreserved goat semen as an example of an alternative extender to 20% of fresh egg yolk for situations where an animal's origin represents a microbiological risk.

Synergistic effects of myo-inositol and melatonin on cryopreservation of goat spermatozoa.

Overproduction of reactive oxygen species (ROS) during sperm cryopreservation has a detrimental effect on sperm parameters. Therefore, the use of antioxidants in the sperm freezing extender can reduce ROS destructive effects. In this study, we investigated whether co-supplementation of melatonin and myo-inositol into the semen extender can improve the post-cryopreservation quality of goat spermatozoa. After the freeze-thawing process, sperm motility, viability, plasma membrane and acrosome intact morphology were improved in the combined myo-inositol and melatonin group compared to both individual and the control groups (p<.05). In addition, the mean of sperm ROS, DNA damage and lipid peroxidation were reduced in co-supplementation of myo-inositol and melatonin compared to their individual counterparts (p<.05). Therefore, the synergistic effects of myo-inositol and melatonin on the cryopreserved spermatozoa are highly likely mediated through the reduction in important factors involved in the sperm lipid peroxidation. Finally, we used the cryopreserved spermatozoa for in vitro production of embryos. Results showed that combined group of myo-inositol and melatonin improved the cleavage rate compared to both individual and control groups, although blastocyst rate was improved using both individual and combined groups. In conclusion, co-supplementation of melatonin and myo-inositol is a promising approach for the improvement of goat sperm cryopreservation.

Vitamin B7 protects DNA damage and stabilizes mitochondrial transmembrane potential from cryoinjury

The proposed study was undertaken with the objective to enhance the life and fertility potential of cryopreserved goat sperm by fortification of various concentrations of biotin in sperm medium before freezing and to evaluate its effects on oxidative status, DNA integrity and mitochondrial membrane potential (MMP). Sixty ejaculates were collected from Barbari bucks and were extended with Tris- Citric acid- Fructose diluent and biotin in different concentrations were added in sperm preparation medium (0, 1, 2, 3 and 4 µM). Sperm concentrations were adjusted to 1 × 108 mL−1 and diluted semen samples were equilibrated at 5 °C for 4 h before being cryopreserved in liquid nitrogen. The sperm motility, live sperm count, acrosome integrity, hypo-osmotic swelling positive spermatozoa, malondialdehyde (MDA), protein carbonyl content (PCC), DNA integrity and MMP were significantly (P < 0.05) improved in 3 µM biotin. The fortification of biotin reduced the detrimental effects of freezing-stress on motility, viability, plasma membrane and acrosome integrity at all concentrations compared to control and optimal response was at 3 µM concentration. The kidding percentage through artificial insemination was significantly higher (35.00%) in supplemented group (3 µM) than in non-supplemented group (30.00%) on the basis of actual kidding rate. In conclusion, biotin may be used for routine goat semen freezing protocol to get better post- thaw quality and fertility.

The Effectiveness of Time Equilibration Before Freezing in Sapera Goat Spermatozoa After Electric Separating Sperm

The purpose of this study was to determine the effectiveness of time equilibration before freezing sapera goat spermatozoa after ESS (Electric Separating Sperm) on motility, viability, membrane integrity, normality and abnormality of spermatozoa. Sapera goat spermatozoa were collected using an artificial vagina and then separated using the ESS method. This study used a completely randomized formula with three treatments, which are the equilibration time of P1 (1 hour), P2 (3 hours), and P3 (4 hours), with six replications. The analysis in this study used the ANOVA test followed by the Duncan test to determine the real difference between treatments. The conclusion of this study showed the highest percentage of motility, viability, membrane integrity, normality of sapera goat spermatozoa at 1-hour treatment, for the lowest percentage of abnormalities at 1-hour treatment. Significantly different results (p &lt;0.05) on the anode and cathode sides in all equilibration time treatments.

The addition of vitamin C in tris–egg yolk extender maintained Sapera goat semen quality in 5° C storage

Goats are small ruminants that are reared by the rural community for financial income and nutrition. This study was aimed to determine the effect of vitamin C addition in tris–egg yolk extender on the lifespan of Sapera goat spermatozoa stored in 5o C. Semen was collected from 1.5 years old Sapera male goat. The ejaculates were diluted in Tris-egg yolk (T-EY) extender added with 0, 0.2, 0.3, and 0.4 g vitamin C/mL extender for T0, T1, T2, and T3 groups, respectively. The spermatozoa motility, viability, and morphological abnormality were assessed along with storage at 5° C. The result showed that spermatozoa motility was the highest (p &lt;0.05) in T1 compared to other groups and qualified for AI use for up to 72 hours. In 24 hours storage, the spermatozoa viability was not significantly different (p &gt;0.05) among the groups. The spermatozoa viability in the T1 group was the highest (p &lt;0.05) among the groups along 48-96 hours of storage. The spermatozoa morphological abnormalities of the T1 group was the lowest (p &lt;0.05) compared to other treatment groups in the range of 24-72 hour storage. In the control group (T0), the less than 5% spermatozoa morphological abnormalities (qualified for artificial insemination) were only in the 24 hours storage, while those of the T1 group were up to 72 hours. It could be concluded that the addition of 0.2 g vitamin C/100 mL T-EY extender maintained the quality of Sapera goat semen for 72 hours at 5°C.