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1.
A new device for storage and shipping of cell cultures – the Petaka G3 cell management device – was tested for its applicability for cooled‐storage of equine semen. Semen from three stallions was processed with EquiPro extender either without antibiotics (three ejaculates per stallion) or with gentamicin (250 mg / l; three ejaculates per stallion). Semen was either stored at five (anaerobic conditions) or 15°C (aerobic conditions) in syringes or cell culture devices. Total and progressive motility, as well as membrane integrity of spermatozoa, were evaluated from days 1 to 7 after collection with computer‐assisted semen analysis. In experiment 1 (extender without antibiotics), total motility, progressive motility and viability of spermatozoa significantly decreased over time (p < 0.05). The decrease was significantly faster at 15°C than at 5°C (p < 0.05). In the presence of gentamicin (experiment 2), this difference was no longer present. It can be concluded that cooled‐storage of equine semen in sophisticated devices for cell culture is not advantageous to syringes for successful maintenance of semen longevity.  相似文献   

2.
A specific problem in the preservation of goat semen has been the detrimental effect of seminal plasma on the viability of spermatozoa in extenders containing egg yolk or milk. The use of chemically defined extenders will have obvious advantages in liquid storage of buck semen. Our previous study showed that the self‐made mZAP extender performed better than commercial extenders, and maintained a sperm motility of 34% for 9 days and a fertilizing potential for successful pregnancies for 7 days. The aim of this study was to extend the viability and fertilizing potential of liquid‐stored goat spermatozoa by optimizing procedures for semen processing and storage in the mZAP extender. Semen samples collected from five goat bucks of the Lubei White and Boer breeds were diluted with the extender, cooled and stored at 5°C. Stored semen was evaluated for sperm viability parameters, every 48 h of storage. Data from three ejaculates of different bucks were analysed for each treatment. The percentage data were arcsine‐transformed before being analysed with anova and Duncan’s multiple comparison test. While cooling at the rate of 0.1–0.25°C/min did not affect sperm viability parameters, doing so at the rate of 0.6°C/min from 30 to 15°C reduced goat sperm motility and membrane integrity. Sperm motility and membrane integrity were significantly higher in semen coated with the extender containing 20% egg yolk than in non‐coated semen. Sperm motility, membrane integrity and acrosomal intactness were significantly higher when coated semen was 21‐fold diluted than when it was 11‐ or 51‐fold diluted and when extender was renewed at 48‐h intervals than when it was not renewed during storage. When goat semen coated with the egg yolk‐containing extender was 21‐fold diluted, cooled at the rate of 0.07–0.25°C/min, stored at 5°C and the extender renewed every 48 h, a sperm motility of 48% was maintained for 13 days, and an in vitro‐fertilizing potential similar to that of fresh semen was maintained for 11 days.  相似文献   

3.
Cooled stallion semen has a short viable life, which ranges with acceptable motility and viability from 24 up to 48 hours. The purpose of this study was to compare the effects of storage pH, the ability of three different zwitterionic buffers, and cholesterol-loaded cyclodextrins (CLC) to preserve the motility and integrity of stallion sperm cooled to 5°C for 48 hours. Fourteen ejaculates were collected and split to receive CLC or not (control group). After incubation, each sample was split into six subsamples and diluted in KMT extender containing 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid (HEPES), N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (BES), or 2-(N-morpholino)ethanesulfonic acid (MES) buffers, and the final pH was adjusted to either 7.0 or 6.6, totalizing 12 experimental groups as a function of CLC, buffer, and pH variables (2 × 3 × 2 factorial). The motility parameters and integrity of plasma and acrosome membranes (live cell index) were determined using computer-automated semen analysis and epifluorescence microscopy at 3, 6, 24, and 48 hours of cooling period. According to results, pH was not a significant source of variation for motility and live sperm over different cooling periods. However, samples diluted in BES exhibited higher progressive motility within 3 hours and higher percentages of total motile cells after 48 hours of incubation at 5°C (P < .05). After 24 hours of storage, CLC-treated sperm samples presented higher motility than control group (P < .05), and after 48 hours of incubation, CLC-treated sperm exhibited higher percentages of live, motile, and progressively motile sperms (P < .05). We inferred that equine semen diluted in KMT containing BES as buffer and CLC treatment improve the equine sperm survival during storage at 5°C for 48 hours.  相似文献   

4.
This study evaluated whether pentoxifylline (PTX) present in the flushing extender influenced the function of equine epididymal spermatozoa after recovery and after thawing. For this experiment, 58 testicles from 29 Brazilian Jumping Horses were used. Cauda epididymides of each stallion were separated and flushed with a skim milk extender, with or without 7.18 mM PTX and then subjected to the freezing process. Samples flushed with the extender containing PTX showed a significant increase in total motility, progressive motility, straight line velocity, curvilinear velocity, and percentage of rapid sperm immediately after the recovery of epididymal sperm and after 15 minutes of incubation at 37°C (P < .05). However, the presence of PTX in the flushing extender did not affect the post-thaw motility parameters or plasma membrane integrity (P > .05). The results of this study showed that the PTX present in the flushing extender improved motility parameters of recently recovered epididymal sperm and had no deleterious effects on plasma membrane integrity and freezability of equine epididymal sperm.  相似文献   

5.
This study was undertaken to investigate the effects of storage of stallion semen in a defined milk protein extender at 5 and 15°C under either anaerobic or aerobic conditions, with or without addition of the antibiotic gentamicin. Semen samples were collected from eight fertile stallions and stored for 96 h (day 0–4) and assessed daily for motility, velocity and membrane integrity (viability) using a CASA system. Samples for bacteriology assessment were taken on day 2 of storage. No significant (p > 0.05) differences in motility, velocity or viability were observed between treatments on days 0–2. On days 3 and 4, semen stored without gentamicin at 5°C had a significantly (p < 0.05) better semen quality compared with storage at 15°C without gentamicin, irrespective of oxygen exposure. On days 3 and 4, motility and velocity were greater in samples stored at 15°C with gentamicin, compared with the corresponding treatments without antibiotic (p < 0.05). This effect was also evident for viability on day 4. The decline in semen quality observed at 15°C most likely resulted from the effect of bacterial growth. Bacterial growth was the greatest in samples stored at 15°C without gentamicin, under both anaerobic and aerobic conditions (p < 0.05). Bacterial growth was inhibited by adding of gentamicin at 15°C, which accordingly reduced the decline in semen quality. Addition of antibiotic to samples stored at 5°C had no significant effect on any parameter analysed. In conclusion, storage at 15°C can be achieved by using an extender containing the antibiotic gentamicin. Storage at 5°C tended to maintain better semen quality irrespective of oxygen exposure, and did not necessitate an antibiotic treatment.  相似文献   

6.
As the preservation of the fertilizing capacity of rabbit spermatozoa for several days after semen collection remains a major target for the artificial insemination programs of rabbit breeding, a study was conducted to compare the efficacy of 5 or 15°C as holding temperature in lengthening the preservability of rabbit semen quality during 192 h of storage both in a solid (Cunigel) and a liquid (Tris-Citric acid-Glucose; TCG) extender. Six pooled semen samples (two ejaculates/male; two-three males/pool) were taken and made four aliquots: two aliquots were tenfold diluted with the TCG extender, whereas the other two were tenfold diluted with the Cunigel extender. One aliquot per diluent was stored at 5°C and the second one at 15°C. Sperm motility (light microscope), viability (SyBr-PI staining), plasma membrane functional integrity (Hypo-osmotic swelling test) and acrosome integrity (PSA-FITC staining) were recorded at 0, 48, 120 and 192 h of storage. In liquid-stored spermatozoa, mass motility and viability were significantly higher (p ≤ 0.05) in samples stored at 5°C than at 15°C at all the storage times; at 5°C resulted also higher (p ≤ 0.05) the percentages of both forward motility at 48 h and sperm functional integrity at 120 and 192 h of storage, whereas chilling temperature did not affect acrosome integrity. With the Cunigel extender, all the semen qualitative parameters were significantly higher in sample stored at 5 than 15°C over storage time (p ≤ 0.05); only acrosome integrity at 192 h was not different according to the chilling temperatures. In conclusion, 5°C were better than 15°C for the long-term storage of rabbit semen both in the TCG and Cunigel extender.  相似文献   

7.
The aim of this study was to assess extenders for cooling equine semen at 5°C and to be used in assisted reproductive technologies (ARTs). Four ejaculates were obtained from each of four stallions. Gel-free semen was diluted in three different extenders: (1) SMK, an opaque skim milk–based extender; (2) SMT, a skim milk (65%) and Tyrode medium (35%); and (3) BSAG, a clear extender containing 1% bovine serum albumin. Samples were packaged (10 mL; 50 × 106 sperm/mL) and stored in a cooling device at 5°C for 12 hours. Analyses were done at 0, 4, 8, and 12 hours after cooling. Semen was analyzed for sperm motility characteristics using a computer-assisted sperm analysis, for plasma membrane and acrosome integrity and mitochondrial membrane potential, using fluorescent probes (propidium iodide, Hoechst 33342, fluorescein isothiocyanate–conjugated Pisum sativum agglutinin, and 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolyl carbocyanine iodide [JC-1]). Morphology was evaluated with differential interference contrast microscopy and sperm chromatin integrity by the toluidine blue technique. Data were analyzed by analysis of variance and by Tukey test, with time as a repeated measure (SAS, 1998), when P < .05 was significant. In general, milk-based extenders (SMT and SMK) showed improved maintenance of semen quality compared with BSAG. Finally, the addition of skim milk to equine semen extender for cooling at 5°C for 12 hours seems to play a crucial role in sperm preservation. Although, optically clearer extenders are desired for use in ARTs, such as sperm sexing, the milk-free extender (BSAG) is less efficient for cooled-stored equine semen.  相似文献   

8.
The aim of the present study was to assess the effect of manganese (III) meso-tetrakis (4-benzoic acid) porphyrin (Mn-TBAP) on stallion sperm quality during storage at 5°C. In the present study, 18 ejaculates from three stallions were collected and diluted by INRA82 extender containing 0 (Mn-0), 100 (Mn-100), 200 (Mn-200), and 300 (Mn-300) μM of Mn-TBAP. Sperm motility, plasma membrane integrity and functionality, and lipid peroxidation as indicated by malondialdehyde (MDA) in the spermatozoa of diluted semen were evaluated in vitro at 2, 24, and 48 hours after storage at 5°C. The results showed that all evaluated sperm parameters, except MDA concentration, decreased significantly during the storage period. Total and progressive motility of spermatozoa were higher in Mn-200 extender (46.75 ± 0.58 and 27.62 ± 0.6, respectively) compared with Mn-0 (44.43 ± 0.58 and 25.13 ± 0.6, respectively) and Mn-300 (43.95 ± 0.58 and 25.28 ± 0.6, respectively) after 48 hours of storage at 5°C (P < .05). In addition, sperm plasma integrity and functionality were higher in Mn-200 extender (53.12 ± 0.6 and 46.63 ± 0.78, respectively) compared with Mn-0 (47.74 ± 0.6 and 40.96 ± 0.78, respectively), Mn-100 (48.63 ± 0.6 and 41.99 ± 078, respectively), and Mn-300 (46.11 ± 0.6 and 3.75 ± 0.78, respectively) after 48 hours of storage at 5°C (P < .05). The result showed also that MDA level was lower in Mn-100 extender (3.91 ± 0.06) compared with Mn-0 (4.51 ± 0.06), Mn-200 (4.25 ± 0.06), and Mn-300 (4.75 ± 0.06) after 48 hours of storage at 5°C (P < .05). In conclusion, INRA82 extender supplemented with 200-μM Mn-TBAP could efficiently preserve Caspian stallion spermatozoa after 48 hours of storage at 5°C.  相似文献   

9.
Insemination with chilled transported semen has become distinctly important in the horse-breeding industry. To ensure cell survival during cooled storage, semen is diluted with an appropriate extender and the concentration of seminal plasma (SP) is reduced. Nevertheless, SP plays an important immunomodulatory role in the female genital tract and supports sperm fertility. The aim of the present study was to evaluate the effect of the addition of autologous SP after cooled storage to highly concentrated stallion semen. Therefore, SP was removed by simple centrifugation of extended semen, aspiration of the supernatant, and resuspension of the sperm pellet with semen extender. Motion characteristics were evaluated after cooled storage for 48 hours at concentrations of 333 × 106 sperm/mL in comparison with stored samples at concentration of 25 × 106 sperm/mL (control). The highly concentrated semen samples were diluted with an extender containing 0%, 5%, 20%, and 80% SP directly before motility analysis. Dilution of the cooled semen with a fresh semen extender without SP (0%) increased kinematic parameters (curvilinear velocity [VCL] 137.3 vs. 151.8; straight-line velocity [VSL] 49.0 vs. 57.5; average path velocity [VAP] 69.5 vs. 79.4 μm/second; amplitude of lateral head [ALH] 3.1 vs. 3.3 μm; beat cross frequency [BCF] 31.6 vs. 33.5 Hz; P < .05) but not total motility (51% vs. 43%) and progressive motility (46% vs. 36%) compared with controls. The addition of SP after storage for 48 hours decreased sperm total motility and progressive motility regardless of SP concentration: 5 (38% and 34%), 20 (37% and 33%), and 80% SP (27% and 22%; P < .05). In contrast, kinematic parameters were enhanced by extenders containing 5% and 20% SP (VCL: 148.0 and 155.6; VSL: 59.2 and 60.9; VAP: 78.7 and 81.9; BCF: 33.4 and 35.7; ALH: 3.4 and 3.4; P < .05). However, using an extender containing 80% SP was detrimental to kinematic parameters (VCL: 151.2; VSL: 52.2; VAP: 76.9; BCF: 34.8; P < .05) except for ALH, which increased (3.5; P < .05). In conclusion, cooled storage at concentrations of 333 × 106 sperm/mL did not affect sperm motility. The addition of a fresh extender or an extender containing small concentrations of SP to highly concentrated ejaculated sperm increased kinematic values after storage; however, increasing concentrations of SP decreased sperm motility.  相似文献   

10.
The purpose of this study was to investigate the effects of semen collection into tubes containing extender supplemented with BSA on the cryosurvival of goat spermatozoa. Semen was collected from two goats into empty tubes or tubes containing 10 ml extender supplemented with 0, 0.1, 1, or 5% BSA, and the washed spermatozoa were frozen as pellets in egg yolk-trehalose extender with the addition of 0.04% SDS and 4% glycerol. Sperm motion parameters were evaluated after post-thawing and during a thermal resistance test. The acrosome status of frozen-thawed spermatozoa was also observed using FITC-PNA staining. In frozen semen that was collected into tubes containing extender supplemented with 5% BSA, the post-thawed spermatozoa exhibited a significant improvement in motion parameters and maintained high motility throughout incubation and acrosome integrity, as compared with semen collected into tubes containing extender supplemented with lower concentrations of BSA. In conclusion, semen collection into tubes with a large volume of extender containing high concentrations of BSA dramatically improves the motility and acrosome integrity of frozen-thawed spermatozoa. This suggests that the in vitro functional freezability of spermatozoa is abruptly modified by reducing contact with seminal plasma and by flash contact with BSA at ejaculation.  相似文献   

11.
Addition of Glutathione to an Extender for Frozen Equine Semen   总被引：1，自引：0，他引：1  
The manipulation of equine semen during cryopreservation reduces sperm viability and fertility because of, among other factors, membrane lipid peroxidation that makes cells highly susceptible to free radicals and reactive oxygen species (ROS). The oxidative effect caused by the generation of ROS can be reduced by the addition of antioxidants to the seminal plasma or to the extenders used for freezing. The current study was performed to test the in vitro effect of exogenous glutathione added in five different concentrations (control, 2.5 mM, 5.0 mM, 7.5 mM, and 10 mM [treatments 1-5, respectively]) to the extender for 12 stallions. Analyzed parameters were sperm motility, viability, and acrosome and plasmatic membrane integrity. Total motility was higher in treatments 1 and 2 (P < .05); viability, progressive motility, and plasmatic membrane integrity were higher in treatment 2 (P < .001). As for acrosome membrane integrity, treatment 3 showed the best results (P < .05). The addition of 2.5 mM glutathione to the freezing extender preserves total motility and increases sperm viability, progressive motility, and plasmatic membrane integrity. Concentrations above 2.5 mM were deleterious to spermatozoa.  相似文献   

12.
Flow cytometrically sex sorted spermatozoa are reduced in their fertilizing capacity, particularly when stored either in cooling extender or after freezing in liquid nitrogen. So far, preservation methods for sorted spermatozoa have differed only marginally from procedures used for unsorted semen. In the present study, a TRIS extender was modified to balance major cell damage caused by the sorting process and by liquid storage of the sorted spermatozoa. The new extender, containing a combination of antioxidants (AO) and bovine serum albumin (BSA), significantly increased the lifespan and fertilizing capacity of sex sorted spermatozoa. No significant differences were observed between unsorted controls and sorted samples for motility and status of sperm membranes as tested by fluorescein-isothiocyanat-peanut agglutinin/propidium iodide (FITC-PNA/PI). Acrosome integrity of spermatozoa was significantly better when semen was stored at 15 degrees C for 24 and 48 h in an extender containing AO with or without BSA as compared with controls (p < 0.05). There were no significant differences, in pregnancy rates of heifers inseminated at a natural oestrus, between unsorted controls (16/24, 66.7%) and both sorted groups (AO + BSA: 18/31, 58.1% and AO-BSA: 12/22, 54.5%). Additionally, it was shown for the first time that artificial insemination (AI) with liquid sexed bull spermatozoa stored for 72 h after sorting can result in pregnancy rates similar to AI with non-sorted semen.  相似文献   

13.
The objective of this experiment was to investigate whether the motility parameters and acrosome integrity of goat ejaculated spermatozoa are affected by collecting semen into tubes containing an extender, and thereby determine the significance of reducing contact between seminal plasma and the sperm membrane at ejaculation. Semen were collected from three goats into tubes containing 0, 1 or 10 ml extender, or collected into tubes containing 10 ml extender supplemented with 0.1, 1 or 5% BSA. Sperm motion parameters were evaluated immediately after collection, after washing, and during a 3-h thermal resistance test. Acrosome integrity was assessed using FITC-PNA staining. Semen collection into tubes containing 10 ml extender produced higher sperm motility, progressive motility, and acrosome integrity than that using a smaller volume of extender. Furthermore, collection into 5% BSA-containing extender exhibited higher sperm characteristics and maintained high sperm motility and progressive motility throughout incubation. In conclusion, semen collection into tubes with a large volume of extender, especially extender containing higher concentrations of BSA, improved the quality of ejaculated spermatozoa, strongly suggesting that the in vitro functional characteristics of the spermatozoa were abruptly modified by flash sperm contact with accessory sex gland fluid at ejaculation.  相似文献   

14.
This study assessed the effect of oral supplementation with the primary antioxidants and fatty acids involved in spermatogenesis (L-carnitine, selenium, vitamin E, omega-3, and omega-6) on the seminal quality in fresh, cooled, and frozen semen of stallions (n = 8), using a randomized design. The animals were divided into Group I (n = 4) and Group II (n = 4) for a 30-week experiment. The two groups alternated between nutraceutical supplementation and a placebo over the course of the experiment. Semen collections were performed in two sets: once in the middle of the experiment, before the two groups switched treatments, and once at the end. The volume, appearance, sperm concentration, spermatozoa kinetics, and membrane integrity of fresh semen were evaluated. The spermatozoa kinetics and membrane integrity of cooled (for 24, 36, and 48 hours) and frozen semen were also evaluated. No differences were observed in volume, appearance, and sperm concentration between treatment and control. However, compared with placebo, nutraceutical supplementation increased (P < .05) total motility, trajectory speed, as well as plasma and acrosomal membrane integrity in spermatozoa from fresh semen. In cooled semen, nutraceutical treatment also increased (P < .05) total motility, speed, and membrane integrity of spermatozoa compared with the control. In frozen semen, supplementation increased (P < .05) spermatozoa progressive motility and plasma membrane integrity. Our results suggest a positive, synergistic effect of the antioxidant L-carnitine and selenium on spermatozoa kinetics. Similarly, the increase in plasma and acrosomal membrane integrity could be attributed to higher concentrations of polyunsaturated fatty acids (a key cell-membrane component), combined with the prevention of excess lipid peroxidation by antioxidants. In conclusion, supplementation with nutraceuticals containing fatty acids and antioxidants improved the quality of fresh, cooled, and frozen stallion semen. Therefore, nutraceutical use should increase the success of artificial insemination with cooled and cryopreserved semen.  相似文献   

15.
1. The effects of lycopene-enriched extenders on the in vitro quality of turkey semen including lipid peroxidation were examined after chilled and frozen storage.

2. Five pools of semen diluted in extenders containing 0, 0·05 or 0·1?mg/ml of lycopene were stored at 5°C for 48?h or cryopreserved as pellets and the following variables determined in fresh samples and samples stored chilled or frozen: sperm motility, viability, osmotic resistance, DNA integrity and lipid peroxidation (as malonaldehyde production).

3. Semen quality was generally compromised after storage, especially post-freezing. However, in the presence of the highest dose of lycopene, both the viability and osmotic-resistance of chilled spermatozoa and the DNA integrity of frozen spermatozoa were similar to those of fresh spermatozoa.

4. Greater lipid peroxidation was detected in refrigerated compared to fresh or cryopreserved spermatozoa. However, spermatozoa chilled in lycopene-enriched extenders showed significantly lower malonaldehyde levels than those chilled without lycopene, while the addition of lycopene to the freezing medium served to maintain the lipid peroxidation levels observed in fresh semen.

5. In conclusion, the presence of lycopene in the extender improved the survival of turkey spermatozoa after liquid-storage and protected DNA integrity against cryodamage. The beneficial effects of lycopene observed could be related to its capacity to diminish sperm lipid peroxidation during refrigeration or cryopreservation.  相似文献   

16.
The objective of this study was to compare semen parameters and embryo recovery rates of cooled stallion semen extended with INRA 96 or BotuSemen Gold. In experiment 1, 45 ejaculates from nine mature stallions were collected, assessed, and equally split between both extenders and then extended to 50 million sperm/mL. Then, the extended semen was stored in three passive cooling containers (Equitainer, Equine Express II, and BotuFlex) for 48 hours. In experiment 2, the same ejaculates extended in experiment 1 were cushion-centrifuged, the supernatant was discarded, and the pellets were resuspended at 100 million sperm/mL with their respective extender. Semen was then cooled and stored as in experiment 1. In both experiments, sperm motility parameters, plasma membrane integrity, and high mitochondrial membrane potential were assessed at 0, 24, and 48 hours post cooling. For experiment 3, 12 mares (n = 24 cycles) were bred with 48 hour–cooled semen from one stallion. Semen was processed as described in experiment 1. Mares had embryo flushing performed by 8-day post-ovulation. In experiment 1, BotuSemen Gold displayed superior total and progressive motility relative to INRA 96 (P < .05). There were no significant differences between the types of containers in any experiment. In experiment 2, INRA 96 and BotuSemen Gold extenders had similar total and progressive motility, but BotuSemen Gold had superior sperm velocity parameters at all timepoints. Embryo recovery was identical for both extenders (50%). Finally, the results obtained herein suggest that BotuSemen Gold is a suitable alternative to be included in semen cooling tests against INRA 96 in clinical practice.  相似文献   

17.
Egg yolk-Tris is most commonly used semen extender; however, its use involves hygienic risk, interference with fertility and poor microscopic examination. Therefore, replacement of egg yolk with a plant-based component with protective effects on spermatozoa would be advantageous. In present study, we observed effect of soya milk-based extenders on dilution and liquid preservation of Murrah buffalo bull semen at 5°C up to 72 h in comparison with conventional egg yolk-Tris extender (Ext.1). In experiment one, a total of 32 buffalo semen ejaculates from four animals were extended and preserved at 5°C for 72 h in soya milk-based extender (Ext.2) with different percentages (10%, 15%, 20%, 25% and 30%) of soya milk for optimization of soya milk concentration. Semen quality was assessed for individual motility, viability, membrane integrity and acrosome integrity at 0, 24, 48 and 72 h of liquid preservation. The results of experiment one indicated that 25% soya milk is an optimum concentration for buffalo bull semen extender preparation. A modified method was used to prepare another soya milk-based extender (Ext.3). In the second experiment, two soya extenders (Ext.2 and 3) with optimized concentration (25%) of soya milk were comparatively assessed with egg yolk-Tris extender (Ext.1) for semen quality parameters at 0, 24, 48 and 72 h of liquid preservation. The individual sperm motility at 0 and 24 h following dilution were found non-significant among extenders. However, after 48 h of dilution, individual motility in Ext.3 was observed significantly (p < 0.05) higher than Ext.1. After 24, 48 and 72 h of dilution sperm membrane integrity in Ext.3 was found significantly (p < 0.05) higher than Ext.1. Overall, comparative evaluation of sperm parameters obtained revealed that Ext.3 containing 25% soya milk can be used as a substitute of egg yolk-based extender for buffalo semen liquid preservation.  相似文献   

18.
Activity of the anti-oxidative enzymes glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT), content of thiobarbituric acid reactive substances (TBARS) and SH-groups were determined in native stallion semen (n = 8 stallions). Semen was then diluted in Kenney extender, EquiPro((R)) extender either with or without addition of N-acetyl cysteine or phosphate-buffered saline (PBS) and stored for 72 h at 5 degrees C. Correlations between initial activity of enzymes and development of semen motility and membrane integrity were calculated. Activities of GSH-Px, SOD and CAT immediately after semen collections were 10.0 +/- 0.6 picokatals, 0.40 +/- 0.03 SOD units and 0.70 +/- 0.05 nanokatals/10(6) spermatozoa respectively. TBARS content was 0.06 +/- 0.01 nmol and SH-group content 1.7 +/- 0.5 mmol/10(6) spermatozoa. The loss of motile spermatozoa during storage did not differ between extenders. N-acetyl cysteine had no effect on semen motility and membrane integrity. The loss in membrane-intact spermatozoa was highest (P < 0.05) in semen diluted in PBS. Motility and membrane integrity after addition of extender were positively correlated with GSH-Px and CAT, indicating that anti-oxidative mechanisms contribute to the initial high percentage of motile and membrane-intact spermatozoa. However, in these samples the decrease in semen quality was most pronounced. No correlations existed between initial activity of anti-oxidative enzymes, peroxidation products and semen quality during storage. This indicates that once extender has been added, peroxidative damage to sperm membranes is not the predominant cause of losses in semen quality.  相似文献   

19.
Polyphenols (PFs) extracted from green tea, known to be potent anti‐oxidants, have been reported to be effective in increasing the motility and viability of mammalian sperm, preserved in a liquid form. Therefore, we tested whether PFs might also be effective for maintaining the integrity of frozen‐thawed boar spermatozoa. Ejaculates, collected from Clawn miniature pigs, were diluted in a semen extender containing various amounts of PFs (0, 0.01, 0.05, 0.1 and 0.2% w/v) and then stored at 15°C overnight. The semen samples were processed, using the straw freezing procedure, and then frozen in liquid nitrogen. After rapid thawing at 40°C, the spermatozoa were subjected to several assays to evaluate semen quality. Spermatozoa frozen in a medium containing 0.01% w/v PFs exhibited significantly (P < 0.05) higher degrees of post‐thawed viability and acrosomal integrity than those stored in the absence of PFs. However, no change in the mitochondrial activity was noted between the two groups. The inclusion of 0.01% PFs in the semen extender was significantly (P < 0.05) effective in increasing both the rates of monospermic oocyte formation and of blastocyst formation. These findings indicate that preincubation with the semen extender, containing 0.01% PFs prior to freezing, exerts a protective effect on boar sperm by preventing injuries associated with freezing‐thawing.  相似文献   

20.
Semen was collected with an artificial vagina from 4 one-year-old rams, in order to study the changes in sperm motility and membrane integrity of spermatozoa split-diluted and stored at 5 degrees C during 7 days in sodium citrate, Tris, and milk-based extenders, respectively. Sperm motility was assessed subjectively and sperm membrane integrity was determined using the fluorescent probes Calcein-AM and Ethidium homodimer. Representative samples were studied using scanning electron microscopy (SEM). The average incidence of sperm motility decreased over time in all the extenders (p < 0.001). The incidence of spermatozoa showing progressive motility and intact plasma membrane was significantly higher in semen diluted with sodium citrate than in the other 2 extenders following 4 days of dilution until the end of the study. Evaluation with SEM confirmed the findings obtained with the supra vital fluorescent dyes. The results of the present study indicated that there were no differences between sodium citrate-, Tris- or milk-based extenders when ovine liquid semen was stored at 5 degrees C during a short period (2 days). However, when semen was stored for longer time, spermatozoa in the sodium citrate-based extender sustained its viability better.  相似文献   

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