The objective of the present study was to investigate the effects of rose (Rosa damascena) extract on the quality of frozen-thawed rooster spermatozoa. Sixteen roosters divided into four aliquots and received a diet containing 0, 50, 100, 150 mg/kg of rose extract. On the 0, 14, 28, 42, 56 and 70th day of experiment, semen fluid were collected and sperm motility characteristics including total motility, progressive motility, linearity (LIN), straight line velocity (VSL), amplitude of lateral head displacement (ALH), curvilinear velocity (VCL), average path velocity (VAP), and sperm viability, acrosome integrity and lipid peroxidation were evaluated after cooling-thawing procedure. The treatments with 100 and 150 mg of rose extract significantly improved post-thaw total motility, ALH, VSL, and sperm viability, and reduced lipid membrane peroxidation (P

The highest damage to the sperm is caused by oxidative stress due to the production of reactive oxygen species (ROS). Semen cryopreservation causes some structural, biochemical and functional changes, which leads to various problems in sperm transport, survival and fertility rate in domestic animals. Also, sperm metabolism produces ROS, which is potentially harmful to the sperm plasma membrane integrity loss of motility .High concentrations of ROS have negative effects on sperm quality and increased degradation of DNA, lipid peroxidation and oxidative stress which inhibits sperm motility and changes in sperm infrastructure and finally the reduction of fertility.
Seminal plasma contains enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase (CAT) which have an important role in the inhibition of the deleterious effects of ROS. Lipid peroxidation may be due to lack of coordination between SOD, GPX, and CAT in seminal plasma or deficiency of total antioxidant capacity of the cell. Ellagic acid is a polyphenol compound that can be found in green tea and other natural resources such as pomegranate, strawberry, raspberry, walnut and eucalyptus bark. Ellagic acid showed antioxidant and anti-apoptotic effects, it can delay or prevent cellular oxidation; ultimately reduce oxidative stress.Ellagic acid increases the activity of enzymes SOD, CAT and GPx by preventing free radical attack to cells. This research was conducted to evaluate the antioxidant effect of different levels of Elligic acid on microscopic, biochemical parameters, antioxidant enzyme activities and total antioxidant capacity of ram semen after freezing-thawing process.
Five Ghezel rams were used. Semen samples were collected twice a week using artificial vagina. In order to remove individual effects, the semen samples were pooled together. Different levels of ellagic acid (0.25, 0.5, 1, 1.5 mM) were added in diluent of tris-lecithin based. After processing and freezing, the samples were stored in liquid nitrogen until the time of evaluation. After thawing of semen samples, sperm motility parameters were evaluated using CASA system, viability by with Eosin-nigrosin staining, membrane integrity with a solution of hypo-osmotic, sperm abnormalities using a solution of Hancock, lipid peroxidation by measuring malondialdehyde and the seminal plasma antioxidant enzymes of glutathione peroxidase, superoxide dismutase and total antioxidant capacity using the RANDOX Laboratories kit. The data were analyzed with SAS (9.1.3) software using GLM procedures.
The results showed that the levels of 0.25, 0.5, 1 and 1.5 mM ellagic acid improved survival and total motility parameters (P The findings of this study showed that the diluent containing 0.5 mM ellagic acid significantly improved sperm parameters compared to other levels, after freezing-thawing process.

One of the freezing semen disadvantages is increasing of free radicals leading to enhanced sperm membrane peroxidation then consequently causes lowered semen fertility after thawing. Some plants bearing high level of natural antioxidants can help to decrease semen peroxidation. The objective of the present study is to evaluate the ability of rosemary extract to protect ram sperm from freeze-thaw damages. In this study, ejaculates were collected from 4 Zandi rams. Semen samples were pooled after each sampling. Experimental treatment included five levels of rosemary aqueous extract (0, 2.5, 5, 7.5 and 10 percent). Then samples were frozen and transferred to liquid nitrogen tanks for three weeks. After thawing sperm quality parameters including total and progressive motility, abnormalities, sperm membrane integrity and MDA production were evaluated. The results showed that rosemary extract significantly improved total and progressive motility, sperm membrane integrity and viability, and decreased sperm abnormalities and MDA production at levels of 5 and 7.5 percent (p<0.05). According to the obtained data, adding specific levels of rosemary extract in the diluent will have a protective effect in the freezing-thawing process of Zandi rams sperm.

Oxidative stress during freezing-thawing reduces motility, viability, membrane functions, antioxidant capacity and finally sperm fertility. Salvia sahendica has antioxidant properties due to phenolic, flavonoids, phenolic acids and phenolic diterpenes compounds. The purpose of this study was to investigate the effect of Salvia sahendica extract as a natural antioxidant on quality of frozen-thawed Holstein bull sperms. In this study, semen samples were collected from three Holstein bulls twice a week using artificial vagina and ejaculates were pooled in order to eliminate the individual effects of bulls. Different levels of Salvia sahendica ethanol extract (0, 2, 4, 8, 12, 16 and 20 ml in dl diluent solution) (which determined based on previous tests) were added to diluents based on egg yolk-citrate. Following cooling and equilibration stage of semen samples, the samples were exposed to nitrogen vapor to be frozen and stored in liquid nitrogen until evaluation. Motility, viability, membrane integrity and the lipid peroxidation parameters of the samples were evaluated after thawing. The results of evaluation showed that, the addition of 2 and 4 ml/dl of Salvia sahendica extract improved motility parameters significantly and addition of 4 ml/dl extract improved viability and plasma membrane integrity of sperms significantly process compared to the control group following freeze-thawing process (P <0.05). Addition of 4 ml/dl extract reduced malondialdehyde concentrations compared to the other groups, however, the differences were not statistically significant. Also, addition of 16 and 20 ml/dl extract had a significantly negative effect on all evaluated traits (P<0.05).

The cryopreservation of spermatozoa has provided special opportunities for the preservation of genetic resources and improving breed programs by the artificial insemination technique (Holt, 1996). Sperm cryopreservation stimulates intracellular ice crystals formation, increasing osmotic and chilling injury that causes sperm cell damage (Isachenko et al. 2003). Freezing and thawing processes impose physical and chemical insults on the sperm membrane that decrease sperm viability and fertilizing ability (Alvarez and Storey, 1992). Both damages are associated with excessive generation of reactive oxygen species (ROS) and peroxidation of the phospholipids in the membrane (Wang et al. 1997; Lasso et al. 1994). High concentrations of ROS have a negative effect on sperm quality and increased degradation of DNA, lipid peroxidation and oxidative stress which inhibits sperm motility and changes the structure of sperm and finally the reduces the fertility (Aitken et al., 1997; Agarwal and Saleh, 2002; Khosrowbeygi and Zarghami, 2007; Zalata et al., 1995). Seminal plasma contains enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase (CAT) which have an important role in the inhibition of the deleterious effects of ROS. Lipid peroxidation may be done due to lack of coordination between SOD, GPX, and CAT in seminal plasma or deficiency of total antioxidant capacity of the cell (Tavilani et al., 2008). Ascorbic acid is the main water-soluble antioxidant which acts as a scavenger for a whole range of reactive oxygen species. Ascorbic acid acts as a strong electron donor reacts with superoxide, peroxide and hydroxyl to form dehydroascorbic acid. Ascorbic acid protects sperm DNA against damage caused by H2O2 radical and reduces the amount of nitrite. This study was conducted to study the effects of adding ascorbic acid on sperm quality, plasma lipid peroxidation and antioxidant enzyme activities of ram semen after freeze- thawing process.
Material and Five sexually mature Ghezel rams (3 to 4 years of age) were used. Semen samples were collected every three days using an artificial vagina. In order to eliminate the individual differences, ejaculates containing sperm with >80% progressive motility, volume of 0.75 to 2 mL, sperm concentrations greater than 3 × 109 sperm/mL and sperm abnormalities of less than 10%, were pooled. Different levels of Ascorbic acid (0, 0.5, 1, 1.5 and 2 mg mL-1) were added in tris-egg yolk based diluent. After processing and freezing, the samples were stored in liquid nitrogen until the time of evaluation. After thawing of semen samples, Sperm motility characteristics were analyzed using computer-assisted sperm analysis. The sperm viability was determined by means of the nigrosin–eosin staining method. The hypo-osmotic swelling test (HOS-test) was used to evaluate functional sperm plasma membrane integrity after freeze-thawing. Sperm abnormalities were assessed using Hancock solution. Lipid peroxidation was measured with determining malondialdehyde and the seminal plasma antioxidant enzymes of glutathione peroxidase, superoxide dismutase and total antioxidant capacity using the RANDOX Laboratories kit. Statistical analyses were performed using SAS software (9.1.3) software using GLM procedures. The least square means were calculated to determine the differences between the experimental treatments for the post-thaw evaluation times.
The results showed that the 1 and 1.5 mg/mL ascorbic acid significantly improved the total motility (TM), progressive motility (PM), curvilinear velocity (VCL) compared to the control group (P

Introduction Semen cryopreservation has detrimental effects on sperm cell organelles, including cell membranes, mitochondria, and DNA due to the increased level of reactive oxygen species (ROS). Oxidation of polyunsaturated fatty acids in the sperm cell membranes is caused by high levels of ROS produced during the freezing-thawing process. This problematic condition causes decreased sperm motility, membrane integrity, increased metabolic changes and, ultimately, decreased fertility of the sperm. The addition of antioxidant compounds to the semen extender before semen cryopreservation can decrease ROS levels and their deleterious effects on spermatozoa. This study was conducted to assess the antioxidant effect of different levels of Rosa canina extract on microscopic and biochemical parameters and antioxidant enzyme activities of Ghezel ram semen after freezing-thawing process.
Materials and Methods Semen samples were collected from five Ghezel rams. Ejaculates were collected twice a week. To eliminate individual effects, ejaculates containing sperm with >80% progressive motility, volume of 0.75-2 mL, sperm concentrations greater than 3×109 sperm/mL and sperm abnormalities of less than 10% were pooled. Different concentrations of Rosa canina extract (0, 100, 150, 200 μL/mL) were added into the tris-egg yolk based diluent. After processing and freezing, the samples were stored in liquid nitrogen until the time of evaluation. Sperm motility characteristics were analyzed using computer-assisted sperm analysis (CASA). Sperm motility parameters including total motility, progressive motility, average path velocity, straight-line velocity, curvilinear velocity, linearity, straightness, amplitude of lateral head displacements and beat/cross frequency of sperm, viability, membrane integrity, sperm abnormalities, lipid peroxidation, antioxidant enzymes of glutathione peroxidase, superoxide dismutase and total antioxidant capacity were evaluated after thawing. Statistical analyses were performed using SAS software. The data were analyzed using the GLM procedure. Tukey–Kramer test were used to determine the significance differences between the experimental treatments. Significant differences were reported at the level of 5 percent.
Results and Discussion The results of lipid peroxidation (LPO) measurements show that adding 100 μl / ml Rosa canina extract to the diluent medium reduced the amount of malondialdehyde, which was not significant in comparison to the control group. This could be indicative of the beneficial effect of Rosa canina extract to reduce the process of lipid peroxidation of the sperm membrane. The results showed no significant difference in malondialdehyde and morphology of sperm in treatments containing Rosa canina extract compared to the control group. The addition of Rosa canina extract was not significantly effect in morphology and structure of sperms compared to the control group. However, the percentage of abnormal sperms decreased at levels of 100 and 150 μL/mL compared to the control group. The addition of 150 μL/mL of Rosa canina extract significantly improved the viability and plasma membrane integrity of sperms after freezing-thawing compared to the control and other treatment groups (P

The aim of this study was to determine the effects of different pre-freezing equilibration times (2, 4 and 6 h) on quality parameters if post-thawed buck semen using soybean lecithin supplemented extender. In this study, semen samples were collected from four Mahabadi bucks. After collection, semen samples were primarily evaluated and pooled together. Afterward, pooled semen samples were divided into three equal parts and diluted in semen extender containing soybean lecithin. Then, after spending different equilibration times diluted semen samples were frozen. After thawing, sperm motility characteristics, plasma membrane integrity and functionality, abnormality and lipid peroxidation were evaluated. Base on the obtained results showed that six h-equilibration time resulted in higher progressive motility (25.6 ± 1.39 %) compared to other groups (P

The aim of this study was to investigate the antioxidant effect of coenzyme Q10 on ram semen quality after the freezing-thawing process. Five Ghezel rams were used for sperm collection twice a week in five replicates. This experiment consisted of 5 treatments, coenzyme Q10 at four levels (0.5, 1, 2 and 2.5 μmol) and control group (without antioxidant). In this study, motility, viability, membrane integrity, abnormality parameters of sperm, malondialdehyde, total antioxidant capacity, glutathione peroxidase and superoxide dismutase activity were measured following freeze-thawing. The results showed that the total motility in samples with 0.5 and 1 μM of coenzyme Q10 antioxidant was significantly higher than the control group (p

The objective of this study was to investigate the effect of different levels of L-Carnitine on Ghezel ram semen characteristics after freeze-thawing. Semen samples were collected from 5 healthy and mature rams using an artificial vagina. Different concentrations of 35, 70, 105, mM L-Carnitine were diluted with lecithin-based semen extender and cooled to 5°C and then placed in liquid nitrogen vapor and finally were stored in liquid nitrogen tank. The experiment was carried out on the basis of completely randomized design with 5 replicates. The results showed that the highest total motility and progressive motility were observed in groups containing 70 and 105 mM L-Carnitine that were not significantly different from the control group. The lowest motility recorded in this experiment was the first level of L-Carnitine. In other parameters only VAP, ALH, VCL and LIN were significantly different from the control group. The first level of L-Carnitin was statistically significant in morphology compared to the control group. Addition of 105 mM of L-Carnitine caused higher membrane integrity and lower MDA level compared to 35 mM group (p

Oxidative stress during freeze-thawing process causes reduction in motility, viability, membrane functions and finally sperm fertility. The aim of this study was to determine the effect of Chamomile extract as natural antioxidant on quality of cryopreserved ram sperm. In this study semen was collected from five mature rams twice a week using an artificial vagina and the ejaculates were pooled equally in order to eliminate the individual effects. Different levels of extract of Chamomile (0, 50, 66.66, 100 and 200 ml/dl of diluent solution) were added to diluent based tris-egg yolk. After cooling, filling and sealing of the samples, they were frozen with nitrogen vapor and immersed in liquid nitrogen and were stored until evaluation time. After thawing, results showed that addition of 66.66 ml/dl of extract increased total motility, progressive motility, curvilinear velocity, straight–line velocity, linearity, viability and plasma membrane integrity of sperm compared to the control and other treatment groups (p<0.05). Also addition of 200 ml/dl extract had significant negative effects on motility parameters, viability and plasma membrane integrity of sperm (p<0.05). Inclusion of different doses of Chamomile extract caused no significant reduction on malondialdehyde concentration in comparison to control but this reduction was significant at 66.66 ml/dl in comparison to 100 and 200 ml/dl of extract. Conflict of interest: None declared.