The present study was undertaken to assess the effect of taurine on sperm motility, viability, total sperm abnormalities, acrosomal and plasma membrane integrity, enzymatic profiles such as reduced glutathione (GSH), glutathione peroxidase (GPX), superoxide dismutase (SOD), and catalase (CAT), and biochemical profiles such as cholesterol efflux and malondialdehyde (MDA) production. A total of 50 ejaculates were collected twice a week from 8 mithun bulls, and semen was split into 4 equal aliquots and diluted with the TEYC extender. Group 1: semen was without additives (control); groups 2 to 4: semen was diluted with 25 mM, 50 mM, and 100 mM of taurine, respectively. Seminal parameters and enzymatic and biochemical profiles were assessed at 5°C. Inclusion of taurine into diluent resulted in significant (
Mithun (
Cold storage of semen is used to reduce metabolism and maintain sperm viability over an extended period of time. Research into extender development has focused on membrane stabilizing compounds, antioxidants, and cryoprotectants. It was reported that fresh mithun semen can be preserved successfully at 5°C for 3 days using a tris-egg yolk-based extender without glycerol [
The addition of antioxidants such as taurine to ovine sperm [
Eight apparently healthy mithun bulls, approximately 4 to 6 years of age, were selected from the herd derived from various hilly tracts of the NEH region of India. The average body weight of the bulls was 501 kg (493 to 507 kg) at 4 years, which increased to 530 kg (523 to 538 kg) at 6 years of age with good body condition (score 5-6) maintained under uniform feeding, housing, and lighting conditions at the National Research Centre on Mithun, Jharnapani, Nagaland, India, which lies at 25°54′30′′ North Latitude and 93°44′15′′ East Longitude at an altitude range of 250–300 MSL. Each experimental animal was offered ad libitum drinking water and 30 kg mixed jungle forages (18.4% dry matter and 10.2% crude protein) and 4 kg concentrates (87.1% dry matter and 14.5% crude protein) fortified with mineral mixture and salt daily. Semen was collected from the animals by rectal massage. Oxytocin (5 IU, intramuscular) was injected just prior to rectal palpation. Briefly, seminal vesicles were massaged centrally and backwardly for 5 min followed by the gentle milking of ampullae one by one for 3–5 min, which resulted into erection and ejaculation. During collection, the initial transparent secretions were discarded, and neat semen drops were collected in a graduated test tube with the help of a funnel. During the study, all the experimental protocols met the Institutional Animal Care and Use Committee regulations.
A total of 50 ejaculates were collected via rectal massage from the mithun twice a week from the experimental animals, and semen was pooled to eliminate individual differences. Immediately after collection, the samples were kept in a water bath at 37°C and evaluated for volume, colour, consistency, mass activity, and pH. After the preliminary evaluations, samples were subjected to the initial dilution with prewarmed (37°C) Tris egg yolk citrate extender (TEYC) (tris-hydroxymethyl aminomethane 3.028% (
Each pooled ejaculate was split into four equal aliquots and diluted with the TEYC extender with taurine. Group 1: semen was without additives (control); groups 2 to 4: semen, with 25 mM, 50 mM, and 100 mM of taurine, respectively at a final concentration of 25 × 106 spermatozoa per mL. However, pH of diluents was adjusted to 6.8–7.0 by using phosphate buffered solution. Sperm concentrations were determined with the aid of a haemocytometer [
The count of live spermatozoa was determined using eosin-nigrosin stain 5% (
The HOST was used as a complementary test to the viability assessment protocol to evaluate the functional integrity of the sperm plasma membrane. HOST relies on the resistance of the membrane to loss of permeability under stress condition of swelling in a hypo-osmotic medium [
Antioxidant profiles such as GSH, GPX, CAT, SOD, TAC, and biochemical profiles such as total cholesterol, glucose-6-phosphate dehydrogenase (G6PD or G6PDH), aspartate amino transaminase (AST), and alanine amino transaminase (ALT) were estimated by commercially available diagnostic kits, whereas LPO of sperm and seminal plasma was measured by determining MDA production, using thiobarbituric acid (TBA) as per the method of Buege and Aust [
Results were analyzed statistically and expressed as the mean ± SEM. Means were analyzed by one way analysis of variance, followed by the Tukey’s post hoc test to determine significant differences between the four experimental groups, that is, with additives or no additive on the sperm parameters using the SPSS/PC computer program (version 15.0; SPSS, Chicago, IL). Differences with values of
Effects of various doses of taurine on sperm motility, viability, and acrosomal and plasma membrane integrity in liquid storage (5°C) are presented in Figure
Effect of diluent supplementation with taurine on seminal parameters of mithun semen (* indicates
Effect of diluent supplementation with taurine on GPX, CAT, SOD, and MDA production in mithun semen (* indicates
Effect of diluent supplementation with taurine on GSH, AST, ALT, and cholesterol efflux in mithun semen (* indicates
Effect of diluent supplementation with taurine on TAC in mithun semen (* indicates
Effect of diluent supplementation with taurine on G6PDH in mithun semen (* indicates
In the present study, the results revealed that addition of taurine has improved the seminal parameters, antioxidant and biochemical profiles of mithun semen, and thus it protects the structures and functions of spermatozoa efficiently. Thus, it may enhance the quality of semen by preserving efficiently during artificial insemination procedure. To the best of our knowledge, this is the first report of the effect of taurine on seminal parameters, antioxidative enzymatic and biochemical profiles in mithun semen. Analysis of various seminal parameters such as forward progressive motility, livability, and acrosomal and plasma membrane integrity are important for extensive utilization of semen in artificial insemination, and these parameters revealed significant difference between the treatment groups. The beneficial effects of taurine in the semen preservation are due to that it is a very potent antioxidant [
Because the mammalian sperm membrane has high polyunsaturated fatty acids, it renders the sperm very susceptible to LPO, which occurs as a result of the oxidation of the membrane lipids by partially reduced oxygen molecules, such as
The results of the present study were showed that addition of 50 mM taurine improves keeping quality of mithun semen preserved at 5°C. The sperm motility was declined by the time of storage and remained over 50% for up to 30 hours. In contrast, decline rate in the motility percentage was higher in semen samples treated with 100 mM taurine or without taurine. It has been reported that the quality of chilled semen decreased with time and remained suitable for use up to 30 hours as judged by motility and morphology [
Taurine helps maintaining the integrity of normal acrosome [
Moreover, it maintains plasma, mitochondrial membrane integrity, and cytoskeleton structure of flagella of sperm as cell protecting effects. Taurine also protects SOD and CAT level in the semen extender [
AST and ALT are essential for metabolic processes which provide energy for survival, motility, and fertility of spermatozoa, and these transaminase activities in semen are good indicators of semen quality because they measure sperm membrane stability [
Glutathione (L-g-glutamyl-L-cysteinylglycine; GSH) is the most abundant nonprotein thiol in mammalian cells and is present mainly in reduced form (GSH), and only a small amount is in oxidized form (GSSG). GSH antioxidant system consists of reduced GSH, oxidized GSSG, glutathione reductase (GRD), GPX, and glutathione-s-transferase. GRD stimulates the reduction of GSSG to GSH. This ensures a steady supply of the reductive substrate (NADPH) to GPX. G6PD is required for the conversion of NADP to NADPH, which is called as GSH oxidizing-reducing cycle in sperm and seminal plasma. In the present study, GSH and GPX were higher in the seminal plasma of taurine added semen [
Catalase is a tetramer of 4 polypeptide chain antioxidant which is found in nearly all living organisms exposed to oxygen. It is derived from the epididymis, seminal vesicle, and detoxifies both intra- and extracellular H2O2 by reduction to H2O and O2 [
G6PD or G6PDH is a cytosolic enzyme in the pentose phosphate pathway, a metabolic pathway that supplies reducing energy to cells by maintaining the level of the coenzyme nicotinamide adenine dinucleotide phosphate (NADPH). NADPH in turn maintains the level of glutathione in these cells that helps to protect the cells against oxidative damage. In the present study, the G6PDH level was higher in the taurine treated semen as indicates it increases the function of antioxidant glutathione and glucose/fructose utilization by the sperm cells leads to good semen quality [
Taurine prevents efflux of cholesterol from the sperm membrane and MDA production in diluents which indicates it prevents premature capacitation and acrosomal reaction that act as an antioxidant. Along with phospholipids, cholesterol is necessary for cell physical integrity and ensures fluidity of the cell membrane. Cholesterol plays a special role in the sperm membrane because its release from the sperm membrane initiates the key step in the process of capacitation and acrosome reaction that is crucial for fertilization [
In this study, improvements observed in sperm quality may be attributed to prevent excessive generation of free radicals, produced by spermatozoa themselves, by means of their antioxidant property of taurine. It was concluded that the possible protective effects of taurine supplementation enhance the antioxidant enzyme content and prevent efflux of cholesterol and phospholipids from cell membrane and MDA production. Thus, it may protect the spermatozoa during preservation and enhance the fertility in this species. Future sperm preservation/cryoprotective studies are warranted to confirm the present findings.