The cryoprotective effects of ascorbic acid supplementation on bovine semen quality.
To investigate the effects of ascorbic acid supplementation on standard semen quality parameters and antioxidant activities after thawing of bovine frozen semen, antioxidant ascorbic acid was added at concentrations of 2.5, 4.5, 6.5 and 8.5 mg/ml to bovine semen cryoprotective medium. The results showed that the sperm motility and motion characteristics were improved in the presence of ascorbic acid in extender, as compared to the control. The motility and straight linear velocity (VSL), linearity index (LIN), average path velocity (VAP), wobble coefficient (WOB), lateral head displacement (ALH) values and the percentage of "grade A" sperm in the extender supplemented with 4.5 mg/ml ascorbic acid were significantly higher than that of other treatment groups (P<0.05). The acrosome integrity and membrane integrity were significantly improved (P<0.05) by supplementing with 4.5 mg/ml ascorbic acid in the extender compared with a control. The extender supplemented with ascorbic acid did not lead to any improvement in superoxide dismutase (SOD) levels. The catalase (CAT) activity was higher in the extender supplemented with ascorbic acid at 4.5 mg/ml, when compared with other groups (P<0.05) and the extender supplemented with ascorbic acid significantly decreased glutathione peroxidase (GSH-Px) activity, whereas reduced glutathione (GSH) activities were significantly enhanced, compared with the control (P<0.05). Increasing the doses level of ascorbic acid decreased GSH-Px and GSH activity, the supplementation of 8.5 mg/ml ascorbic acid produced the lowest level of GSH-Px and GSH activity among groups (P<0.05). The extender supplemented with ascorbic acid could reduce the oxidative stress provoked by freezing-thawing and improve bovine semen quality. The particular properties of ascorbic acid are poorly related to its effectiveness in membrane cryopreservation. Further studies are required to determine lipid peroxidation and antioxidant capacities of ascorbic acid in cryopreserved bovine semen.
Hu JH
，Tian WQ
，Zhao XL
，Zan LS
，Wang H
，Li QW
，Xin YP
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The cryoprotective effects of vitamin B12 supplementation on bovine semen quality.
The present study aimed to investigate the effects of vitamin B(12) supplementation on standard bovine semen quality parameters and anti-oxidative enzyme activities. Vitamin B(12) was supplemented at concentrations of 1.25, 2.5, 3.75 and 5.0 mg/ml to bovine semen cryoprotective medium. The results indicated that the motility and straight line velocity, curvilinear velocity, mean coefficient, velocity of the average path values of sperm supplemented with 2.50 mg/ml vitamin B(12) were significantly higher than that of other groups (p<0.05). No significant difference was observed for linearity index, lateral head displacement values and the percentage of grade A spermatozoa between the extenders containing 2.50 and 3.75 mg/ml vitamin B(12) (p>0.05). The percentages of acrosome-intact and plasma membrane-intact spermatozoa were significantly improved (p<0.05) by supplementing with 2.50 mg/ml vitamin B(12) . The results of biochemical assay revealed that vitamin B(12) supplementation did not cause significant changes in superoxide dismutase levels compared with control (p>0.05). However, the catalase levels were higher in the treatment supplemented with vitamin B(12) at 2.50 mg/ml, when compared with other groups (p<0.05). The extender supplemented with vitamin B(12) significantly decreased glutathione peroxidase activity compared with the control (p<0.05). The supplementation of 3.75 mg/ml vitamin B(12) caused the highest value of glutathione reductase activity, compared with other groups (p<0.05). In conclusion, the extender supplemented with vitamin B(12) could reduce the oxidative stress provoked by freezing-thawing and improve bovine semen quality. Further studies are required to obtain more concrete results on the determination of lipid peroxidation and antioxidant capacities of vitamin B(12) in cryopreserved bovine semen.
Hu JH
，Tian WQ
，Zhao XL
，Zan LS
，Xin YP
，Li QW
... -
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Cryopreservation of epididymal cat spermatozoa: effects of in vitro antioxidative enzymes supplementation and lipid peroxidation induction.
Reactive oxygen species and lipid peroxidation reaction, causes of sperm damage, can be diminished by action of antioxidative enzymes. This study aimed to investigate effects of (1) the antioxidative enzymes; catalase, glutathione peroxidase and superoxide dismutase, on epipididymal cat sperm quality and (2) the lipid peroxidation reaction induced by a transition metal (ferrous ion (II); Fe(2+)) on sperm quality during the cryopreservation process. Epididymal spermatozoa harvested from 39 male cats were pooled and divided into 13 aliquots (n=13). Each aliquot was resuspended with either a Tris egg yolk extender I (control; EE-I), or the Tris egg yolk extender I supplemented with 200 U/mL catalase (EE-CAT), or 10 U/mL glutathione peroxidase (EE-GPx), or 600 U/mL superoxide dismutase (EE-SOD), and then cryopreserved. After thawing, each sperm sample was subdivided into two groups; with and without lipid peroxidation induction (EE-I plus Fe(2+), EE-CAT plus Fe(2+), EE-GPx plus Fe(2+) and EE-SOD plus Fe(2+)). Subjective sperm motility, membrane, and acrosome integrity were evaluated at the time of collection, after cooling, and at 0, 2, 4, and 6h after thawing. Motility patterns assessed by computer-assisted sperm analysis (CASA), mitochondrial activity, and DNA integrity were evaluated during post-thaw incubation, whereas percentage of lipid peroxidation was detected at 0 and 6h after thawing. The results demonstrate that catalase supplementation reduced linear motility and subjective motility immediately and 2h after thawing (P<0.05). Catalase supplementation, however, improved DNA integrity at 4h (P<0.05). Supplementation with glutathione peroxidase, compared to the control group, had a statistically significant positive effect on subjective motility at 0 and 6h, linear motility at 6h, mitochondrial activity at 6h, membrane integrity at 2 and 6h, and DNA integrity at 4h after thawing. Although superoxide dismutase had a positive effect on sperm membrane integrity at 2h after thawing (P<0.05), it significantly reduced membrane integrity after cooling, linear motility at thawing, and acrosome integrity at 2h after thawing. None of the three selected antioxidative enzymes significantly influenced acrosome integrity and none reduced the level of lipid peroxidation. Furthermore, induction of the lipid peroxidation reaction by Fe(2+) negatively affected most of the sperm quality parameters, i.e., motility and DNA integrity, during post-thaw sperm incubation (P<0.05). After thawing, there were, however, no significant differences between the control plus Fe(2+) and the antioxidative enzymes supplementation plus Fe(2+) groups. We can conclude that (1) glutathione peroxidase exhibits positive effects on post-thaw epididymal cat spermatozoa; but (2) none among the selected antioxidative enzymes could improve all sperm quality parameters; and (3) the lipid peroxidation reaction may be one cause of post-thaw epididymal sperm damage in cats, but the concentrations of antioxidative enzymes used in this study could not protect cat spermatozoa from lipid peroxidation induction.
Thuwanut P
，Chatdarong K
，Johannisson A
，Bergqvist AS
，Söderquist L
，Axnér E
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ResearchGate
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钛学术
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