Anti-oxidative Effects of Ethanol Extract of Origanum vulgare on Kinetics, Microscopic and Oxidative Parameters of Cryopreserved Holstein Bull Spermatozoa
محورهای موضوعی : Camelح. دقیق کیا 1 , ر. فرهادی 2 , ا. اشرفی 3 , م. مهدی‌پور 4
1 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
2 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
3 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
4 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
کلید واژه: Antioxidant, cryopreservation, bull sperm, <, i>, Origanum vulgare<, /i>, ,
چکیده مقاله :
Origanum vulgare contains high levels of phenolic compounds such as gallic acid and polyphenols such as rosmarinic acid and quercetin. The purpose of this study was to investigate the effects of Origanum vulgare extract, as a natural antioxidant, on freezing-thawing semen quality in Holstein bulls. Three Holstein bulls (5-6 years old, mean live weight 800 kg) were used for semen collection twice a week for two months. Ethanol extract of Origanum vulgare (2, 4, 8, 12, 16 and 20 mL/dL extender) was added to a citrate-yolk-base extender. After freezing-thawing sperm motility parameters, viability and membrane integrity were determined using a CASA system, eosin-nigrosin staining, and hypo-osmotic swelling test, respectively; malondialdehyde concentration and activity of superoxide dismutase and catalase were also measured. The percentages of motility were higher (P<0.05) in the freezing extender containing 4 mL/dL Origanum vulgare extract (72.34±7.98). Addition of 2 and 4 mL/dL extract of Origanum vulgare significantly improved the motility, viability and plasma membrane integrity of the spermatozoa following freezing-thawing process compared to the control group. Addition of 4 mL/dL extract significantly reduced the concentration of malondialdehyde compared to the control group (P<0.05). The activity of superoxide dismutase and catalase increased significantly by inclusion of 4 and 8 mL/dL extract of Origanum vulgare to the extender (1.86±0.18 and 1.92±0.28 U/mg protein; 4.54±0.13 and 4.28±0.28 U/mg protein, respectively). The activity of superoxide dismutase and catalase was significantly increased by inclusion of 4 and 8 mL/dL extract to the extender. In conclusion, addition of 2 and 4 mL/dL extract of Origanum vulgare to the semen extender improved the post-thawed quality of semen, which may be due to increasing in antioxidant enzyme activity and reduction in lipid peroxidation.
گیاه مرزنجوش دارای سطوح بالایی از ترکیبات فنولی نظیر اسید گالیک و فنلهایی نظیر اسید رزمارینیک و کوئرسیتین است. هدف از مطالعه حاضر بررسی اثرات عصاره مرزنجوش به عنوان آنتیاکسیدان طبیعی، بر روی کیفیت منی منجمد-یخگشایی شده گاو نر هلشتاین بود. اسپرمگیری دو بار در هفته، به مدت دو ماه و از سه گاو نر هلشتاین (6-5 ساله با میانگین وزنی 800 کیلوگرم) انجام گرفت. عصاره اتانولی مرزنجوش در مقادیر 20، 16، 12، 8، 4، 2 میلیلیتر در دسیلیتر رقیقکننده بهرقیق کننده بر پایه سیترات-زرده تخم مرغ افزوده شد. بعد از یخگشایی نمونههای منی، پارامترهای تحرکی اسپرم، زندهمانی و یکپارچگی غشاء به ترتیب توسط سیستم کاسا، رنگ آمیزی ائوزین-نیگروزین و تست التهاب هیپواسمتیک مورد ارزیابی قرار گرفتند. همچنین مقادیر مالون دی آلدهاید و فعالیت سوپراکسید دیسموتاز و کاتالاز اندازهگیری شدند. نتایج بیانگر بالا بودن درصد تحرک (98/7±34/72) در نمونه دریافت کننده 4 میلیلیتر در دسیلیتر عصاره مرزنجوش بود (05/0>P). افزودن 2 و 4 میلیلیتر در دسیلیتر عصاره مرزنجوش باعث بهبود معنیدار میزان تحرک، یکپارچگی غشاء پلاسمایی اسپرمها بعد از فرآیند انجماد-یخگشایی نسبت به گروه شاهد شد. افزودن 4 میلیلیتر در دسیلیتر عصاره مرزنجوش باعث کاهش غلظت مالون دی آلدهاید نسبت به گروه شاهد شد (05/0>P). میزان فعالیت سوپراکسید دیسموتاز و کاتالاز در نمونههای دریافت کننده 8 و 4 میلیلیتر در دسیلیتر عصاره مرزنجوش به طور معنیداری افزایش یافت (به ترتیب 18/0±86/1 و 28/0±92/1 واحد بر میلیگرم پروتئین؛ 13/0±54/4 و 28/0±28/4 واحد بر میلیگرم پروتئین) (05/0>P). میزان فعالیت سوپراکسید دیسموتاز و کاتالاز به طور معنیداری در نمونههای دریافت کننده 4 و 8 میلیلیتر در دسیلیتر عصاره مرزنجوش افزایش یافت (05/0>P). به طور کلی، افزودن 2 و 4 میلیلیتر در دسیلیتر عصاره مرزنجوش به رقیقکننده منی باعث بهبود کیفیت منی بعد از انجماد-یخگشایی شد که ممکن است به دلیل افزایش فعالیت آنزیمهای آنتیاکسیدانی و کاهش پراکسیداسیون لیپیدی باشد.
Aebi H. (1984). Catalase in vitro. Method. Enzymol. 105, 121-126.
Aitken R.J., Fisher H.M., Fulton N., Gomez E., Knox W., Lewis B. and Irvine S. (1997). Reactive oxygen species generation by human spermatozoa is induced by exogenous NADPH and inhibited by the flavoprotein inhibitors diphenylene iodonium and quinacrine. Mol. Reprod. Dev. 47, 468-482.
Alvarez J.G. and Storey B.T. (1992). Evidence for increased lipid peroxidative damage and loss of superoxide dismutase activity as a mode of sublethal cryodamage to human sperm during cryopreservation. J. Anthol. 13, 232-241.
Amini M.R., Kohram H., Zare-Shahaneh A., Zhandi M., Sharideh H. and Nabi M.M. (2015). The effects of different levels of catalase and superoxide dismutase in modified Beltsville extender on rooster post-thawed sperm quality. Cryobiology. 70, 226-232.
Armstrong J.S., Rajasekaran M., Chamulitrat W., Gatti P., Hellstrom W.J. and SikkaS.C. (1999). Characterization of reactive oxygen species induced effects on human spermatozoa movement and energy metabolism. Free Radic. Biol. Med. 26, 869-880.
Arora A., Byrem T.M., Nair M.G. and Strasburg G.M. (2000). Modulation of liposomal membrane fluidity by flavonoids and isoflavonoids. Arch. Biochem. Biophys. 373, 102-109.
AshrafiI., Kohram H. and Ardabili F.F. (2013). Anti-oxidative effects of melatonin on kinetics, microscopic and oxidative parameters of cryopreserved bull spermatozoa. Anim. Reprod. Sci. 139, 25-30.
AshrafiI., Kohram H., Naijian H., Bahreini M. and Mirzakhani H. (2011). Effect of controlled and uncontrolled cooling rate on motility parameters of cryopreserved ram spermatozoa. Afican J. Biol. 10, 8965-8969.
Aurich J., Schönherr U., Hoppe H. and Aurich C. (1997). Effects of antioxidants on motility and membrane integrity of chilled-stored stallion semen. Theriogenology. 48, 185-192.
Barbas J.P. and Mascarenhas R.D. (2009). Cryopreservation of domestic animal sperm cells. Cell. Tissue. Bank. 10, 49-62.
Bilodeau J.F., Blanchette S., Gagnon C. and Sirard M.A. (2001). Thiols prevent H2O 2-mediated loss of sperm motility in cryopreserved bull semen. Theriogenology. 56, 275-286.
Bilodeau J.F., Chatterjee S., Sirard M.A. and Gagnon C. (2000). Levels of antioxidant defenses are decreased in bovine spermatozoa after a cycle of freezing and thawing. Mol. Reprod. Dev. 55, 282-288.
Blokhina O., Virolainen E. and Fagerstedt K.V. (2003). Antioxidants, oxidative damage and oxygen deprivation stress: a review. Ann. Bot. 91, 179-194.
Buyukleblebici S., Tuncer P.B., Bucak M.N., Tasdemir U., Eken A., Buyukleblebici O., Durmaz E., Sariozkan S. and Endirlik B.U. (2014). Comparing ethylene glycol with glycerol and with or without dithiothreitol and sucrose for cryopreservation of bull semen in egg-yolk containing extenders. Cryobiology. 69, 74-78.
Cervato G., Carabelli M., Gervasio S., Cittera A., Cazzola R. and Cestaro B. (2000). Antioxidant properties of oregano (Origanum vulgare) leaf extracts. J. Food Biochem. 24, 453-465.
Cocchia N., Pasolini M., Mancini R., Petrazzuolo O., Cristofaro I., Rosapane I., Sica A., Tortora G., Lorizio R. and Paraggio G. (2011). Effect of sod (superoxide dismutase) protein supplementation in semen extenders on motility, viability, acrosome status and ERK (extracellular signal-regulated kinase) protein phosphorylation of chilled stallion spermatozoa. Theriogenology. 75, 1201-1210.
Daghigh-Kia H., Olfati-Karaji R., Hoseinkhani A. and AshrafiI. (2014). Effect of rosemary (Rosmarinus officinalis) extracts and glutathione antioxidants on bull semen quality after cryopreservation. Spanish J. Agric. Res. 12, 98-105.
Dehghan G., Shafiee A., Ghahremani M.H., Ardestani S.K. and Abdollahi M. (2007). Antioxidant potential of various extracts from Ferula szovitsiana. in relation to their phenolic content. Pharm. Biol. 45, 691-699.
Guthrie H. and Welch G. (2012). Effects of reactive oxygen species on sperm function. Theriogenology. 78, 1700-1708.
Heo H.J., Cho H.Y., Hong B., Kim H.K., Heo T.R., Kim E.K., Kim S.K., Kim C.J. and Shin D.H. (2002). Ursolic acid of Origanum majorana reduces amyloid beta-induced oxidative injury. Mol. cells. 13, 5-11.
Holt W. (1996). Alternative strategies for the long-term preservation of spermatozoa. Reprod. Fertil. Dev. 9, 309-319.
Isachenko E., Isachenko V., Katkov I.I., Dessole S. and Nawroth F. (2003). Vitrification of mammalian spermatozoa in the absence of cryoprotectants: from past practical difficulties to present success. Reprod. Biomed. Online. 6, 191-200.
Kikuzaki H. and Nakatani N. (1989). Structure of a new anti-oxidative phenolic acid from oregano (Origanum vulgare). Agric. Biol. Chem. 53, 519-524.
Kulišić T., Kriško A., Dragović-Uzelac V., Miloš M. and Pifat G. (2007). The effects of essential oils and aqueous tea infusions of oregano (Origanum vulgare), thyme (Thymus vulgaris) and wild thyme (Thymus serpyllum) on the copper-induced oxidation of human low-density lipoproteins. Int. J. Food Sci. Nutr. 58, 87-93.
Lasso J.L., Noiles E.E., Alvarez J.G. and Storey B.T. (1994). Mechanism of superoxide dismutase loss from human sperm cells during cryopreservation. J. Androl. 15, 255-255.
Mahmood S.A. and Ijaz A. (2006). Effect of cold shock on frozen-thawed spermatozoa of buffalo and cow. Pp. 701-708 in Proc. 5th Asian Buffalo. Congr. Naning, China.
Malo C., Gil L., Cano R., Martínez F. and Galé I. (2011). Antioxidant effect of rosemary (Rosmarinus officinalis) on boar epididymal spermatozoa during cryopreservation. Theriogenology. 75, 1735-1741.
Malo C., Gil L., Gonzalez N., Martinez F., Cano R., De Blas I. and Espinosa E. (2010). Anti-oxidant supplementation improves boar sperm characteristics and fertility after cryopreservation: comparison between cysteine and rosemary (Rosmarinus officinalis). Cryobiology. 61, 142-147.
Matsuura H., Chiji H., Asakawa C., Amano M., Yoshihara T. and Mizutani J. (2003). DPPH radical scavengers from dried leaves of oregano (Origanum vulgare). Biosci. Biotechnol. Biochem. 67, 2311-2316.
Michalak A. (2006). Phenolic compounds and their antioxidant activity in plants growing under heavy metal stress. Polish J. Environ. Stud. 15, 523-601.
Najafi A., Daghigh Kia H., Mohammadi H., Najafi M.H., Zanganeh Z., Sharafi M., Martinez-Pastor F. and Adeldust H. (2014). Different concentrations of cysteamine and ergothioneine improve microscopic and oxidative parameters in ram semen frozen with a soybean lecithin extender. Cryobiology. 69, 68-73.
Osawa T. (1994). Novel natural antioxidants for utilization in food and biological systems. Pp. 241-251 in Postharvest Biochemistry of Plant Food Materials in the Tropics. I. Uritani, V.V. Garcia and E.M. Mendoza, Eds. Tokyo, Japan.
Placer Z.A., Cushman L.L. and Johnson B.C. (1966). Estimation of product of lipid peroxidation (malonyl dialdehyde) in biochemical systems. Anal. Biochem. 16, 359-364.
Purdy P. (2006). A review on goat sperm cryopreservation. Small Rumin. Res. 63, 215-225.
Revell S. and Mrode R. (1994). An osmotic resistance test for bovine semen. Anim. Reprod. Sci. 36, 77-86.
Roca J., Gil M.A., Hernandez M., ParrillaI., Vazquez J.M. and Martinez E.A. (2004). Survival and fertility of boar spermatozoa after freeze thawing in extender supplemented with butylated hydroxytoluene. J. Androl. 25, 397-405.
Sanocka D. and Kurpisz M. (2004). Reactive oxygen species and sperm cells. Reprod. Biol. Endocrinol. 2, 1-7.
Sariozkan S., Bucak M.N., Tuncer P.B., Buyukleblebici S., Eken A. and Akay C. (2015). Influence of fetuin and hyaluronan on the post-thaw quality and fertilizing ability of Holstein bull semen. Cryobiology. 71, 119-124.
SAS Institute. (2001). SAS®/STAT Software, Release 9.1. SAS Institute, Inc., Cary, NC. USA.
SikkaS.C. (1996). Oxidative stress and role of antioxidants in normal and abnormal sperm function. Front. Biosci. 1, 78-86.
Ukeda H., Maeda S., Ishii T. and Sawamura M. (1997). Spectrophotometric assay for superoxide dismutase based on tetrazolium salt 3′-{1-[(phenylamino)-carbonyl]-3, 4-tetrazolium}-bis (4-methoxy-6-nitro) benzenesulfonic acid hydrate reduction by xanthine-xanthine oxidase. Anal. Biochem. 251, 206-209.
Vekiari S., Oreopoulou V., Tzia C. and Thomopoulos C. (1993). Oregano flavonoids as lipid antioxidants. J. Am. Oil Chem. Soc. 70, 483-487.
Wang Y., Sharma R. and Agarwal A. (1997). Effect of cryopreservation and sperm concentration on lipid peroxidation in human semen. Urology. 50, 409-413.
Wishart G. (1982). Maintenance of ATP concentrations in and of fertilizing ability of fowl and turkey spermatozoa in vitro. J. Reprod. Fertil. 66, 457-462.
Yanishlieva N.V. and Marinova E.M. (1996). Anti-oxidative effectiveness of some natural antioxidants in sunflower oil. Z. Lebensm. Unters. For. 203, 220-223.
Zanganeh Z., Zhandi M., Zare-Shahneh A., Najafi A., Nabi M.M. and Mohammadi-Sangcheshmeh A. (2013). Does rosemary aqueous extract improve buck semen cryopreservation? Small Rumin. Res. 114, 120-125.
Zhao H.W., Li Q.W., Ning G.Z., Han Z.S., Jiang Z.L. and Duan Y.F. (2009). Rhodiola sacra aqueous extract (RSAE) improves biochemical and sperm characteristics in cryopreserved boar semen. Theriogenology. 71, 849-857.
Zini A., Lamirande E. and Gagnon C. (1993). Reactive oxygen species in semen of infertile patients: levels of superoxide dismutase and catalase like activities in seminal plasma and spermatozoa. Int. J. Androl. 16, 183-188.
