Year 2025,
Volume: 9 Issue: 2, 123 - 130, 30.10.2025
Ahmet Eser
,
Kemal Bağcı
,
Abdurrahman Alakuş
,
Aslıhan Çakır Cihangiroğlu
,
Selin Yağcioğlu
,
Ramazan Arıcı
,
Kamber Demir
References
-
Abramowitz JS. The psychological treatment of obsessive- Ahmed A. Cryopreservation of stallion semen: a review. Zagazig Vet J. 2023; 51(3):263-278.
-
Agustinus A, Pakpahan C. Computer assisted sperm analysis: a review. IABJ. 2020; 1(2):60-66.
-
Akhtar MF, Ma Q, Li Y, et al. Effect of sperm cryopreservation in farm animals using nanotechnology. Animals. 2022; 12(17): 2277.
-
Akhter S, Ansari MS, Rakha BA, et al. In vitro evaluation of liquid‐stored buffalo semen at 5ºC diluted in soya lecithin based extender (Bioxcell®), tris‐Citric egg yolk, skim milk and egg yolk‐citrate extenders. Reprod Domest Anim. 2011; 46(1):45-49.
-
Alamo A, Luca C, Mongioì L, et al. Mitochondrial membrane potential predicts 4-hour sperm motility. Biomedicines. 2020; 8(7):196.
-
Alcay S, Aktar A, Koca D, Kilic MA, Akkasoglu M, Sagirkaya H. Positive effect of autologous platelet rich plasma on Saanen buck semen cryopreservation in non-breeding season. Cryobiology. 2021; 103:45-48.
-
Alcay S, Gokce E, Toker MB, et al. Freeze-dried egg yolk based extenders containing various antioxidants improve post-thawing quality and incubation resilience of goat spermatozoa. Cryobiology. 2016; 72(3):269-273.
-
Almeida Filho RAD, Silva LSD, Lima LGFD, et al. Egg-yolk- and a liposome- based extender: refrigeration time and effects on ram semen quality. Anim Reprod. 2025; 22(2):e20240052.
-
Ansari MS, Rakha BA, Akhter S. Cryopreservation of bull semen in OptiXcell® and conventional extenders: Comparison of semen quality and fertility. Anim Sci Pap Rep. 2017; 35(3):317-328.
-
Bathgate R, Maxwell WM, Evans G. Studies on the effect of supplementing boar semen cryopreservation media with different avian egg yolk types on in vitro post-thaw sperm quality. Reprod Domest Anim. 2006; 41(1):68-73.
-
Bousseau S, Brillard JP, Marguant-Le Guienne B, Guerin B, Camus A, Lechat M. Comparison of bacteriological qualities of various egg yolk sources and the in vitro and in vivo fertilizing potential of bovine semen frozen in egg yolk or lecithin based diluents. Theriogenology. 1998; 50(5):699-706.
-
Câmara D, Silva S, Almeida F, Nunes J, Guerra M. Effects of antioxidants and duration of pre-freezing equilibration on frozen-thawed ram semen. Theriogenology. 2011; 76(2):342-350.
-
Dariush G, Gholamhossein R, Rouhollah F, et al. The application of ultrasonic vibration in human sperm cryopreservation as a novel method for the modification of physicochemical characteristics of freezing media. Sci Rep. 2019; 9(1):10066.
-
Eser A, Yağcıoğlu S, Arıcı R, Demir K, Ak K. Effects of resveratrol-loaded cyclodextrin on the quality characteristics of ram spermatozoa following cryopreservation. Animals. 2024; 14(18):2745.
-
Gillan L, Evans G, Maxwell W. Flow cytometric evaluation of sperm parameters in relation to fertility potential. Theriogenology. 2005; 63(2):445-457.
-
Graham JK, Foote RH. Effect of several lipids, fatty acyl chain length and degree of unsaturation on the motility of bull spermatozoa after cold shock and freezing. Cryobiology. 1987; 24(1):42-45.
-
Intawicha P, Moonmanee T, Saengwong S, Lumsangkul C, Liao Y, Inyawilert W. The replacement of fresh egg yolk by lyophilized egg yolk in tris‐base extender in cryopreserved boer and saanen semen. Reprod Domest Anim. 2022; 57(8):928-935.
-
Karaşör ÖF, Bucak MN, Cenariu M, Bodu M, Taşpınar M, Taşpınar F. The effects of different doses of ROCK inhibitor, antifreeze protein III, and boron added to semen extender on semen freezeability of Ankara bucks. Molecules. 2022; 27(22):8070.
-
Kumar P, Saini M, Kumar D, et al. Liposome-based semen extender is suitable alternative to egg yolk-based extender for cryopreservation of buffalo (Bubalus bubalis) semen. Anim Reprod Sci. 2015; 159:38-45.
-
Layek SS, Mohanty TK, Kumaresan A, Parks JE. Cryopreservation of bull semen: Evolution from egg yolk based to soybean based extenders. Anim Reprod Sci. 2016; 172:1-9.
-
Leboeuf B, Restall B, Salamon S. Production and storage of goat semen for artificial insemination. Anim Reprod Sci. 2000; 62(1-3):113-141.
-
Lipar JL, Ketterson ED, Nolan V. Intra- clutch variation in testosterone content of red- winged blackbird eggs. Auk. 1999; 116(1):231-235.
-
Longobardi V, Zullo G, Cotticelli A, et al. Crocin improves the quality of cryopreserved goat semen in different breeds. Animals. 2020; 10(6):1101.
-
Manjunath P. New insights into the understanding of the mechanism of sperm protection by extender components. Anim Reprod. 2012; 9(4):809-815.
-
Marco-Jiménez F, Puchades S, Gadea J, Vicente J, Viudes-de-Castro M. Effect of semen collection method on pre-and post-thaw Guirra ram spermatozoa. Theriogenology. 2005; 64(8):1756-1765.
-
Miguel-Jimenez, S., Del Alamo, M. M. R., Álvarez-Rodríguez, et al. In vitro assessment of egg yolk-, soya bean lecithin-and liposome-based extenders for cryopreservation of dairy bull semen. Anim Reprod Sci. 2020; 215:106315.
-
Moussa M, Martinet V, Trimeche A, Tainturier D, Anton M. Low density lipoproteins extracted from hen egg yolk by an easy method: cryoprotective effect on frozen- thawed bull semen. Theriogenology. 2002; 57(6):1695-1706.
-
Narwade BM, Mohanty TK, Bhakat M, Rahim A. Goat semen cryopreservation using egg yolk and soya based extenders containing trehalose. Indian J Anim Sci. 2017; 87(7):851-855.
-
Naz S, Umair M, Iqbal S. Comparison of Tris egg yolk‐based, Triladyl® and Optixell® extender on post‐thaw quality, kinematics and in vivo fertility of Nili Ravi buffalo (Bubalus bubalis) bull spermatozoa. Andrologia. 2018; 50(8):e13063.
-
Pace MM, Graham EF. Components in egg yolk which protect bovine spermatozoa during freezing. J Anim Sci. 1974; 39(6):1144-1149.
-
Paoli D, Gallo M, Rizzo F, et al. Mitochondrial membrane potential profile and its correlation with increasing sperm motility. Fertil Steril. 2021; 95(7):2315-2319.
-
Perumal P, De A, Ramesh V, et al. Effect of superoxide dismutase on cryopreservation of Mithun semen. Indian J Anim Sci. 2022; 92(6):712-724.
-
Posevkin D, Khussein A. Tracking the swimming of self-propelling spermatozoa using computer assisted semen analysis techniquesto determine the effect of vibration on spermatozoids. Interact Sci. 2023; 2 (78):66-73.
-
Rather H, Islam R, Malik A, Lone F, Banday M. Effect of different extenders for preservation of ram semen at 4°C. Indian J Anim Res. 2017; 51(6):982-987.
-
Ropke T, Oldenhof H, Leiding C, Sieme H, Bollwein H, Wolkers WF. Liposomes for cryopreservation of bovine sperm. Theriogenology. 2011; 76(8):1465-1472.
-
Shin SJ, Lein DH, Patten VH, Ruhnke HL. A new antibiotic combination for frozen bovine semen 1. Control of mycoplasmas, ureaplasmas, Campylobacter fetus subsp. venerealis and Haemophilus somnus. Theriogenology. 1988; 29(3):577-591.
-
Stewart JL, Shipley CF, Katich AS, et al. Cryopreservation of white-tailed deer (Odocoileus virginianus) semen using soybean-, liposome-, and egg yolk-based extenders. Anim Reprod Sci. 2016; 171:7-16.
-
Sun W, Jiang S, Su J, et al. The effects of cryopreservation on the acrosome structure, enzyme activity, motility, and fertility of bovine, ovine, and goat sperm. Anim Reprod. 2020; 17(4):e20200219.
-
Swelum AA, Saadeldin IM, Ba-Awadh H, et al. Efficiency of commercial egg yolk-free and egg yolk-supplemented tris-based extenders for dromedary camel semen cryopreservation. Animals. 2019; 9(11):999.
-
Tang S, Lu J, Xu Y, et al. Analysis of selected sperm samples by a computer‐assisted system with high frame rate: a prospective study. Health Sci Rep. 2023; 6(5):e1217.
-
Tekin K, Daşkın A. Effect of different extenders on motility and some sperm kinematics parameters in Norduz goat semen. Turk J Vet Anim Sci. 2016; 40(4):490-495.
-
Thurston LM, Watson PF. Semen cryopreservation: a genetic explanation for species and individual variation?. CryoLetters. 2002; 23(4):255-262.
-
Van Wagtendonk-de Leeuw AM, Haring RM, Kaal-Lansbergen LMTE, Den Daas JHG. Fertility results using bovine semen cryopreserved with extenders based on egg yolk and soybean extract. Theriogenology. 2000; 54:57-67.
-
Watson PM. The protection of ram and bull spermatozoa by low density lipoprotein fraction of egg yolk during storage at 5°C and deep freezing. J of Therm Biol. 1976; 1:137-141.
-
Whaley D, Damyar K, Witek R, et al. Cryopreservation: an overview of principles and cell-specific considerations. Cell Transplant. 2021; 30:1-12.
-
Wilhelm KM, Graham JK, Squires EL. Effects of phosphatidylserine and cholesterol liposomes on the viability, motility and acrosomal integrity of stallion spermatozoa prior to and after cryopreservation. Cryobiology. 1996; 33:320-329.
Effects of tris–egg yolk and OptiXcell® extenders on post-thaw spermatological parameters in goat semen
Year 2025,
Volume: 9 Issue: 2, 123 - 130, 30.10.2025
Ahmet Eser
,
Kemal Bağcı
,
Abdurrahman Alakuş
,
Aslıhan Çakır Cihangiroğlu
,
Selin Yağcioğlu
,
Ramazan Arıcı
,
Kamber Demir
Abstract
Objective: The objective of this study was to comparatively evaluate the post-thaw sperm quality of Boer and Anglo-Nubian goats using semen samples in which seminal plasma was removed by centrifugation and subsequently diluted with a Tris-based egg yolk extender, or directly diluted with the commercial liposome-based extender OptiXcell® without prior centrifugation.
Materials and Methods: Semen samples were collected from five adult Boer and five adult Anglo-Nubian goats using an artificial vagina, with collections performed five times per week. The collected semen samples were divided into two equal portions. The first portion was centrifuged to remove seminal plasma prior to dilution with a Tris–egg yolk (TEY) extender (Boer-TEY and Anglo-Nubian-TEY), whereas the remaining portion was directly diluted with OptiXcell® (Boer- OptiXcell and Anglo-Nubian- OptiXcell). All samples were frozen, and post-thaw evaluations were performed. Sperm motility parameters were assessed using computer-assisted sperm analysis (CASA), while plasma membrane integrity, acrosome integrity, mitochondrial activity, viability, and reactive oxygen species (ROS) levels were evaluated by flow cytometry.
Results: Post-thaw CASA analysis revealed that total motility (MOT, %) was significantly higher in Boer goat spermatozoa diluted with the TEY extender compared to those diluted with OptiXcell® (p<0.05). However, MOT values in the Boer–TEY group did not differ from those in the Anglo-Nubian TEY or Anglo-Nubian OptiXcell® groups (p>0.05). Progressive motility (%) and all velocity-related kinematic parameters (VAP, VSL, VCL, ALH, BCF, STR, LIN, and WOB) showed no significant differences among groups (p>0.05). Flow cytometric assessments indicated
no significant differences in plasma membrane integrity (%), acrosome integrity (%), high mitochondrial activity (%), or viability (%) between groups (p>0.05). In contrast, the proportion of spermatozoa with intracellular reactive oxygen species (ROS) was significantly lower in Boer goat spermatozoa diluted with the TEY extender (p<0.05)
Conclusion: Breed variation should be considered when selecting extenders for goat semen cryopreservation. In Boer goats, the Tris–egg yolk extender was found to be more effective for semen cryopreservation compared to the OptiXcell® extender.
Ethical Statement
This study was approved by the Animal Research Local Ethics Committee at the Siirt University with decision number 30.04.2025-2025/17.
Supporting Institution
No financial support was provided for this study.
Thanks
The authors would like to acknowledge the invaluable contributions of the staff at the Siirt University Reproductive Biotechnology Center.
References
-
Abramowitz JS. The psychological treatment of obsessive- Ahmed A. Cryopreservation of stallion semen: a review. Zagazig Vet J. 2023; 51(3):263-278.
-
Agustinus A, Pakpahan C. Computer assisted sperm analysis: a review. IABJ. 2020; 1(2):60-66.
-
Akhtar MF, Ma Q, Li Y, et al. Effect of sperm cryopreservation in farm animals using nanotechnology. Animals. 2022; 12(17): 2277.
-
Akhter S, Ansari MS, Rakha BA, et al. In vitro evaluation of liquid‐stored buffalo semen at 5ºC diluted in soya lecithin based extender (Bioxcell®), tris‐Citric egg yolk, skim milk and egg yolk‐citrate extenders. Reprod Domest Anim. 2011; 46(1):45-49.
-
Alamo A, Luca C, Mongioì L, et al. Mitochondrial membrane potential predicts 4-hour sperm motility. Biomedicines. 2020; 8(7):196.
-
Alcay S, Aktar A, Koca D, Kilic MA, Akkasoglu M, Sagirkaya H. Positive effect of autologous platelet rich plasma on Saanen buck semen cryopreservation in non-breeding season. Cryobiology. 2021; 103:45-48.
-
Alcay S, Gokce E, Toker MB, et al. Freeze-dried egg yolk based extenders containing various antioxidants improve post-thawing quality and incubation resilience of goat spermatozoa. Cryobiology. 2016; 72(3):269-273.
-
Almeida Filho RAD, Silva LSD, Lima LGFD, et al. Egg-yolk- and a liposome- based extender: refrigeration time and effects on ram semen quality. Anim Reprod. 2025; 22(2):e20240052.
-
Ansari MS, Rakha BA, Akhter S. Cryopreservation of bull semen in OptiXcell® and conventional extenders: Comparison of semen quality and fertility. Anim Sci Pap Rep. 2017; 35(3):317-328.
-
Bathgate R, Maxwell WM, Evans G. Studies on the effect of supplementing boar semen cryopreservation media with different avian egg yolk types on in vitro post-thaw sperm quality. Reprod Domest Anim. 2006; 41(1):68-73.
-
Bousseau S, Brillard JP, Marguant-Le Guienne B, Guerin B, Camus A, Lechat M. Comparison of bacteriological qualities of various egg yolk sources and the in vitro and in vivo fertilizing potential of bovine semen frozen in egg yolk or lecithin based diluents. Theriogenology. 1998; 50(5):699-706.
-
Câmara D, Silva S, Almeida F, Nunes J, Guerra M. Effects of antioxidants and duration of pre-freezing equilibration on frozen-thawed ram semen. Theriogenology. 2011; 76(2):342-350.
-
Dariush G, Gholamhossein R, Rouhollah F, et al. The application of ultrasonic vibration in human sperm cryopreservation as a novel method for the modification of physicochemical characteristics of freezing media. Sci Rep. 2019; 9(1):10066.
-
Eser A, Yağcıoğlu S, Arıcı R, Demir K, Ak K. Effects of resveratrol-loaded cyclodextrin on the quality characteristics of ram spermatozoa following cryopreservation. Animals. 2024; 14(18):2745.
-
Gillan L, Evans G, Maxwell W. Flow cytometric evaluation of sperm parameters in relation to fertility potential. Theriogenology. 2005; 63(2):445-457.
-
Graham JK, Foote RH. Effect of several lipids, fatty acyl chain length and degree of unsaturation on the motility of bull spermatozoa after cold shock and freezing. Cryobiology. 1987; 24(1):42-45.
-
Intawicha P, Moonmanee T, Saengwong S, Lumsangkul C, Liao Y, Inyawilert W. The replacement of fresh egg yolk by lyophilized egg yolk in tris‐base extender in cryopreserved boer and saanen semen. Reprod Domest Anim. 2022; 57(8):928-935.
-
Karaşör ÖF, Bucak MN, Cenariu M, Bodu M, Taşpınar M, Taşpınar F. The effects of different doses of ROCK inhibitor, antifreeze protein III, and boron added to semen extender on semen freezeability of Ankara bucks. Molecules. 2022; 27(22):8070.
-
Kumar P, Saini M, Kumar D, et al. Liposome-based semen extender is suitable alternative to egg yolk-based extender for cryopreservation of buffalo (Bubalus bubalis) semen. Anim Reprod Sci. 2015; 159:38-45.
-
Layek SS, Mohanty TK, Kumaresan A, Parks JE. Cryopreservation of bull semen: Evolution from egg yolk based to soybean based extenders. Anim Reprod Sci. 2016; 172:1-9.
-
Leboeuf B, Restall B, Salamon S. Production and storage of goat semen for artificial insemination. Anim Reprod Sci. 2000; 62(1-3):113-141.
-
Lipar JL, Ketterson ED, Nolan V. Intra- clutch variation in testosterone content of red- winged blackbird eggs. Auk. 1999; 116(1):231-235.
-
Longobardi V, Zullo G, Cotticelli A, et al. Crocin improves the quality of cryopreserved goat semen in different breeds. Animals. 2020; 10(6):1101.
-
Manjunath P. New insights into the understanding of the mechanism of sperm protection by extender components. Anim Reprod. 2012; 9(4):809-815.
-
Marco-Jiménez F, Puchades S, Gadea J, Vicente J, Viudes-de-Castro M. Effect of semen collection method on pre-and post-thaw Guirra ram spermatozoa. Theriogenology. 2005; 64(8):1756-1765.
-
Miguel-Jimenez, S., Del Alamo, M. M. R., Álvarez-Rodríguez, et al. In vitro assessment of egg yolk-, soya bean lecithin-and liposome-based extenders for cryopreservation of dairy bull semen. Anim Reprod Sci. 2020; 215:106315.
-
Moussa M, Martinet V, Trimeche A, Tainturier D, Anton M. Low density lipoproteins extracted from hen egg yolk by an easy method: cryoprotective effect on frozen- thawed bull semen. Theriogenology. 2002; 57(6):1695-1706.
-
Narwade BM, Mohanty TK, Bhakat M, Rahim A. Goat semen cryopreservation using egg yolk and soya based extenders containing trehalose. Indian J Anim Sci. 2017; 87(7):851-855.
-
Naz S, Umair M, Iqbal S. Comparison of Tris egg yolk‐based, Triladyl® and Optixell® extender on post‐thaw quality, kinematics and in vivo fertility of Nili Ravi buffalo (Bubalus bubalis) bull spermatozoa. Andrologia. 2018; 50(8):e13063.
-
Pace MM, Graham EF. Components in egg yolk which protect bovine spermatozoa during freezing. J Anim Sci. 1974; 39(6):1144-1149.
-
Paoli D, Gallo M, Rizzo F, et al. Mitochondrial membrane potential profile and its correlation with increasing sperm motility. Fertil Steril. 2021; 95(7):2315-2319.
-
Perumal P, De A, Ramesh V, et al. Effect of superoxide dismutase on cryopreservation of Mithun semen. Indian J Anim Sci. 2022; 92(6):712-724.
-
Posevkin D, Khussein A. Tracking the swimming of self-propelling spermatozoa using computer assisted semen analysis techniquesto determine the effect of vibration on spermatozoids. Interact Sci. 2023; 2 (78):66-73.
-
Rather H, Islam R, Malik A, Lone F, Banday M. Effect of different extenders for preservation of ram semen at 4°C. Indian J Anim Res. 2017; 51(6):982-987.
-
Ropke T, Oldenhof H, Leiding C, Sieme H, Bollwein H, Wolkers WF. Liposomes for cryopreservation of bovine sperm. Theriogenology. 2011; 76(8):1465-1472.
-
Shin SJ, Lein DH, Patten VH, Ruhnke HL. A new antibiotic combination for frozen bovine semen 1. Control of mycoplasmas, ureaplasmas, Campylobacter fetus subsp. venerealis and Haemophilus somnus. Theriogenology. 1988; 29(3):577-591.
-
Stewart JL, Shipley CF, Katich AS, et al. Cryopreservation of white-tailed deer (Odocoileus virginianus) semen using soybean-, liposome-, and egg yolk-based extenders. Anim Reprod Sci. 2016; 171:7-16.
-
Sun W, Jiang S, Su J, et al. The effects of cryopreservation on the acrosome structure, enzyme activity, motility, and fertility of bovine, ovine, and goat sperm. Anim Reprod. 2020; 17(4):e20200219.
-
Swelum AA, Saadeldin IM, Ba-Awadh H, et al. Efficiency of commercial egg yolk-free and egg yolk-supplemented tris-based extenders for dromedary camel semen cryopreservation. Animals. 2019; 9(11):999.
-
Tang S, Lu J, Xu Y, et al. Analysis of selected sperm samples by a computer‐assisted system with high frame rate: a prospective study. Health Sci Rep. 2023; 6(5):e1217.
-
Tekin K, Daşkın A. Effect of different extenders on motility and some sperm kinematics parameters in Norduz goat semen. Turk J Vet Anim Sci. 2016; 40(4):490-495.
-
Thurston LM, Watson PF. Semen cryopreservation: a genetic explanation for species and individual variation?. CryoLetters. 2002; 23(4):255-262.
-
Van Wagtendonk-de Leeuw AM, Haring RM, Kaal-Lansbergen LMTE, Den Daas JHG. Fertility results using bovine semen cryopreserved with extenders based on egg yolk and soybean extract. Theriogenology. 2000; 54:57-67.
-
Watson PM. The protection of ram and bull spermatozoa by low density lipoprotein fraction of egg yolk during storage at 5°C and deep freezing. J of Therm Biol. 1976; 1:137-141.
-
Whaley D, Damyar K, Witek R, et al. Cryopreservation: an overview of principles and cell-specific considerations. Cell Transplant. 2021; 30:1-12.
-
Wilhelm KM, Graham JK, Squires EL. Effects of phosphatidylserine and cholesterol liposomes on the viability, motility and acrosomal integrity of stallion spermatozoa prior to and after cryopreservation. Cryobiology. 1996; 33:320-329.