The optimum concentration of soya-lecithin for replacing egg yolk in INRA-82-modified semen extenders improved the cryopreserved semen quality and DNA integrity of Arabian stallions

Amal Aboelmaaty; Islam El Sayed El-Seadawy; Hazem Ahmad Aldebakey; Heba F. Hozyen; Diya A. El-Badry; Abeer M. Anwer; Tsvetan Tsvetkov; Teodora Daneva

doi:10.31893/avr.2023019

Keywords:

modified INRA-82

postthaw DNA

semen cryopreservation

soy lecithin

stallion

Abstract

The cryopreservation of stallion spermatozoa varies greatly between animals. Different equine semen extender supplements were invented to maximize the postthaw semen quality. Soy lecithin is one of those additives that contain plant origin phospholipids used to replace egg yolk to minimize disease transmission. This study aimed to determine which concentrations of soy lecithin can replace egg-yolk in modified INRA-82 semen extenders for the cryopreservation of horse semen. Semen was collected from six Arabian stallions and extended with modified INRA-82 semen extender supplemented with 0.0, 1.25, 2.50, and 5.0% soya lecithin. Frozen-thawed sperm motility was measured at 0, 1, 2, and 3 h. Viability index, sperm plasma membrane integrity (SPMI), and acrosome integrity in addition to the comet assay nonfragmented DNA, head DNA, tail DNA, tail length, and tail moment were statistically analyzed. The addition of 2.5% soya lecithin had the highest postthaw sperm motility at 0, 1, 2 h (P < 0.001) and 3 h (P < 0.0001), the optimum viability index (P < 0.0001), SPMI (P < 0.05), acrosome integrity (P < 0.0001), nonfragmented DNA (P < 0.01), DNA in the comet head (P < 0.05), minimum DNA in the comet tail (P < 0.05), shortest comet tail length (P < 0.0001), and lowest comet tail moment (P < 0.01). Soy lecithin at concentrations other than 2.5% deteriorated all postthaw semen parameters. Soy lecithin (2.5%) can replace egg yolk in the modified INRA-82 semen extender for the cryopreservation of good-quality stallion semen.
References
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2. Akhter S, Ansari M, Rakha B, Andrabi S, Iqbal S, Ullah N (2010) Cryopreservation of buffalo (Bubalus bubalis) semen in Bioxcell® extender. Theriogenology 74:951-955.
3. Akhter S, Ansari MS, Rakha BA, Ullah N, Andrabi SM, Khalid M (2011) In vitro evaluation of liquid-stored buffalo semen at 5°C diluted in soya lecithin based extender (Bioxcell®), tris-citric egg yolk, skim milk and egg yolk-citrate extenders. Reprod. Domest. Anim. 46:45-9. doi: 10.1111/j.1439-0531.2009.01561.x. 34
4. Arifiantini RII, Yusuf STL (2010) Developing of tris soy milk diluent for Frisian Holstein bull frozen semen. Hayati J. Biol.17: 91-94
5. Axnér E, Lagerson E (2016) Cryopreservation of Dog Semen in a Tris Extender with 1% or 2% Soya Bean Lecithin as a Replacement of Egg Yolk. Reprod. Domest. Anim. 51:262-8.
6. Bousseau S, Brillard JP, Marguant-Le Guienne B, Guérin B, Camus A, Lechat M (1998) 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, 50: 699-706
7. Cochran JD, Amann RP, Froman DP, Pickett BW (1984) Effects of centrifugation, glycerol level, cooling to 5C, freezing rate and thawing rate on the postthaw motility of equine sperm. Theriogenology 22:25-38.
8. Cristanelli MJ, Amann RP, Squires EL, Pickett BW (1985) Effects of egg yolk and glycerol level in lactose-EDTA-egg yolk extender and of freezing rate on the motility of frozen-thawed stallion spermatozoa. Theriogenology 23:25-38.
9. Crockett EC, Graham JK, Bruemmer JE, Squires EL (2001) Effect of cooling of equine spermatozoa before freezing on postthaw motility: preliminary results. Theriogenology 55:793-803.
10. Duan H, Zhang X, Deng L, Zeng S, Han G (2014) Soybean lecithin in stallion semen extender maintains frozen-thawed sperm membrane integrity. J. Equine Vet. Sci. 34: 101.https://doi.org/10.1016/j.jevs.2013.10.068
11. Duan H, Zhang X, Sun Y, Deng L, Han G (2013) Study on the Effect of Soybean Lecithin in the Cryopreservation of Stallion Semen. China Anim. Husbandry Vet. Med. 40:150-154.
12. El-Azzazi FE, Yaseen MA (2016) Soybean lecithin-based extender for cryopreservation of ram semen. Egyptian J. Anim. Prod. 53:43-48.
13. El-Badry DA, Abo El-Maaty AM, El Sisy GA (2017) The Effect of Trehalose Supplementation of INRA-82 Extender on Quality and Fertility of Cooled and Frozen-Thawed Stallion Spermatozoa. J. Equine Vet. Sci. 48: 86-92. DOI: 10.1016/j.jevs.2016.08.020.
14. EL-Badry DA, Anwer AM, Rawash ZM (2014) Effect of different concentrations of sodium dodecyl sulfate, egg yolk and glycerol on the freezability and DNA integrity of Arabian stallion spermatozoa. Assiut Vet. Med. J. 60:29-35.
15. Fathi M, Rawash Z, Dohreig R, Gamal I, Mohamed A, Abu-El Naga E, Badr M (2019) Soybean lecithin-based extender improves Damascus goat sperm cryopreservation and fertilizing potential following artificial insemination. Asian Pacific J. Reprod. 8: 174-180.
16. Fraser L (2004) Structural damage to nuclear DNA in mammalian spermatozoa: its evaluation techniques and relationship with male infertility. Pol. J. Vet. Sci. 7:311-21
17. Hinsch E, Hinsch KD, Boehm JG, Schill WB, Mueller-Schloesser F (1997) Functional parameters and fertilization success of Bovine semen cryopreserved in egg-yolk free and egg-yolk containing extenders. Reprod. Domest. Anim. 32: 143-149.
18. Hughes CM, Lewis SE, McKelvey-Martin V, Thompson WA (1996) Comparison of baseline and induced DNA damage in human spermatozoa from fertile and infertile men, using a modified comet assay. Mol. Hum. Reprod. 2:613-9
19. Khalifa EI, Abdel-Hafez MAM (2014) Effect of soybean lecithin-based semen extender on freezability and fertility of Rahmani ram spermatozoa. Egyptian J. Sheep Goat Sci. 9:59- 66.
20. Layek SS, Mohanty TK, Kumaresan A, Parks JE (2016) Cryopreservation of bull semen: Evolution from egg yolk based to soybean-based extenders. Anim. Reprod. Sci. 172:1-9. doi: 10.1016/j.anireprosci.2016.04.013.
21. Medica AJ, Aitken RJ, Nicolson GL, Sheridan AR, Swegen A, De Iuliis GN, Gibb Z (2021) Glycerophospholipids protect stallion spermatozoa from oxidative damage in vitro. Reprod Fertil. 2:199-209. doi: 10.1530/RAF-21-0028.
22. Mehdipour M, Daghigh Kia H, Najafi A, Vaseghi Dodaran H, García-Álvarez O (2016) Effect of green tea (Camellia sinensis) extract and pre-freezing equilibration time on the post-thawing quality of ram semen cryopreserved in a soybean lecithin-based extender. Cryobiology, 73:297-303. doi: 10.1016/j.cryobiol.2016.10.008.
23. Miguel-Jimenez S, Rivera Del Alamo MM, Álvarez-Rodríguez M, Hidalgo CO, Peña AI, Muiño R, Rodríguez-Gil JE, Mogas T (2020) In vitro assessment of egg yolk-, soya bean lecithin- and liposome-based extenders for cryopreservation of dairy bull semen. Anim. Reprod. Sci. 215:106315. doi: 10.1016/j.anireprosci.2020.106315.
24. Milovanov VK. Biology of reproduction and artificial insemination of farm animals. Mosc: Monogr Selkohz Lit J Plakatov; 1962.
25. Muiño R, Fernández M, Peña AI. Post-thaw survival and longevity of bull spermatozoa frozen with an egg yolk-based or two egg yolk-free extenders after an equilibration period of 18 h. Reprod. Domest. Anim. 2007; +3.42:305-11. doi: 10.1111/j.1439-0531.2006.00784.x.
26. Najafi A, Daghigh-Kia H, Dodaran HV, Mehdipour M, Alvarez-Rodriguez M (2017) Ethylene glycol, but not DMSO, could replace glycerol inclusion in soybean lecithin-based extenders in ram sperm cryopreservation. Anim. Reprod. Sci. 177:35-41. doi: 10.1016/j.anireprosci.2016.12.004.
27. Nie GJ, Wenzel JGW (2001) Adaptation of the hypo-osmotic swelling test to assess functional integrity of stallion spermatozoal plasma membranes. Theriogenology 55:1005-18.
28. Nishijima K, Kitajima S, Koshimoto C, Morimoto M, Watanabe T, Fan J, Matsuda Y (2015) Motility and fertility of rabbit sperm cryopreserved using soybean lecithin as an alternative to egg yolk. Theriogenology 84:1172-5. doi: 10.1016/j.theriogenology.2015.06.018.
29. Ondřej Š, Jiří Š, Jan B, Pavla MP, Lucie T, Doležalová M, Petra F, Luděk S, Radko R (2019) Low Density Lipoprotein - important player in increasing cryoprotective efficiency of soybean lecithin-based bull semen extenders. Anim. Reprod. 16:267-276. doi: 10.21451/1984-3143-AR2018-0107
30. Papa FO, Felício GB, Melo-Oña CM, Alvarenga MA, De Vita B, Trinque C, Puoli-Filho JN, Dell'Aqua JA Jr (2011) Replacing egg yolk with soybean lecithin in the cryopreservation of stallion semen. Anim. Reprod. Sci. 129:73-7. doi: 10.1016/j.anireprosci.2011.10.006.
31. Salmani H, Towhidi A, Zhandi M, Bahreini M, Sharafi M (2014) In vitro assessment of soybean lecithin and egg yolk-based diluents for cryopreservation of goat semen. Cryobiology, 68:276-80. doi: 10.1016/j.cryobiol.2014.02.008.
32. Singh AK, Kumar A, Honparkhe M, Kaur S, Kaur H, Ghuman S, Brar PS (2018) Comparison of in vitro and in vivo fertilizing potential of buffalo bull semen frozen in egg yolk-, soya bean lecithin- and liposome-based extenders. Reprod Domest Anim. 53:195-202. doi: 10.1111/rda.13092.
33. Singh AK, Singh VK, Narwade BM, Mohanty TK, Atreja SK (2012) Comparative quality assessment of buffalo (Bubalus bubalis) semen chilled (5°C) in egg yolk- and soya milk-based extenders. Reprod. Domest. Anim. 47:596-600. doi: 10.1111/j.1439-0531.2011.01928.x.
34. Speroni F, Anón MC, de Lamballerie M (2010) Effects of calcium and high pressure on soybean proteins: a calorimetric study. Food Res. Int. 43: 1347-1355.
35. Taladrid D, Marín D, Alemán A, Álvarez-Acero I, Montero P, Gómez-Guillén M (2017) Effect of chemical composition and sonication procedure on properties of food-grade soy lecithin liposomes with added glycerol. Food Res. Int. 100:541-550. doi: 10.1016/j.foodres.2017.07.052.
36. van Wagtendonk-de Leeuw AM, Haring RM, Kaal-Lansbergen LM, den Daas JH (2000) Fertility results using bovine semen cryopreserved with extenders based on egg yolk and soy bean extract. Theriogenology 54:57-67. doi: 10.1016/S0093-691X(00)00324-1.
37. Wells ME, Awa OA (1970) New technique for assessing acrosomal characteristics of spermatozoa. J. Dairy Sci. 53:327-32
38. Zhao JQ, Xiao GL, Zhu WL, Fang D, Li N, Han CM, Gao QH (2021) Ram semen preserved at 0°C with soybean lecithin Tris-based extender substituted for egg yolk. Anim. Biosci. 34:192-197. doi: 10.5713/ajas.20.0118.

This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

How to cite

Aboelmaaty, A., El-Seadawy, I. E. S., Aldebakey, H. A., Hozyen, H. F., El-Badry, D. A., Anwer, A. M., Tsvetkov, T., & Daneva, T. (2023). The optimum concentration of soya-lecithin for replacing egg yolk in INRA-82-modified semen extenders improved the cryopreserved semen quality and DNA integrity of Arabian stallions. Applied Veterinary Research, (| Accepted Articles), 2023019. https://doi.org/10.31893/avr.2023019

[1] Akhter S, Ansari M, Andrabi S, Rakha B, Ullah N, Khalid M (2012) Soya-lecithin in extender improves the freezability and fertility of buffalo (Bubalus bubalis) bull spermatozoa. Reprod. Domest. Anim. 47:815-819.

[2] Akhter S, Ansari M, Rakha B, Andrabi S, Iqbal S, Ullah N (2010) Cryopreservation of buffalo (Bubalus bubalis) semen in Bioxcell® extender. Theriogenology 74:951-955.

[3] Akhter S, Ansari MS, Rakha BA, Ullah N, Andrabi SM, Khalid M (2011) In vitro evaluation of liquid-stored buffalo semen at 5°C diluted in soya lecithin based extender (Bioxcell®), tris-citric egg yolk, skim milk and egg yolk-citrate extenders. Reprod. Domest. Anim. 46:45-9. doi: 10.1111/j.1439-0531.2009.01561.x. 34

[4] Arifiantini RII, Yusuf STL (2010) Developing of tris soy milk diluent for Frisian Holstein bull frozen semen. Hayati J. Biol.17: 91-94

[5] Axnér E, Lagerson E (2016) Cryopreservation of Dog Semen in a Tris Extender with 1% or 2% Soya Bean Lecithin as a Replacement of Egg Yolk. Reprod. Domest. Anim. 51:262-8.

[6] Bousseau S, Brillard JP, Marguant-Le Guienne B, Guérin B, Camus A, Lechat M (1998) 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, 50: 699-706

[7] Cochran JD, Amann RP, Froman DP, Pickett BW (1984) Effects of centrifugation, glycerol level, cooling to 5C, freezing rate and thawing rate on the postthaw motility of equine sperm. Theriogenology 22:25-38.

[8] Cristanelli MJ, Amann RP, Squires EL, Pickett BW (1985) Effects of egg yolk and glycerol level in lactose-EDTA-egg yolk extender and of freezing rate on the motility of frozen-thawed stallion spermatozoa. Theriogenology 23:25-38.

[9] Crockett EC, Graham JK, Bruemmer JE, Squires EL (2001) Effect of cooling of equine spermatozoa before freezing on postthaw motility: preliminary results. Theriogenology 55:793-803.

[10] Duan H, Zhang X, Deng L, Zeng S, Han G (2014) Soybean lecithin in stallion semen extender maintains frozen-thawed sperm membrane integrity. J. Equine Vet. Sci. 34: 101.https://doi.org/10.1016/j.jevs.2013.10.068

[11] Duan H, Zhang X, Sun Y, Deng L, Han G (2013) Study on the Effect of Soybean Lecithin in the Cryopreservation of Stallion Semen. China Anim. Husbandry Vet. Med. 40:150-154.

[12] El-Azzazi FE, Yaseen MA (2016) Soybean lecithin-based extender for cryopreservation of ram semen. Egyptian J. Anim. Prod. 53:43-48.

[13] El-Badry DA, Abo El-Maaty AM, El Sisy GA (2017) The Effect of Trehalose Supplementation of INRA-82 Extender on Quality and Fertility of Cooled and Frozen-Thawed Stallion Spermatozoa. J. Equine Vet. Sci. 48: 86-92. DOI: 10.1016/j.jevs.2016.08.020.

[14] EL-Badry DA, Anwer AM, Rawash ZM (2014) Effect of different concentrations of sodium dodecyl sulfate, egg yolk and glycerol on the freezability and DNA integrity of Arabian stallion spermatozoa. Assiut Vet. Med. J. 60:29-35.

[15] Fathi M, Rawash Z, Dohreig R, Gamal I, Mohamed A, Abu-El Naga E, Badr M (2019) Soybean lecithin-based extender improves Damascus goat sperm cryopreservation and fertilizing potential following artificial insemination. Asian Pacific J. Reprod. 8: 174-180.

[16] Fraser L (2004) Structural damage to nuclear DNA in mammalian spermatozoa: its evaluation techniques and relationship with male infertility. Pol. J. Vet. Sci. 7:311-21

[17] Hinsch E, Hinsch KD, Boehm JG, Schill WB, Mueller-Schloesser F (1997) Functional parameters and fertilization success of Bovine semen cryopreserved in egg-yolk free and egg-yolk containing extenders. Reprod. Domest. Anim. 32: 143-149.

[18] Hughes CM, Lewis SE, McKelvey-Martin V, Thompson WA (1996) Comparison of baseline and induced DNA damage in human spermatozoa from fertile and infertile men, using a modified comet assay. Mol. Hum. Reprod. 2:613-9

[19] Khalifa EI, Abdel-Hafez MAM (2014) Effect of soybean lecithin-based semen extender on freezability and fertility of Rahmani ram spermatozoa. Egyptian J. Sheep Goat Sci. 9:59- 66.

[20] Layek SS, Mohanty TK, Kumaresan A, Parks JE (2016) Cryopreservation of bull semen: Evolution from egg yolk based to soybean-based extenders. Anim. Reprod. Sci. 172:1-9. doi: 10.1016/j.anireprosci.2016.04.013.

[21] Medica AJ, Aitken RJ, Nicolson GL, Sheridan AR, Swegen A, De Iuliis GN, Gibb Z (2021) Glycerophospholipids protect stallion spermatozoa from oxidative damage in vitro. Reprod Fertil. 2:199-209. doi: 10.1530/RAF-21-0028.

[22] Mehdipour M, Daghigh Kia H, Najafi A, Vaseghi Dodaran H, García-Álvarez O (2016) Effect of green tea (Camellia sinensis) extract and pre-freezing equilibration time on the post-thawing quality of ram semen cryopreserved in a soybean lecithin-based extender. Cryobiology, 73:297-303. doi: 10.1016/j.cryobiol.2016.10.008.

[23] Miguel-Jimenez S, Rivera Del Alamo MM, Álvarez-Rodríguez M, Hidalgo CO, Peña AI, Muiño R, Rodríguez-Gil JE, Mogas T (2020) In vitro assessment of egg yolk-, soya bean lecithin- and liposome-based extenders for cryopreservation of dairy bull semen. Anim. Reprod. Sci. 215:106315. doi: 10.1016/j.anireprosci.2020.106315.

[24] Milovanov VK. Biology of reproduction and artificial insemination of farm animals. Mosc: Monogr Selkohz Lit J Plakatov; 1962.

[25] Muiño R, Fernández M, Peña AI. Post-thaw survival and longevity of bull spermatozoa frozen with an egg yolk-based or two egg yolk-free extenders after an equilibration period of 18 h. Reprod. Domest. Anim. 2007; +3.42:305-11. doi: 10.1111/j.1439-0531.2006.00784.x.

[26] Najafi A, Daghigh-Kia H, Dodaran HV, Mehdipour M, Alvarez-Rodriguez M (2017) Ethylene glycol, but not DMSO, could replace glycerol inclusion in soybean lecithin-based extenders in ram sperm cryopreservation. Anim. Reprod. Sci. 177:35-41. doi: 10.1016/j.anireprosci.2016.12.004.

[27] Nie GJ, Wenzel JGW (2001) Adaptation of the hypo-osmotic swelling test to assess functional integrity of stallion spermatozoal plasma membranes. Theriogenology 55:1005-18.

[28] Nishijima K, Kitajima S, Koshimoto C, Morimoto M, Watanabe T, Fan J, Matsuda Y (2015) Motility and fertility of rabbit sperm cryopreserved using soybean lecithin as an alternative to egg yolk. Theriogenology 84:1172-5. doi: 10.1016/j.theriogenology.2015.06.018.

[29] Ondřej Š, Jiří Š, Jan B, Pavla MP, Lucie T, Doležalová M, Petra F, Luděk S, Radko R (2019) Low Density Lipoprotein - important player in increasing cryoprotective efficiency of soybean lecithin-based bull semen extenders. Anim. Reprod. 16:267-276. doi: 10.21451/1984-3143-AR2018-0107

[30] Papa FO, Felício GB, Melo-Oña CM, Alvarenga MA, De Vita B, Trinque C, Puoli-Filho JN, Dell'Aqua JA Jr (2011) Replacing egg yolk with soybean lecithin in the cryopreservation of stallion semen. Anim. Reprod. Sci. 129:73-7. doi: 10.1016/j.anireprosci.2011.10.006.

[31] Salmani H, Towhidi A, Zhandi M, Bahreini M, Sharafi M (2014) In vitro assessment of soybean lecithin and egg yolk-based diluents for cryopreservation of goat semen. Cryobiology, 68:276-80. doi: 10.1016/j.cryobiol.2014.02.008.

[32] Singh AK, Kumar A, Honparkhe M, Kaur S, Kaur H, Ghuman S, Brar PS (2018) Comparison of in vitro and in vivo fertilizing potential of buffalo bull semen frozen in egg yolk-, soya bean lecithin- and liposome-based extenders. Reprod Domest Anim. 53:195-202. doi: 10.1111/rda.13092.

[33] Singh AK, Singh VK, Narwade BM, Mohanty TK, Atreja SK (2012) Comparative quality assessment of buffalo (Bubalus bubalis) semen chilled (5°C) in egg yolk- and soya milk-based extenders. Reprod. Domest. Anim. 47:596-600. doi: 10.1111/j.1439-0531.2011.01928.x.

[34] Speroni F, Anón MC, de Lamballerie M (2010) Effects of calcium and high pressure on soybean proteins: a calorimetric study. Food Res. Int. 43: 1347-1355.

[35] Taladrid D, Marín D, Alemán A, Álvarez-Acero I, Montero P, Gómez-Guillén M (2017) Effect of chemical composition and sonication procedure on properties of food-grade soy lecithin liposomes with added glycerol. Food Res. Int. 100:541-550. doi: 10.1016/j.foodres.2017.07.052.

[36] van Wagtendonk-de Leeuw AM, Haring RM, Kaal-Lansbergen LM, den Daas JH (2000) Fertility results using bovine semen cryopreserved with extenders based on egg yolk and soy bean extract. Theriogenology 54:57-67. doi: 10.1016/S0093-691X(00)00324-1.

[37] Wells ME, Awa OA (1970) New technique for assessing acrosomal characteristics of spermatozoa. J. Dairy Sci. 53:327-32

[38] Zhao JQ, Xiao GL, Zhu WL, Fang D, Li N, Han CM, Gao QH (2021) Ram semen preserved at 0°C with soybean lecithin Tris-based extender substituted for egg yolk. Anim. Biosci. 34:192-197. doi: 10.5713/ajas.20.0118.

Abstract

References

How to cite