Effect of Glycerol Concentration, Freezing Rate and Thawing Rate on Semen Characteristics in PoongSan-dog

풍산개 정자의 동결보존에 있어서 Glycerol 농도, 동결 및 융해속도가 정자성상에 미치는 영향

  • Ji, D.Y. (Seoul Grand Park Zoo) ;
  • Yoon, T.C. (Korea University Institute of life Science and Natural Resources) ;
  • Rho, J.R. (Seoul Grand Park Zoo) ;
  • Cho, S.R. (National Institute of Animal Science, RDA) ;
  • Kim, C.K. (Department of Animal Science & Technology, Chung-Ang University) ;
  • Pang, M.G. (Department of Animal Science & Technology, Chung-Ang University) ;
  • Kim, Bo-Sook (Seoul Grand Park Zoo)
  • Published : 2007.10.31


This research was carried out in order to establish the production technique for Poong-san dog’s frozen semen, by examining the semen characteristic and the volume of glycerol added to the dilution solution, thawing temperature and sperm motility and viability as well as the motility using CASA according to time variation. Average semen volume was 5.9ml, sperm concentration 116.3×106 sperm/ml, total sperm number 789.3×106 sperm, motility 88.7±1.7% and viability 87.6±7.8%. When it was cryopreservation and thawed at different glycerol concentrated extender, it showed 52.7% motility and 57.7±10.3% viability at 7% glycerol, compared to other treatments. For semen cryogeny, at conditions of 5, 7cm and a height of 10cm for pre-cryogeny and maintaining the semen at 7cm from the surface of liquid nitrogen resulted in profitable motility and viability.


PoongSan-dog;Semen characteristic;Glycerol;Freezing;Thawing


  1. Gill, H. P., Kaufman, C. F., Foote, R. H. and Kirk, R. W. 1960. Artificial insemination of beagle bitches with freshly collected, liquid-stored and frozen semen. Am. J. Vet. Res. 31:1807-1813
  2. Harrop, A. E. 1962. Artificial insemination in the dog. In: The semen of animals and artificial insemination. J. P. Maule, ed. Commonwealth Agri. Bureaux, Farnham Royal, England. 186-189
  3. Hewitt, D. A., Leahy, R., Shwldon, I. M. and England, G. C. W. 2001. Cryopreservation of epididymal dog sperm. Anim. Reprod. Sci 67:101-111
  4. Ivanova-Kicheva, M. G., Subev, M. S., Bobadov, N. D., Dacheva, D. P. and Rouseva, L. A. 1995. Effect of thawing regimens on the morphofunctional state of canine spermatozoa. Theriogenology 44:563-569
  5. Morton, D. B. and Bruce, S. G. 1989. Semen evalution, cryopreservation and factors relevant to the use of frozen semen in dog. J. Reprod. Fertil. (Suppl.) 39:311-316
  6. Okano, T., Murase, T., Asano, M. and Tsubota, T. 2004. Effects of final dilution rate, sperm concentration and times for cooling and glycerol equilibration on post-thaw characteristics of canine spermatozoa. J. Vet. Med. Sci. 66:1359-1364
  7. Petrunkina, A. M., Gropper, B., Topfer-Petersen. E. and Gunzel-Apel. A. P. 2005. Volume regulatory function and sperm membrane dynamics as parameters for evaluating cryoprotective efficiency of a freezing extender. Theriogenology 15;63:1390-406
  8. Pena, A., Johannisson, A. and Linde-Forsberg, C. 1999. Post-thaw evaluation of dog spermatozoa using new triple fluorescent staining and flow cytometry. Theriogenology 52:965-960
  9. Platz, C. C. and Seager, S. W. 1977. Successful pregnancies with concentrated frozen canine semen. Lab. Anim. Sci. 27:1013-1016
  10. Rota, A., Frishling, A., Vannozzi, I., Camillo, F. and Romagnoli, S. 2001. Effect of the inclusion of skimmed milk in freezing extenders on the viability of canine spermatozoa after thawing. J. Reprod. Fertil(Suppl.) 57:377-381
  11. Rota, A., Strom, B., Linde-Forsberg, C. and Rodriguez-Martinez, H. 1997. Effects of equex STM paste on viability of frozen-thawed dog spermatozoa during in vitro incubation at $38^{\circ}C$. Theriogenology 47:1093-1101
  12. Seager, S. W. 1969. Successful pregnancies utilizing frozen dog semen. AI Digest. 17:6-7
  13. Silva, L. D. M. and Verstegen, J. P. 1995. Comparisons between three different extenders for canine intrauterine insemination with frozen-thawed spermatozoa. Theriogenology 44:571-579
  14. Takagi, H., Kurihara, A., Inoue, T., Nakamura, I. and Kimura, M. 2001. Investigation of usefulness of sperm analyses in dog for male fertility study. Toxicological Sci. 25:313-321
  15. Tsutsui, T., Hase, M., Tanaka, A., Fujimura, N., Hori, T. and Kawakami, E. 2000. Intrauterine and intravaginal insemination with frozen canine semen using an extender consisting of Orvus ES Paste-supplemented egg yolk tris-fructose citrate. J. Vet. Med. Sci. 62(6):603-6006
  16. Tsutsui, T., Tezuka, T., Mikasa, Y., Sugisawa, H., Kirihara, N., Hori, T. and Kawakami, E. 2003. Artificial insemination with canine semen stored at a low temperature. J. Vet. Med. Sci. 65(3):307-312
  17. Voelkel, S. A. and Hu, Y. X. 1992. Use of ethylene glycol as a cryoprotectant for bovine embryos allowing direct transfer of frozen-thawed embryos to recipient females. Theriogenology 37:687-697
  18. Yildiz, C., Kaya, A., Akoy, M. and Tekeli, T. 2000. Influence of sugar supplementation of the extender on motility, viability and acrosomal integrity of spermatozoa during freezing. Theriogenology 54:579-585
  19. 이영락, 이성림, 강태영, 최상용. 2003. 개 정자의 동결융해 후 생존성 및 첨체의 변화. 한국수정란이식학회지. 18:151-159
  20. 이장희, 김인철. 1999. 돼지정자의 동결융해 후 활력 및 생존성에 대한 보존액, 통해보호제, 예비동결 및 동결처리시간의 영향. 한국동물번식학회지. 23:165-174
  21. 이제협, 박향, 박흠대, 김재명. 2003. 개 정자의 동결보존에 있어서 Glycerol 농도, 동결 및 융해 속도가 정자의 생존율 및 운동성에 미치는 영향. 한국수정란이식학회지. 18:195-201
  22. 김흥률, 이계웅, 공일근. 2002. 진도개 동결정액 제조를 위한 정액성상과 동결정액의 운동성에 관한 연구. 한국동물번식학회지. 26:253-261
  23. Notlig, J. O. and Shuttleworth, R. 2005. The effect of straw size, freezing rate and thawing rate upon post-thaw quality of dog semen. Theriogenology 15;63:1469-80
  24. Rota, A., Strom, B. and Linde-Forsberg, C. 1995. Effects of seminal plasma and three extenders on canine semen stored $4^{\circ}C$. Theriogenology 44:885-900