Uso do extrato líquido de melancia (Citrullus lanatus) na criopreservação de sêmen de touros da raça Nelore

• Autor(a) principal: Castilho, Erick Fonseca de
• Data de Publicação: 2012
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/1460
• topic: Antioxidantes; Crioprotetores; Bovinos; Antioxidants; Cryoprotectors; Cattle; CNPQ::CIENCIAS AGRARIAS::MEDICINA VETERINARIA::REPRODUCAO ANIMAL

The objectives of this study were to evaluate the potential of liquid watermelon extract as a natural diluent for cooling and/or freezing semen, and determining the cryoprotectant effect of liquid watermelon extract on the integrity of the plasma membrane of bovine sperm. Semen from four adult bulls were used, totalizing 10 semen samples for each one. After semen collection, the evaluation consisted of physical and morphological exam, live and dead cells (supravital test) and hypoosmotic swelling test. Afterwards, the fresh semen was divided in four equal parts and diluted with respective extenders: D1 (Bovimix® - control); D2 (Bovimix® base + 25 % of liquid watermelon extract); D3 (Bovimix® base + 50 % of liquid watermelon extract); and D4 (100 % of liquid watermelon extract base). After final dilutions, sperm motility and vigor were evaluated in each extender, and posterior seal. The straws were cooled and frozen in the freezing machine (Cryogen Dualflex®). Then, the straws were immersed in nitrogen. The thawing of the samples was done by immersion of the straws in a 37 °C water-bath for 30 minutes to evaluate the sperm motility and vigor, morphological characteristics of sperm, supravital test, hypoosmotic swelling test, epifluorescence test and thermoresistence test. The volume, motility, sperm vigor, sperm concentration per mL, supravital test, hypoosmotic swelling test, major and total sperm defects varied among animals, but showed no difference between them (P>0.05). The appearance, whirlpool and minor sperm defects showed differences among animals (P<0.05). Sperm motility showed high and positive correlation (r = 0.98) with the hypoosmotic swelling test. The supravital test showed high and positive correlation (r = 0.88) with the hypoosmotic swelling test. There was an average and negative correlation (r = -0.48) between motility and sperm concentration per mL, as well as average and negative correlation (r = -0.46) between the hypoosmotic swelling test and sperm concentration per mL. The motility and sperm vigor of the diluted semen (pre-cooling) in all extenders used, presented no difference between them (P>0.05). The motility upon thawing and at the end of thermoresistence test differed between extenders (P<0.05), where higher values were obtained by diluents D1 and D2, and lower values were obtained by diluents D3 and D4. At the beginning of the thermoresistence test, there was no difference between diluents D1 and D2. However, there was a difference when comparing the diluent D1 with D3 and D4. There was no difference between the diluent D2 and D3 or D2 and D4. At the end of the thermoresistence test, there was no difference between the diluents D1 and D2 or D1 and D4, but there was a difference between D1 and D3. There was no difference when comparing the diluent D2 with D3 and D4. The average sperm alive in the supravital test at the time of thawing in all extenders used, presented no difference between them (P>0.05). During the thermoresistence test, the average vigor of postthawed semen, in all extenders used, showed no difference between them (P>0.05). The hypoosmotic swelling test differed between extenders (P<0.05), where higher values were recorded in the diluent D1, an intermediate value was recorded in the diluent D2 and lower values were recorded in the diluents D3 and D4. There was no difference when comparing the diluent D1 with D2 and, no difference in between the diluents D2 and D3 or D2 and D4. However, there was a difference when comparing the diluent D1 with D3 and D4. There was no difference between the diluents D2 and D3 or D4. The epifluorescence test differed between extenders (P<0.05), but there was no difference between diluents D1 and D2. There was no difference between the diluents D2 and D3, or between the diluents D3 and D4. However, there was a difference when comparing D1 with D3 and D4. There were differences between diluents D2 and D4. The major and total sperm defects ranged among extenders, but showed no difference between them (P>0.05). However, minor sperm defects differed between extenders (P<0.05), where the highest values were observed in the diluent D3, and lower values were observed in the other diluents. There was no difference between diluent D1 and the other extenders. However, there was a difference between diluents D3 and D4. In the diluent D1, the values obtained in the supravital test showed average and positive correlation (r = 0.49) with sperm motility. The hypoosmotic swelling test showed high and positive correlation (r = 0.89) with intact sperm and, high and negative correlation (r = -0.80) with damaged spermatozoa observed in epifluorescence test. In the diluent D2, the values obtained in hypoosmotic test showed high and positive correlation (r = 0.86) with intact sperm, high and negative correlation (r = -0.83) with damaged spermatozoa and, average and positive correlation (r = 0.41) with semi-damaged sperm observed in epifluorescence test. In the diluent 3, the values obtained in the supravital test showed low and positive correlation (r = 0.33) with sperm motility. The hypoosmotic test showed high and positive correlation (r = 0.82) with sperm integrity, high and negative correlation (r = -0.80) with damaged sperm and, average and positive correlation (r = 0.51) with semi-damaged sperm observed in epifluorescence test. In the diluent D4, the values obtained in the supravital test showed average and positive correlation (r = 0.46) with sperm motility. The hypoosmotic swelling test showed high and positive correlation (r = 0.83) with intact sperm, high and negative correlation (r = - 0.85) with damaged sperm and, low and positive correlation (r = 0.38) with semidamaged sperm observed in epifluorescence test. At concentrations of 50 % and 100 %, the liquid watermelon extract was not effective in maintaining the integrity and sperm viability after thawing. At the concentration of 25 %, the liquid watermelon extract maintained the structural integrity of the membrane of spermatozoa during the cryopreservation process, and viable after the thermoresistence test, could be an alternative in the composition of diluents for cryopreservation of bovine semen.