One of the critical points in the cryopreservation process is the use of a proper diluent while lowering the temperature following the resuspension and thawing processes. Here, we tested an alternative diluent for the process of freezing boar semen. We used skimmed dried milk (SDM) during the cooling and post-thawed resuspension steps. To do so, we collected semen from 15 Dalland boars using the glovedhand technique, and incubated each ejaculate sample at 30 °C. We then removed two semen aliquots from a pre-dilution. We diluted one of the aliquots in Beltsville thawing solution (BTS -control), and the remaining sample was diluted in SDM. Both aliquots were subsequently held at 30º C for 45 min (1 st period of stabilization). At the end of this period, we analysed vigor and motility to determine sperm metabolic activity. We then held the diluted semen at 25° C for 30 min (2 nd period of stabilization) and at 17° C for 2 h (3 rd period stabilization). We centrifuged the semen at 800 × g and 1600 × g at 5º C for 15 min, discarded the supernatant, and resuspended the sperm pellet in 2 mL of the cooling diluent at 5° C for 1h. We again diluted the samples in 2 mL of the freezing diluent, poured them into straws, and cooled and plunged them into liquid N 2 . The sêmen samples were thawed in a 39º C water bath, and were resuspended in their respective diluents at the same temperature. We determined the following sperm features: vigor, motility, vitality, acrosomal integrity and membrane functionality. During the first phase of temperature cooling (30º C), semen diluted in SDM exhibited a higher vigor (3.4 ± 0.6) and motility (78.6 ± 13.0) than those diluted BTS (vigor: 3.1 ± 0.7; motility: 69.4±14.3). However, after the thawing procedure, the inverse was observed in that: BTS samples exhibited a higher vigor (2.1 ± 0.6) and motility (35.5 ± 21.0) than SDM samples (vigor: 1.7 ± 0.9; motility: 22.8 ± 18.1). Regarding membrane functionality and acrosomal integrity, we did not find a significant difference between BTS and SDM, although SDM provided a higher percentage of living cells at the end of the freeze/thaw procedures (72.3 ± 16.4). In summary, we suggest that BTS should be considered a better option than SDM for the cooling and post-thaw resuspension steps of boar semen freezing process.
ResumoEsse estudo objetivou testar um diluente alternativo, a base de leite em pó desnatado, durante as etapas de resfriamento e ressuspensão pós descongelação durante a congelação do sêmen suíno. Para isso o sêmen de 15 varrões da raça Dalland, foi coletado através da técnica da mão enluvada. Visando o início da curva de congelação, cada ejaculado foi incubado a 30 °C. Em seguida, foram retiradas duas alíquotas de sêmen para uma pré-diluição. Uma das alíquotas foi diluída no diluente Beltsville Thawing Solution (BTS -Controle) e a outra em leite em pó desnatado (LPD), onde permaneceram a essa temperatura por 45 minutos. O sêmen pré-diluído passou por três tempos de estabilização. Ao final do primeiro tempo (30 °C -45 minut...