This study was aimed to analyze the metabolic activity and membrane integrity of boar spermatozoa following storage in long-term semen extenders. Boar semen was diluted with AndrohepR EnduraGuardTM (AeG), DILU-Cell (DC), SafeCell PlusTM (SCP) and Vitasem LD (VLD) extenders and stored for 10 days at 17oC. Parameters of the analyzed sperm metabolic activity included total motility (TMOT), progressive motility (PMOT), high mitochondrial membrane potential (MMP) and ATP content, whereas those of the membrane integrity included plasma membrane integrity (PMI) and normal apical ridge (NAR) acrosome. Extender type was a significant (P < 0.05) source of variation in all the analyzed sperm parameters, except for ATP content. Furthermore, the storage time had a significant effect (P < 0.05) on the sperm metabolic activity and membrane integrity during semen storage. In all extenders the metabolic activity and membrane integrity of the stored spermatozoa decreased continuously over time. Among the four analyzed extenders, AeG and SCP showed the best performance in terms of TMOT and PMI on Days 5, 7 and 10 of storage. Marked differences in the proportions of spermatozoa with high MMP were observed between the extenders, particularly on Day 10 of storage. There were not any marked differences in sperm ATP content between the extenders, regardless of the storage time. Furthermore, the percentage of spermatozoa with NAR acrosomes decreased during prolonged storage, being markedly lower in DC-diluted semen compared with semen diluted with either AeG or SCP extender. The results of this study indicated that components of the long-term extenders have different effects on the sperm functionality and prolonged semen longevity by delaying the processes associated with sperm ageing during liquid storage.

Boar variability affects sperm metabolism activity in liquid stored semen at 5°C: Metabolic activity of boar spermatozoa, liquid stored for three days at 5°C, was measured using bioluminescence for ATP content, fluorescent assay (JC fluorochrome) of mitochondrial activity and oxygen consumption. Sperm motility and plasma membrane integrity (PMI) were simultaneously analyzed. Apart from the statistically significant effect (P < 0.001) of semen storage time, the importance of the individual source of the ejaculate for the analyzed parameters of metabolic efficiency of spermatozoa was shown. This phenomenon was manifested in the interaction of the individual source of the ejaculate with spermatozoa motility, integrity of their membranes and metabolic activity with the passing time of semen preservation. Recorded results indicate that the individual factor may have a significant influence on the technological usefulness of boar spermatozoa for liquid storage. Quality analyses conducted on boar semen stored at 5°C may be used for pre-selection of boars producing sperm with an enhanced tolerance to cold shock.

The aim of this study was to investigate the effect the sperm-rich fraction (F1) and the post-F1 fraction (F2) on the quality of boar spermatozoa stored in a liquid state. Ejaculates were collected from three Polish Landrace boars. Each ejaculate fraction was diluted with BTS short-term extender and Safe-Cell Plus (SCP) long-term extender and stored for seven days (D1-D7) at 17°C. Analyses included sperm motility parameters, normal apical ridge (NAR) acrosomes and plasma membrane integrity (PMI). Prior to the dilution of fractions, marked changes (p<0.05) were noted between F1 and F2 in progressive motility (PMOT), velocity average pathway (VAP) and velocity straight line (VCL). After the ejaculate was diluted, the type of fraction and type of extender significantly affected (p<0.05) PMOT, being markedly higher (p<0.05) for F1 extended in BTS. No marked changes (p<0.05) were observed between F1 and F2 extended in SCP for any of the analyzed sperm quality parameters during seven days of storage. Significantly higher (p<0.05) values of sperm quality parameters were noted in F1 compared with F2 for BTS on D7 of storage. The results of the four-way ANOVA analysis indicate that boar, fraction of ejaculate, extender type and day of storage had significant effects on the quality of boar stored spermatozoa. The F1 was characterised by higher quality of spermatozoa during storage in comparison with F2 in the short-term extender. Using the long-term extender containing the proteins allowed for a better application of F2, which could be important for the pig industry.

The aim of this study was to evaluate the effect of different extenders on the quality of European red deer epididymal sperm stored at 5 °C. Epididymal spermatozoa were collected post mortem from 10 stags and diluted with three extenders (Bovidyl^®, BoviFree^®, and BioXcell^®) and stored at 5 °C. Sperm motility (TMOT), motility parameters (system CASA), plasma membrane integrity (SYBR-14⁺/PI⁻), acrosomal membrane integrity (FITC-PNA⁻/PI⁻), mitochondrial activity (JC-1/PI), viability, and apoptotic-like changes (YOPRO/PI) were evaluated. The analyses were conducted on the first and successive days of storage (D1–D7). The applied extender, storage time, and the interactions between these factors significantly (p < 0.001) affected most of the analyzed parameters whose values were highest in sperm samples stored in Bovidyl^®, regardless of storage time. In Bovidyl^®, BoviFree^®, and BioXcell^® extenders, TMOT values were estimated at 83%, 63%, and 59%, respectively, on D3. The extenders significantly influenced DNA integrity on D7. The percentage of dead sperm increased from D3. The quality of stored sperm cells was significantly influenced by the extenders’ biochemical composition. BoviFree^® and BioXcell^® contain glycerol which could contribute to deteriorating the quality of spermatozoa stored at 5 °C. Sperm cells stored in the egg yolk-based extender (Bovidyl^®) were characterized by the highest viability and functionality.

The aim of the study was to evaluate viability longevity and quality of liquid-stored epididymal sperm of European red deer (Cervus elaphus elaphus). Sperm samples were recovered from epididymides of 15 mature stags. Samples were diluted to 100 × 106 sperm⁄mL in modified Salomon's extender and stored at 5 °C for 25 days. Sperm were analyzed to determine total motility (TMOT), progressive motility (PMOT) and motion variables [CASA system], acrosomes with normal apical ridges (NAR), viability and acrosomal status (FITC-PNA/PI), plasma membrane integrity (PMI) and mitochondrial membrane potential (MMP) on the first day (D1) and every other day of storage (D3-D25). Data were analyzed using a one-way analysis of variance (ANOVA). Spermatozoa remained motile until D25, whereas TMOT exceeded 60%, NAR and PMI exceeded 80%, FITC-PNA/PI exceeded 75%, and MMP approximated 70% until D11. There was a lesser (P < 0.05) value for these variables on D5 (relative to D1). Furthermore, there was a decrease in values for motility variables as duration of storage increased, excluding amplitude of lateral head displacement, on D3. The results indicate that liquid-stored epididymal sperm of European red deer are characterized as having a long viability (25 days) with a retention of sperm quality for as long as 11 days in the liquid storage state. In vitro or in vivo studies, however, are required to confirm the findings in the present study.