# Alginate Silver Nanoparticles and Their Effect on Sperm Parameters of the Domestic Rabbit
## 3 Results and Discussion
### Analysis of Morphological Changes in Rabbit Sperm
They also evaluated the effects of these three agents on rabbit semen parameters. In addition to their antibacterial properties, the compounds showed no effect on total sperm motility, acrosome integrity, membrane functionality or rabbit spermatozoid viability. Also, after 24, 48, or 72 h of exposure, they found no significant changes in these parameters of rabbit semen. Silver nanoparticles suspended in sodium alginate, analyzed in this work, can be considered as a potential alternative to antibiotics in semen diluents. However, these compounds induce a relatively high level of secondary changes in the morphology of rabbit spermatozoa after 24 h of treatment; AgNP AlgX 24 h generated >24% changes and AgNP AlgB 24 h generated 34% changes, despite the small effect of both solutions on chromatin integrity in the acrosome demonstrated by the comet assay. [PERSON] et al. [78], in their study of modified pellet composition, found in a control sample of rabbit ejaculate pH 6.75, sperm viability of 88.75%; and, in a morphological analysis, normal spermatozoids accounted for 82.75%, and spermatozoa with a coiled tail accounted for 2.00%, while the presence of spermatozoa with a looped tail was at 3.75%. [PERSON] et al. [79], examining the abnormality of spermatozoa obtained from three breeds of rabbits, including New Zealand Whites, showed the presence of 83.50 \(\pm\) 0.20% normal spermatozoa and 16.5 \(\pm\) 0.20% abnormal spermatozoa. Within the altered spermatozoids, acrosome lesions accounted for 1.71 \(\pm\) 0.18%, the presence of protoplasmic droplet was at 0.42 \(\pm\) 0.15%, and 1.03% represented spermatozoa with a coiled tail. In addition, it was found that the pH of the ejacate was significantly dependent on the percentage of primary and secondary abnormalities. Sodium alginate is not harmful to health, which has been confirmed by opinion institutions, including FAO/WHO experts and the U.S. FDA have recognized the compound as a safe food additive, including for children [80]. Above that, alginate is not absorbed into the body. Taken in the recommended amounts, as a rule, it does not cause any side effects. Some harmfulness can be spoken of only in the case of non-compliance with the recommended doses. When significantly exceeded, this compound can impair the absorption of proteins and some elements [81; 82]. Despite the lack of toxic properties, it was sodium alginate that showed the strongest effect after 24 h in altering the stability of rabbit sperm chromatin, relative to the other compounds tested. In the comet assay, Alg 24 h showed the highest %Tail DNA and TM values after 24 h exposure compared to the negative control 24 h, AgNP AlgX 24 h and AgNP AlgB 24 h.  
A factor influencing the effectiveness of the silver nanoparticles used in the study, in addition to sodium alginate as a carrier, was the reductant used--xylose. This is a monosaccharide obtained by hydrolyzing hemicellulose-rich plants, such as straw or corn cobs [83]. One of the primary components of semen diluents and cryopreservatives is sugar, usually glucose or fructose. In that medium, sugars have several important functions, as they provide an energy substrate for sperm during storage, maintain the appropriate osmotic pressure of the diluent, and act as a cryoprotectant. [PERSON] et al. [84] studied the effects of various sugars added to the diluent on the viability, motility and rate of acrosomal reaction in canine spermatozoa during dilution, standardization and freezing. At the stage of assessing the quality of the material, they showed that xylose was one of those sugars, along with fructose, galactose and maltose, that provided higher total rates of active spermatozoa compared to the control without added sugar. Samples with xylose addition had higher survival rates and lower percentages of dead and acrosome-damaged cells than the control. The opposite effect of xylose on water buffalo spermatozoa was demonstrated by [PERSON] et al.