# The Effects of Graphene Oxide Nanoparticles on the Cryopreservation of Angora Buck Sperm

## TL;DR

This study explores how graphene oxide nanoparticles affect the freezing and thawing of Angora buck sperm, finding that certain sizes and doses improve sperm quality after freezing.

## Contribution

The study introduces graphene oxide nanoparticles as a potential cryoprotectant for Angora buck sperm, showing size- and dose-dependent effects.

## Key findings

- 50 nm NGO at 50 µg/mL significantly improved motility and plasma membrane and acrosome integrity.
- 500 nm NGO at 50 µg/mL only improved plasma membrane and acrosome integrity.
- All NGO sizes and doses significantly protected sperm DNA integrity after thawing.

## Abstract

Nano-graphene oxide (NGO) is a nanomaterial that has been frequently used in the fields of health and bioengineering in recent years. However, its potential use in semen cryopreservation is still in the exploratory phase. In this study, Angora bucks, a breed with low resistance to cold shock, were used. Sperm was collected from five different Angora bucks, pooled, diluted with a Tris-based egg yolk diluent, and frozen with the addition of NGO at two different sizes (50 and 500 nm) and doses (10 and 50 µg/mL). Nanoparticle characterization was performed using field emission scanning electron microscopy (FE-SEM), dynamic light scattering (DLS), and Fourier-transform infrared spectroscopy (FTIR). Post-thaw sperm analyses were evaluated based on motility and kinematic parameters, mitochondrial membrane potential (MMP), plasma membrane and acrosome integrity (PMAI), and DNA fragmentation. Applying 50 nm NGO at a dose of 50 µg/mL led to statistically significant improvements in motility and PMAI (p < 0.05). The same dose of 500 nm NGO, however, only showed a statistically significant improvement in the PMAI parameter (p < 0.05). No significant differences were observed between the groups for MMP and kinematic parameters (p > 0.05). Conversely, it was found that all sizes and doses of NGO significantly protected post-thaw sperm regarding DNA integrity (p < 0.05). These findings indicate that the NGO, at a size of 50 nm and a dose of 50 µg/mL, improves the post-thaw quality of Angora buck sperm and provides a cryoprotective effect that depends on size and dose. This study provides preliminary data on the potential effects of NGO; however, comprehensive mechanistic and in vivo validation studies are required to establish the biological and clinical validity of these findings.