Optimization of Black Boar Sperm Cryopreservation Efficiency with Antioxidant-Rich Plant Extracts from Djulis (Chenopodium formosanum).

The cryopreservation of boar sperm effectively extends its storage period but often leads to increased reactive oxygen species (ROS) production, compromising sperm quality. Plant extracts, rich in bioactive compounds such as polyphenols and flavonoids, have been shown to reduce ROS. Djulis (Chenopodium formosanum), also known as the "ruby of cereals", is nutritionally rich and holds potential as a cryoprotective additive. This study aimed to determine the optimal concentration of extracts from different parts of djulis, including unhulled seeds and stems, for effective boar semen cryopreservation. Fresh semen from Taiwan indigenous boars was diluted with a modified GLT-cryoprotectant extender containing glycerol, low-density lipoprotein (LDL), and trehalose. The experimental groups included DSS25, DSS50, DS25, and DS50-representing djulis unshelled seed at 25 mg/mL and 50 mg/mL, and djulis stem at 25 mg/mL and 50 mg/mL in distilled water, respectively-alongside a control group without additives. Post-thaw assessments included sperm motility, kinetic parameters, viability, acrosome integrity, and the antioxidant properties of djulis extracts, such as DPPH radical scavenging activity and total phenolic acid content. Results showed that total motility (TM) was significantly higher in the DSS25 (48.8 ± 3.9), DSS50 (49.0 ± 6.7), and DS50 (49.0 ± 2.4) groups compared to the control group (31.3 ± 4.8). Similarly, progressive motility (PM) was significantly improved in DSS25 (27.5 ± 2.7) and DSS50 (26.8 ± 4.1) versus the control (12.8 ± 3.2). However, for straightness (STR), the control group (87.8 ± 1.3) exhibited significantly higher values than the DS50 group (83.5 ± 1.3) (p < 0.05). Viability and acrosome integrity showed no significant differences across groups. In conclusion, djulis extracts positively influence sperm motility and forward movement, with 1% djulis extract confirmed to enhance the quality of cryopreserved semen. Future research will focus on determining the optimal dosage of djulis extract for improved cryopreservation outcomes.