Introduction
The production of agro-industrial by-products has increased in recent years, posing environmental problems and economic challenges regarding their valorization within the circular economy. Over 54 million tons of peanuts (Arachis hypogaea) are produced annually, resulting in approximately 0.8 to 1.6 million tons of peanut peels (PP) that are typically discarded despite their potential for valorization as aquafeed ingredients (FAO, 2025). PPs are rich in polyphenols, including flavonoids, procyanidins, and phenolic acids, that exhibit high antioxidant activity. With aquaculture intensification and global warming, fish are increasingly exposed to thermal challenges, leading to increased oxidative stress and disease susceptibility. Dietary supplementation with antioxidants may help maintain redox balance and improve fish resilience to thermal stress. In this context, identifying sustainable and natural antioxidants is of utmost importance to address this gap. This study aimed to: 1) optimize PP extraction to maximize bioactive compounds yield; 2) evaluate the potential of the selected extract on improving zootechnical performance and survival of European seabass under normal and thermal challenge rearing conditions.
Materials and Methods
PP extraction was performed using 50% (v/v) ethanol and water in a microwave-assisted extraction. A full factorial design with solid-liquid ratio (SLR) and extraction time as independent variables was used to determine the optimal conditions to maximize the yield of bioactive compounds from PP. SLR of 1:35, 1:45, and 1:55 (w/v), and extraction times of 30, 60, and 90 s were tested, followed by a 30-minute soaking. Then, all mixtures were vacuum-filtered, and total antioxidant activity, total phenols, ferric reducing antioxidant power (FRAP), flavonoids, and total protein were evaluated. Data from the full factorial design were analyzed using the response surface methodology in Statistica 10. The theoretical optimum (SLR 1:55 for 90s) yielded the highest bioactive compound content. However, due to practical limitations associated with the high solvent volume, the SLR 1:35 and 90s extract was selected for diet inclusion.
Three isoproteic (46% CP) and isolipidic (18% CL) diets were formulated: a control diet with no extract and two diets containing the selected PP extract (SLR 1:35 and 90s) to enrich the control diet with 500 ppm and 1000 ppm total antioxidants (Diets Extract 1 and 2), respectively. PP extract was mixed with the diets in liquid form, then dried at 40 °C for 48 h to remove water and ethanol. Triplicate groups of European seabass (Dicentrarchus labrax) juveniles (IBW: 74 g) were fed these diets to satiation for 4 weeks in a RAS system at 24 °C. After the feeding trial, fish were subjected to a 7-day thermal challenge (temperature increased to 30 °C over 2 days and oxygen maintained at 4 mg/L) to simulate heatwave conditions. At the end of the feeding trial, zootechnical performance parameters were recorded, and during the thermal challenge, mortality was registered. Data on zootechnical performance were analyzed using one-way ANOVA (p<0.05). Survival was analyzed using the Kaplan–Meier method, and group differences were assessed using the log-rank test in GraphPad 8.
Results
Overall, the experimental and predicted values from the response surface model were consistent, supporting the model's robustness. Increasing SLR and time generally improved DPPH, phenols, soluble protein, FRAP, and flavonoids contents, while DPPH decreased with increasing extraction time. The extract SRL 1:55; 90s had the highest levels of phenolics, soluble protein, FRAP, and flavonoids. Nonetheless, due to the limitation of including high solvent volumes in the diets, the SLR 1:35; 90s extract was selected and scaled up for inclusion, as it also provided a high antioxidant yield. The experimental diets (Extracts 1 and 2) did not affect weight gain, feed intake, feed efficiency, DGI, or PER compared to the control diet. During the thermal challenge, survival of fish fed the Extract 2 diet was higher than that of fish fed the control and Extract 1 diets (Figure 1).
Conclusion
PP extracts have high antioxidant content and the potential to increase fish survival under chronic thermal stress when supplemented to diets at 1000 ppm.
Figure 1 – Kaplan-Meier survival curves of European seabass fed the experimental diets during the 7-day thermal challenge. Data is represented as mean (n=3) and standard deviation. Significant differences are denoted by asterisks ** (p-value = 0.002).
Acknowledgments
LV and RM were supported by the Portuguese Foundation for Science and Technology (FCT) through a doctoral grant (2023.02516.BD) and a CEEC Individual contract (DOI https://doi.org/10.54499/2023.09082.CEECIND /CP2848/CT0017), respectively. This work was funded by the FCT-funded project "Zero-Waste Omega-3: A Green Label Solution for Sustainable Aquaculture Feeds (DOI: https://doi.org/10.54499/2023.13017.PEX).
References
FAO. (2025). FAOSTAT – Production: Crops and livestock products. Retrieved from https://www.fao.org/faostat /en/#data/QCL