Introduction: The zebrafish (Danio rerio) is a widely used model in biological and genetic research, and it is also a popular species in the aquarium trade. Its rapid growth, low maintenance requirements, transparent larval stages, and genetic similarities to humans make it an ideal species for studying physiological and molecular responses to environmental stimuli (Lawrence, 2007; Ulloa et al., 2014). However, like other aquatic organisms, zebrafish are exposed to various environmental stressors that can affect their health, growth, and performance. One common stressor in aquaculture is transportation, which involves sudden environmental changes, temperature fluctuations, and reduced oxygen availability, often leading to biochemical alterations and elevated cortisol levels (Carmichael, 1984; Barton, 2002; Brick and Cech, 2002; Hoseini et al., 2022). Natural feed additives, such as medicinal plants, have gained attention as a strategy to mitigate stress effects in fish. Parsley (Petroselinum crispum) is rich in antioxidants, vitamin C, and anti-inflammatory compounds and has the potential to improve fish health and stress resilience (Citarasu, 2010; Harikrishnan et al., 2011; Nouioura et al., 2024). Although several studies have examined other medicinal plants, limited information exists on the effects of dietary parsley powder on stress indicators in zebrafish. Therefore, the present study aimed to evaluate the impact of different inclusion levels of parsley powder on the physiological responses of zebrafish during transportation stress. The results are expected to provide scientific evidence supporting the use of parsley as a natural additive to enhance fish welfare and stress management in aquaculture.
Materials and Methods: Dried parsley powder, sieved (≤30 μ), was incorporated into a commercial base diet at 0.5%, 1%, and 2% levels and coated with gelatin. A total of 300 zebrafish were distributed into 4 groups of Control, 0.5%, 1%, and 2% (C, T1, T2, and T3, respectively) with 3 replicates and fed the experimental diets for 8 weeks. Fish were subjected to a 2-hour simulated transportation stress at the end of the trial. Post-stress, whole-body homogenates were prepared. Serum biochemical parameters, including glucose, cholesterol, triglycerides, lactate dehydrogenase (LDH), liver enzymes (AST, ALT, ALP), and cortisol were assessed. Data were analyzed by one-way ANOVA and Tukey's test using GraphPad Prism version 8.
Results and Discussion: The present findings demonstrate that dietary parsley supplementation significantly modulated the serum biochemical profile of zebrafish in a dose-dependent manner. The most pronounced effects were observed in the groups receiving T2 and T3 parsley. A marked reduction of approximately 25-30% in serum glucose and cholesterol levels compared to the control group underscores parsley's potential in mitigating metabolic stress (Farzaei et al., 2013) and regulating lipid synthesis, likely through the bioactive inhibition of key enzymatic pathways such as HMG-CoA reductase, as previously suggested (Schumacher and DeBose-Boyd, 2021). Furthermore, the significant 20% decrease in triglyceride levels in the T2 and T3 groups points towards enhanced lipid metabolism and β-oxidation of fatty acids (Punoševac et al., 2021). The notable dose-dependent decline in the activities of the hepatic enzymes AST, ALT, and ALP, along with serum cortisol levels, strongly indicates parsley's hepatoprotective and anti-stress properties. This is consistent with studies on other medicinal herbs, which attribute such effects to antioxidant compounds that stabilize cell membranes and prevent enzyme leakage (Abdel-Tawwab et al., 2024; Ghafarifarsani et al., 2022). The lack of significant change in LDH activity suggests a specific, rather than generalized, effect on metabolic pathways.
Conclusion: In conclusion, this study demonstrates that dietary supplementation with parsley powder, particularly at a 1% inclusion level, effectively enhances the physiological resilience of zebrafish to transportation-induced stress. The significant reductions observed in key stress and metabolic biomarkers, including cortisol, glucose, triglycerides, cholesterol, and hepatic enzymes, are attributed to the potent bioactive compounds in parsley. These compounds appear to act through multiple synergistic mechanisms, primarily by modulating the hypothalamic-pituitary-Interrenal (HPI) axis to mitigate the stress response, bolstering the antioxidant defense system to reduce oxidative damage, and improving hepatic lipid metabolism. Therefore, parsley presents itself as a viable natural feed additive to alleviate the adverse effects of aquaculture-related stressors. For future research, it is recommended to extend these investigations to other economically important fish species, explore potential synergistic effects with other medicinal herbs, and elucidate the precise molecular pathways involved.
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