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Biosynthesis of silver nanoparticles has received considerable attention due to their cost-effective, eco-friendly and medicinal values. In this study, silver nanoparticles (Ag NPs) were synthesised using the aqueous leaf extracts of Piper nigrum. TEM images revealed that the particle is spherical with 20–50 nm in size. Furthermore, to evaluate the toxicity of synthesized Ag NPs, fish Labeo rohita were exposed to two different concentrations (2.5 µg/L as the treatment I and 5 µg/L as treatment II) for 35 days, and antioxidant parameters and histology of gill, liver and kidney were examined. A biphasic response in the activity of glutathione S-transferases (GST) was observed in gill and liver of fish. GST activity in the kidney of fish was significantly increased when compared to control group. Glutathione reductase (GR) activity in organs/tissue of fish were found to be increased while peroxidase (POD) activity was significantly decreased. Histopathological changes such as hyperplasia, proliferation of epithelial cells and fusion of lamellae were observed in both the concentrations. In liver, necrosis, nuclear degeneration and dilation of sinusoids were observed. Subsequently, the representative effects of POD activity were assessed based on the Box–Behnken Equation, 3-D contour plot and ANOVA analysis through response surface methodology analysis. [ABSTRACT FROM AUTHOR]