Introduction: Snake venoms are rich sources of bioactive peptides with therapeutic potential, particularly against neurodegenerative diseases linked to oxidative stress. While the peptide fraction (<10 kDa) from Bothrops jararaca venom has shown in vitro neuroprotection, analogous fractions from related species remain unexplored in vivo. Methods: This study comparatively evaluated the neuroprotective effects of two peptide fractions (pf) from Daboia siamensis (pf-Ds) and B. jararaca (pf-Bj) against H₂O₂-induced oxidative stress using in vitro (PC12 cells) and in vivo (zebrafish, Danio rerio) models. Results: In vitro, pf-Ds (1 µg mL⁻¹) did not protect PC12 cells against H₂O₂-induced cytotoxicity, unlike previously reported effects of pf-Bj. In vivo, neither pf-Ds nor pf-Bj (1–20 µg mL⁻¹) induced significant developmental toxicity in zebrafish larvae up to 120 h post-fertilization (hpf). The neuroprotective effects of both pf were evaluated using two experimental models: (I) Larvae at 96 hpf were exposed to either pf-Ds or pf-Bj (10 µg mL⁻¹) for 4 h, followed by co-exposure to H₂O₂ (0.2 mmol L⁻¹) for an additional 10 h to induce oxidative stress (4–20 h model); (II) Embryos at 4 hpf were treated with pf-Ds or pf-Bj (10 µg mL⁻¹) continuously until 96 hpf, after which they were exposed to H₂O₂ (0.2 mmol L⁻¹) for another 24 h (96–120 h model). In a short-term treatment model, neither fraction reversed H₂O₂-induced deficits in metabolism or locomotor activity. However, in a prolonged treatment model, pf-Bj significantly reversed the H₂O₂-induced locomotor impairment, whereas pf-Ds did not confer protection. Conclusions: These findings demonstrate, for the first time, the in vivo neuroprotective potential of pf-Bj against oxidative stress-induced behavioral deficits in zebrafish, contingent on the treatment regimen. The differential effects between pf-Ds and pf-Bj highlight species-specific venom composition and underscore the value of zebrafish for evaluating venom-derived peptides.

Functional characterization of peptide fraction (PF) from snake venom has provided novel opportunities to investigate possible neuroprotective compounds relevant to pharmaceuticals. This study was performed to investigate the PF-mediated neuroprotection obtained from Naja mandalayensis snake venom, a member of the Elapidae family, using two neuronal cell lines, undifferentiated PC12 and differentiated mHippoE-18, in response to H₂O₂-induced oxidative stress. Cells were pre-treated for 4 h with PF (10, 1, 0.01, and 0.001 μg mL⁻¹), and thereafter exposed to H₂O₂ (0.5 mmol L⁻¹) for 20 h. Then, the oxidative stress markers and label-free differential proteome strategy were analyzed to understand the neuroprotective effects of PF. In PC12 cells, PF showed no neuroprotective effects against oxidative stress. In mHippoE-18 cells, PF at 0.01 and 0.001 μg mL⁻¹ increased the viability and metabolism of cells against H₂O₂-induced neurotoxicity, reducing reactive oxygen species (ROS) generation. Interestingly, PF also exhibited a substantial reduction in baseline ROS levels compared to the control, indicating that PF could have compounds with antioxidant features. The comparative proteomic profiling identified 53 proteins with differential expression related to antioxidant action, catalysis, molecular function regulators, structural molecule activity, translation regulatory activity, ATP, and binding. The PF + H₂O₂ group indicated that protein expression is 6% upregulated, 4% downregulated, and 94% unchanged compared to the H₂O₂ group. Three significant proteins upregulated in the PF + H₂O₂ group, including elongation factor 2 (P58252), proteasome subunit alpha type (E9Q0X0), and E2 ubiquitin-conjugating enzyme (A0A338P786), suggested that PF-mediated neuroprotection happens through translational regulation and the degradation of defective proteins via the proteasome complex. Additionally, differential protein expression in PF changed the metabolism, protein synthesis, synaptic activity, and intracellular transport, suggesting that PF contains the rich mixture of bioactive peptides of interest pharmacologically. Overall, this study offers new opportunities for evaluating whether PF’s neuroprotective features in specific neuronal cells are maintained and to investigate neurodegenerative disease drug development processes.

Snake venoms are rich sources of bioactive peptides with therapeutic potential, particularly against neurodegenerative diseases linked to oxidative stress. While the peptide fraction (< 10 kDa) from Bothrops jararaca venom has shown in vitro neuroprotection, analogous fractions from related species remain unexplored in vivo. This study comparatively evaluated the neuroprotective effects of two peptide fractions (pf) from Daboia siamensis (pf-Ds) and B. jararaca (pf-Bj) against H₂O₂-induced oxidative stress using in vitro (PC12 cells) and in vivo (zebrafish, Danio rerio) models. In vitro, pf-Ds (1 µg⋅mL⁻¹) did not protect PC12 cells against H₂O₂-induced cytotoxicity, unlike previously reported effects of pf-Bj. In vivo, neither pf-Ds nor pf-Bj (1-20 µg⋅mL⁻¹) induced significant developmental toxicity in zebrafish larvae up to 120 hours post-fertilization (hpf). The neuroprotective effects of both pf were evaluated using two experimental models: (I) Larvae at 96 hpf were exposed to either pf-Ds or pf-Bj (10 µg⋅mL⁻¹) for 4 hours, followed by co-exposure to H₂O₂ (0.2 mmol⋅L⁻¹) for an additional 10 hours to induce oxidative stress (4–20 hours model); (II) Embryos at 4 hpf were treated with pf-Ds or pf-Bj (10 µg⋅mL⁻¹) continuously until 96 hpf, after which they were exposed to H₂O₂ (0.2 mmol⋅L⁻¹) for another 24 hours (96–120 hours model). In a short-term treatment model, neither fraction reversed H₂O₂-induced deficits in metabolism or locomotor activity. However, in a prolonged treatment model, pf-Bj significantly reversed the H₂O₂-induced locomotor impairment, whereas pf-Ds did not confer protection. These findings demonstrate, for the first time, the in vivo neuroprotective potential of pf-Bj against oxidative stress-induced behavioral deficits in zebrafish, contingent on the treatment regimen. The differential effects between pf-Ds and pf-Bj highlight species-specific venom composition and underscore the value of zebrafish for evaluating venom-derived peptides.