Impact of sublethal concentrations of monosodium methylarsonate (MSMA) on zebrafish: Insights from embryotoxicity assessment and NMR-based metabolomics.

Organic arsenicals such as monosodium methylarsonate (MSMA) remain widely used, yet their mechanistic toxicity in aquatic systems is poorly understood. Here, we integrated fish embryo toxicity (FET) assays with 1H NMR-based metabolomics to characterize the effects of acute MSMA exposure in zebrafish (Danio rerio) embryos. The 96-h LC50 was 74 mg L-1, and embryos were exposed to sublethal concentrations (10, 30, and 60 mg L-1). Exposure induced concentration-dependent embryotoxicity, with severe phenotypic alterations at 60 mg L-1, including reduced survival (p = 0.0316), delayed hatching (p = 0.0331), tachycardia (p < 0.0001), and a reduction in some morphometric parameters, when compared with the control group. Total arsenic analysis confirmed significant bioaccumulation in larvae, reaching 1.34 ± 0.06, 6.51 ± 0.01, and 11.08 ± 0.22 mg kg-1 across increasing concentrations. Metabolomic profiling revealed marked metabolic reprogramming, and multivariate analyses (PCA, PLS-DA, and OPLS-DA) demonstrated clear separation between the control and exposed groups. Perturbations in energy metabolism were evidenced by decreased levels of lactate, glucose, and oxaloacetate, and increased levels of acetate, suggesting impaired glycolysis, mitochondrial dysfunction, and reduced fatty acid oxidation, while elevated glutathione levels indicated activation of antioxidant defenses. Pathway analysis highlighted disruptions in glycolysis/gluconeogenesis, glutathione metabolism, purine metabolism, and aminoacyl-tRNA biosynthesis. Four metabolites (xanthine, lactate, acetoacetate, and L-tryptophan) consistently discriminated between exposed groups, with high predictive performance (AUC = 0.967) supported by cross-validation and permutation testing (p = 0.001). Overall, these findings provide mechanistic insight into MSMA-induced toxicity and demonstrate the potential of metabolomics-derived biomarkers for environmental risk assessment, supporting more sensitive and integrative strategies for evaluating arsenical contamination in aquatic ecosystems and contributing to the development of improved environmental monitoring and regulatory frameworks for arsenical pollutants.