Common Herbicide Damages DNA and Stunts Growth of Toad Tadpoles at Tiny Doses

A commercial herbicide sprayed across the crop fields of Argentina’s Pampas can inflict measurable genetic damage, trigger oxidative stress, and slow the growth of toad tadpoles at concentrations far below the rates farmers actually apply, according to a new laboratory study published in Environmental Science and Pollution Research. The findings, reported by Celeste Ruiz de Arcaute, Milagros R. R. Laborde, Florencia Cardascia, and Sonia Soloneski of the National University of La Plata and CONICET, mark the first time the prometryn-based formulation Prometrex FW has been tested for sublethal effects on the Neotropical toad Rhinella arenarum, a species whose larvae share the shallow ponds and drainage channels that border intensively farmed land.

Prometryn belongs to the triazine family of herbicides, a chemical class that includes the widely discussed atrazine. Like its relatives, prometryn works by blocking photosystem II in plants, halting the light-driven reactions that weeds need to survive. It is used against broadleaf and grassy weeds in crops such as maize, vegetables, and citrus, and it is registered in Argentina under several commercial names, Prometrex FW among them. The formulation tested in the study contains 50 percent active ingredient, with the remainder made up of so-called inert adjuvants, substances added to help the product mix, spread, and stick to foliage. Those adjuvants, recent research suggests, can contribute their own toxicity, which is one reason ecotoxicologists increasingly test the full commercial product rather than the pure active compound alone.

The Argentine Pampas is one of the most intensively cultivated regions in South America, and agricultural intensification has steadily raised the burden of pesticides entering its waterways. Amphibians are considered particularly vulnerable sentinels in this landscape. Their permeable skin, aquatic eggs, and free-swimming larvae place them in direct and prolonged contact with whatever runs off the surrounding fields. Rhinella arenarum, the common South American toad, is broadly distributed and relatively easy to rear in the laboratory, which has made it a standard model for ecotoxicology in the region for decades. Yet despite the long history of testing other herbicides on this species, no one had previously examined how a prometryn-based product affects its larvae.

To fill that gap, the research team exposed premetamorphic tadpoles to three concentrations of Prometrex FW: 0.0015, 0.015, and 15 milligrams of active ingredient per liter. The lowest dose was chosen to reflect concentrations that could plausibly be found in the environment, while the highest, although dramatically lower than the manufacturer’s recommended application rate, was included to probe the upper range of acute sublethal exposure. Larvae were kept in the solutions for two exposure periods, 48 hours and 96 hours, under controlled laboratory conditions and in accordance with internationally recognized guidelines for acute toxicity testing with amphibians. The experimental protocol was reviewed and approved by the institutional animal care committee at the National University of La Plata.

The most striking result came from the alkaline comet assay, a technique that detects strand breaks in the DNA of individual cells. Cells are embedded in a gel, subjected to electrophoresis, and stained so that damaged DNA migrates away from the nucleus, forming the comet-like tail that gives the method its name. In the treated tadpoles, the genetic damage index rose significantly at every concentration tested, including the lowest, environmentally relevant dose. In other words, there was no observable threshold below which the herbicide formulation stopped harming DNA within the range examined. Because unrepaired DNA damage can propagate as mutations, interfere with cell division, and undermine development, this result carries weight well beyond the laboratory dish.

The biochemical assays told a consistent story. The researchers measured the activity of catalase, an enzyme that decomposes hydrogen peroxide, and glutathione S-transferase, a family of enzymes that conjugate toxic compounds to the cellular antioxidant glutathione so they can be excreted. They also quantified the total glutathione content of the larvae. All three biomarkers were significantly elevated in exposed animals, a pattern the authors interpret as evidence of oxidative stress. When herbicides or their breakdown products enter cells, they can generate reactive oxygen species faster than the normal antioxidant defenses can neutralize them. The tadpoles’ systems responded by ramping up production of catalase and GST and by mobilizing glutathione, effectively fighting a biochemical fire on two fronts. Sustained oxidative stress of this kind is linked in the broader literature to lipid peroxidation, protein damage, and the activation of apoptotic cell death pathways.

Growth told its own grim tale. Across all treatments and both exposure durations, larval growth was significantly inhibited compared with controls. For a tadpole, growth is not merely a matter of getting bigger; it is a race against pond drying, predation, and the physiological demands of metamorphosis. Larvae that fall behind face longer windows of vulnerability in the water, and population-level consequences can follow if enough individuals fail to reach metamorphosis in time. The combination of DNA damage, oxidative stress, and stunted development in the same animals suggests that the formulation attacks larval health at several levels simultaneously, from the molecular to the whole organism.

The study does not stand in isolation. Earlier work has documented prometryn’s capacity to disrupt aquatic life in other species. In zebrafish, prometryn exposure has been associated with mitochondrial dysfunction, oxidative stress, and failed organogenesis during early development, while in marine medaka it induces developmental toxicity at environmentally relevant concentrations. In crustaceans such as the Chinese mitten crab, the compound provokes oxidative damage and apoptosis in the hepatopancreas. Fish studies, including work on common carp and goldfish exposed to prometryn-containing products, have reported disturbances of antioxidant defenses and blood parameters. The new findings extend this picture to a Neotropical amphibian and, crucially, to a complete commercial formulation rather than the purified active ingredient.

Why does the distinction between active ingredient and formulation matter so much? Adjuvants and solvents in commercial products can enhance the penetration of the active compound through biological membranes, including the skin of amphibians, and some adjuvants are themselves toxic. Screening studies using molecular dynamics simulation and machine learning have begun to catalog the environmental hazards posed by pesticide adjuvants, an area historically exempt from the rigorous testing applied to active ingredients. By testing Prometrex FW as sold and sprayed, the Argentine team captured the real-world chemical mixture to which pond-dwelling larvae would actually be exposed after agricultural application, rather than an idealized single molecule.

The authors are careful about the scope of their conclusions. The work was conducted under laboratory conditions with acute exposures, and the researchers note that data supporting the findings are available from the corresponding author upon request. Nevertheless, the convergence of effects at low concentrations, including one chosen for its environmental relevance, leads them to flag a potential ecological risk to non-target species. In the Pampas, where drainage canals, temporary ponds, and crop fields interpenetrate, the boundary between treated land and amphibian habitat is thin. The study, funded by the National University of La Plata, CONICET, and Argentina’s National Agency of Scientific and Technological Promotion, adds prometryn-based formulations to the growing list of agricultural chemicals that can injure native wildlife at doses regulators and applicators may assume are harmless, and it underscores the case for monitoring surface waters in farming regions with the same urgency applied to the crops themselves.

Subject of Research: Sublethal genotoxic and oxidative effects of a prometryn-based herbicide formulation on Rhinella arenarum tadpoles

Article Title: Biochemical and morphological alterations in Rhinella arenarum larvae exposed to the prometryn-based formulation Prometrex FW®

Article References: Ruiz de Arcaute, C., Laborde, M. R. R., Cardascia, F., & Soloneski, S. (2026). Biochemical and morphological alterations in Rhinella arenarum larvae exposed to the prometryn-based formulation Prometrex FW®. Environmental Science and Pollution Research, 33(28), 14590-14604. https://doi.org/10.1007/s11356-026-38194-w

Image Credits: AI Generated

DOI: 10.1007/s11356-026-38194-w

Keywords: prometryn, Rhinella arenarum, amphibian toxicology, comet assay, DNA damage, oxidative stress, catalase, glutathione S-transferase, herbicide formulation, ecotoxicology, Argentine Pampas, tadpole growth