Famoxadone is a chiral fungicide frequently found in the environment and agricultural products. However, the health risks of famoxadone enantiomers are not well understood. This study investigated the stereoselective cytotoxicity and metabolic behavior of famoxadone enantiomers in mammals. Results showed that R-famoxadone was 1.5 times more toxic to HepG2 cells than S-famoxadone. R-famoxadone induced more pronounced ferroptosis compared to S-famoxadone. It caused greater upregulation of genes related to iron transport and lipid peroxidation, and greater downregulation of genes related to peroxide clearance. Furthermore, R-famoxadone induced more severe lipid peroxidation and reactive oxygen species (ROS) accumulation through ACSL4 activation and GPX4 inhibition. Additionally, the bioavailability of R-famoxadone in mice was six times higher than that of S-famoxadone. Liver microsome assays, cytochrome P450 (CYP450) inhibition assays, human recombinant CYP450 assays, and molecular docking suggested that the lower binding affinities of CYP2C8, CYP2C19, and CYP2E1 for R-famoxadone caused its preferential accumulation. Overall, R-famoxadone poses a higher risk than S-famoxadone due to its greater cytotoxicity and persistence. This study provides the first evidence of ferroptosis-induced stereoselective toxicity, offering insights for the comprehensive health risk assessment of chiral famoxadone and valuable references for the application of high-efficiency, low-risk pesticide enantiomers.

In recent years, the awareness that pesticides can have other effects apart from generic toxicity is growing. In particular, several pieces of evidence highlight their influence on human fertility. In this study, we investigated, by a virtual screening approach, the binding between pesticides and proteins present in human gametes or associated with reproduction, in order to identify new interactions that could affect human fertility. To this aim, we prepared ligand (pesticides) and receptor (proteins) 3D structure datasets from online structural databases (such as PubChem and RCSB), and performed a virtual screening analysis using Autodock Vina. In the comparison of the predicted interactions, we found that famoxadone was predicted to bind Cellular Retinol Binding Protein-III in the retinol-binding site with a better minimum energy value of -10.4 Kcal/mol and an RMSD of 3.77 with respect to retinol (-7.1 Kcal/mol). In addition to a similar network of interactions, famoxadone binding is more stabilized by additional hydrophobic patches including L20, V29, A33, F57, L117, and L118 amino acid residues and hydrogen bonds with Y19 and K40. These results support a possible competitive effect of famoxadone on retinol binding with impacts on the ability of developing the cardiac tissue, in accordance with the literature data on zebrafish embryos. Moreover, famoxadone binds, with a minimum energy value between -8.3 and -8.0 Kcal/mol, to the IZUMO Sperm-Egg Fusion Protein, interacting with a network of polar and hydrophobic amino acid residues in the cavity between the 4HB and Ig-like domains. This binding is more stabilized by a predicted hydrogen bond with the N185 residue of the protein. A hindrance in this position can probably affect the conformational change for JUNO binding, avoiding the gamete membrane fusion to form the zygote. This work opens new interesting perspectives of study on the effects of pesticides on fertility, extending the knowledge to other typologies of interaction which can affect different steps of the reproductive process.

Thyroid dysfunction has become a serious public health problem, which is considered a trigger of nonalcoholic fatty liver disease (NAFLD). Pesticide exposure could contribute to thyroid dysfunction and NAFLD, but the relationship between these factors remains unclear. In this study, the effects of subchronic famoxadone exposure on thyroid and liver at no observed adverse effect level (NOEL) related concentrations were investigated using in vivo, in vitro, and in silico models. Famoxadone caused hepatic steatosis, lipid metabolism disorder, and liver oxidative stress and induced NAFLD in male mice. The suppression of hepatic fatty acid β-oxidation was the key factor of NAFLD, which was highly associated with hypothalamic-pituitary-thyroid (HPT) axis hormones disorder. Famoxadone disrupted thyroid hormone biosynthesis by causing thyroid follicle aberrations and abnormal HPT axis-related gene expression. In vitro studies confirmed that famoxadone inhibited the transport of thyroxine (T4) into hepatocytes and the conversion of T4 to triiodothyronine (T3). In silico studies verified that famoxadone interfered with the binding of thyroid hormones to proteins mediating thyroid hormone transport, conversion, and activation. This study comprehensively reported the association between NAFLD and thyroid dysfunction caused by famoxadone, providing new perspectives for the health risk evaluation of pesticides with a similar structure in mammals.

In the present study, we conducted optimization of pyramoxadone and synthesized a series of novel oxazolidinediones. Antifungal assays showed that these compounds exhibited moderate to excellent antifungal activity against various pathogens. Further SAR analysis revealed that the introduction of substituents to the benzene ring of the phenoxy group or the inclusion of bulky groups, such as tert-butyl, on the aniline moiety, had a detrimental effect on the activity. However, the inclusion of fluorine atoms in the aniline moiety significantly enhanced the antifungal efficacy. Notably, compound 2-4 displayed significantly higher activity compared to both pyramoxadone and famoxadone against R. solani, B. cinerea, S. sclerotiorum, and P. oryzae, where it demonstrated EC50 values of 1.78, 2.47, 2.33, and 2.23 μg/mL, respectively. Furthermore, compound 2-4 exhibited potent protective and curative effects against the tomato gray mold in vivo. A mechanistic investigation revealed that compound 2-4 significantly impacted the mycelial morphology, inhibited spore germination, and impeded mycelial respiration, ultimately leading to the inhibition of pathogenic fungus growth. These findings indicate that compound 2-4 has the potential to serve as a cyt bc1 inhibitor and should be further investigated for development.

In accordance with Article 43 of Regulation (EC) No 396/2005, the European Commission requested EFSA to assess whether existing Codex Maximum Residue Limits (CXLs) for famoxadone are safe for consumers in light of the lowered toxicological reference values established following the non-renewal of approval of the active substance famoxadone. Based on the targeted assessment EFSA identified a potential acute concern for the CXL on table grapes. For the other CXLs consumers intake concern was not identified.

The uptake, translocation, and subcellular distribution of oxathiapiprolin and famoxadone in tomato plants were investigated using hydroponic experiments. Oxathiapiprolin and famoxadone mainly accumulated in the tomato roots with limited translocation capacity from the roots to the upper part. The root absorption and inhibitor results noted the dominance of the apoplastic and symplastic pathways in the oxathiapiprolin and famoxadone uptake by the tomato roots, respectively. Furthermore, the uptake process for the two fungicides followed passive and aquaporin-dependent transport. Insoluble cell components (cell organelles and walls) were the dominant storage compartments for oxathiapiprolin and famoxadone. In the protoplast, oxathiapiprolin in the soluble fraction had a higher proportion than that of famoxadone. Finally, the uptake and distribution of the two fungicides by the tomato plants was accurately predicted using a partition-limited model. Thus, this study provides an in-depth understanding of the transfer of oxathiapiprolin and famoxadone from the environment to tomato plants.

In this study, the stereoselective bioactivity, acute toxicity, and environmental fate for famoxadone enantiomers were reported for the first time. Five representative pathogens (e.g., Alternaria solani) were used to investigate enantioselective activity, and three non-target organisms (e.g., Selenastrum bibraianum) were used to evaluate acute toxicity. S-Famoxadone was 3.00-6.59 times more effective than R-famoxadone. R-Famoxadone also showed 1.80-6.40 times more toxicity than S-famoxadone toward S. bibraianum and Daphnia magna. The toxicity of R-famoxadone was 100 times more toxic than S-famoxadone toward Danio rerio. Under aerobic conditions, the half-life (t1/2) for famoxadone enantiomer degradation was 46.2-126 days in different soils and the enantiomeric fraction (EF) ranged from 0.435 to 0.470 after 120 days. R-Famoxadone preferentially degraded in three soils, resulting in an enrichment of S-famoxadone. Under anaerobic conditions, t1/2 of famoxadone enantiomers was 62.4-147 days in different soils and the EF ranged from 0.489 to 0.495, indicating that famoxadone enantiomers were not enantioselective. This study will be useful for the environmental and health risk assessments for famoxadone enantiomers.

Famoxadone enantiomers were separated on Lux Amylose-1 chiral column and determined by ultra-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS). The half-lives of R-(-)-famoxadone and S-(+)-famoxadone were 69.3 and 86.6 h in apple cider, 231.0 and 346.5 h in apple pomace, 69.3 and 77.0 h in grape wine, and 231.0 and 346.5 h in grape pomace, respectively. The enantiomeric fraction (EF) values decreased gradually from 0.498, 0.499, and 0.500 (0 h) to 0.404, 0.374, and 0.427 (144 h) and then increased gradually to 0.474, 0.427, and 0.422 (312 h) in apple cider, grape wine, and grape pomace. The EF value in apple pomace decreased gradually from 0.499 (0 h) to 0.450 (168 h) and then increased gradually to 0.482 (312 h). The processing factors (PFs) for famoxadone ranged from 0.014 to 0.024 in the overall process. The residue of famoxadone reduced 94.7-97.4% after the fermentation process.

The chiral pesticide famoxadone is mainly applied to control fungal diseases on fruiting vegetables. The fungicidal activity, ecotoxicological effects, and degradation behavior of famoxadone enantiomers are less well known. In this study, a systemic assessment of the stereoselectivity of famoxadone was performed in cucurbits and soil. Famoxadone enantiomers presented distinct inhibitory activities among different fungal species. The bioactivities of R-(-)-famoxadone were 2.7-178 times higher than S-(+)-famoxadone toward five phytopathogens. Based on the obtained LC50 values, famoxadone was super toxic to Eisenia foetida (E. foetida). Moreover, the acute toxicity of R-(-)-famoxadone presented 167 times greater to E. foetida than that of S-(+)-famoxadone, indicating that R-(-)-famoxadone showed higher bioactivity toward target organisms and non-target organisms than S-(+)-famoxadone. In addition, a simple high-performance liquid chromatography (HPLC) method was established to determine the stereoselective degradation of famoxadone in two species of cucurbits (cucumber and chieh-qua) and in field soil. The half-life values of famoxadone degradation were from 5.4 to 14.1 days, indicating that famoxadone was easily degraded. Additionally, no stereoselective degradation was found in cucurbits and soil. The results may provide promising implications for comprehensive environmental and ecological risk assessments of famoxadone.

The dissipation of famoxadone as well as the behaviour of its metabolites in environmental samples such as water and soil is a major concern. In this study, the dissipation of the target compound in both matrices was carried out applying an analytical method based on ultra-high performance liquid chromatography coupled to Orbitrap mass spectrometry (UHPLC-Orbitrap-MS). The dissipation of famoxadone was monitored over a period of 100 days after the plant protection product, Equation Pro®, was administered to the target matrices. This study was performed at two doses, normal and double in the case of soils and fivefold instead of double dose in water. The concentration of famoxadone steadily decreased during the monitoring period in both matrices. Half-life (DT50) values were lower than 30 days in most cases except for loam soils, for which it was 35 days. Therefore, persistence of this pesticide in both matrices was low. Famoxadone metabolites such as IN-KF015 ((5RS)-5-methyl-5-(4-phenoxyphenyl)-1,3- oxazolidine-2,4-dione) and IN-JS940 ((2RS)-2-hydroxy-2-(4- phenoxyphenyl)propanoic acid) were detected in both matrices and their concentration increased while the concentration of the parent compound decreased. Metabolite IN-JS940 was the compound detected at highest concentration for both matrices. In water the maximum concentration was 20% of the initial famoxadone content and in soils it was 50% of initial famoxadone content. In addition, another metabolite, IN-MN467 ((5RS)-5-methyl-3-[(2-nitrophenyl)amino]- 5-(4-phenoxyphenyl)-1,3-oxazolidine-2,4-dione), was detected in soils, following the same behaviour as the other metabolites. These results provided ample information about the behaviour of metabolites and the necessity of knowing their toxicity in both matrices in order to detect possible risks for living beings.