PFASs are highly persistent in both natural and living environment, and pose a significant risk for wildlife and human beings. The present study was carried out to determine the inhibitory behaviours of fourteen PFASs on metabolic activity of two major isoforms of carboxylesterases (CES). The probe substrates 2-(2-benzoyl-3-methoxyphenyl) benzothiazole (BMBT) for CES1 and fluorescein diacetate (FD) for CES2 were utilized to determine the inhibitory potentials of PFASs on CES in vitro. The results demonstrated that perfluorododecanoic acid (PFDoA), perfluorotetradecanoic acid (PFTA) and perfluorooctadecanoic acid (PFOcDA) strongly inhibited CES1 and CES2. The half inhibition concentration (IC₅₀) value of PFDoA, PFTA and PFOcDA for CES1 inhibition was 10.6 μM, 13.4 μM and 12.6 μM, respectively. The IC₅₀ for the inhibition of PFDoA, PFTA and PFOcDA towards CES2 were calculated to be 9.56 μM, 17.2 μM and 8.73 μM, respectively. PFDoA, PFTA and PFOcDA exhibited noncompetitive inhibition towards both CES1 and CES2. The inhibition kinetics parameters (Kᵢ) were 27.7 μM, 26.9 μM, 11.9 μM, 4.04 μM, 29.1 μM, 27.4 μM for PFDoA-CES1, PFTA-CES1, PFOcDA-CES1, PFDoA-CES2, PFTA-CES2, PFOcDA-CES2, respectively. In vitro-in vivo extrapolation (IVIVE) predicted that when the plasma concentrations of PFDoA, PFTA and PFOcDA were greater than 2.77 μM, 2.69 μM and 1.19 μM, respectively, it might interfere with the metabolic reaction catalyzed by CES1 in vivo; when the plasma concentrations of PFDoA, PFTA and PFOcDA were greater than 0.40 μM, 2.91 μM, 2.74 μM, it might interfere with the metabolic reaction catalyzed by CES2 in vivo. Molecular docking was used to explore the interactions between PFASs and CES. In conclusion, PFASs were found to cause inhibitory effects on CES in vitro, and this finding would provide an important experimental basis for further in vivo testing of PFASs focused on CES inhibition endpoints.

Penconazole is one of the most widely used fungicides all over the world, and since it spreads to large environments, its toxic effects on non-target organisms are of great concern. The toxic effects of penconazole on crayfish (Astacus leptodactylus), which is a bioindicator in freshwater ecosystems and consumed economically, are not known. Therefore, in this study, the purpose was to contribute to the literature on the potential harmful effects of penconazole on a non-target species, Astacus leptodactylus. For this aim, the acute toxicity (96 h) of penconazole was examined. The 96-h LC₅₀ value of penconazole was detected as 18.7 mg L⁻¹. Four concentrations of penconazole (18.7 mg L⁻¹, 9.35 mg L⁻¹, 4.68 mg L⁻¹, 2.34 mg L⁻¹) were applied to crayfish for 96 h. The results showed that penconazole had destructive effects on esterase mechanisms by inhibiting acetylcholinesterase (AChE) and carboxylesterase (CaE) activities. Significant increases were observed in all antioxidant parameters (superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR), glutathione S-transferase (GST), reduced glutathione (GSH), malondialdehyde (MDA)) in all doses except the lowest concentration (2.34 mg L⁻¹). All adenosine triphosphatase (ATPase) activities (Na⁺/K⁺-ATPase, Mg²⁺-ATPase, Ca²⁺-ATPase, total ATPase) had significant dose-related inhibition in both gill and muscle tissues. In summary, our findings show that acute penconazole administration to crayfish causes significant toxic effects on esterase, antioxidative parameters, and metabolic enzymes.

Mussels are worldwide bioindicators in pollution monitoring since they fulfil the requirements for being good sentinels. However, some methodological concerns arise in the use of particular biomarkers, particularly those displaying low enzymatic rates and/or limited responsiveness to chemicals and biological-related variability. In the present study, the suitability of oxidative stress and detoxification parameters when using mussels as sentinels of polycyclic aromatic hydrocarbon (PAH) pollution is addressed. Present results show that the S9 subcellular fraction of the digestive gland in mussels is an adequate and convenient matrix where to measure most pollution-related biomarkers. Furthermore, this work constitutes the first evidence of the potential suitability of using particular carboxylesterase (CE) activities in determining PAHs exposure in mussels. This fact could imply the replacement of more controversial cytochrome P450 components (phase I oxidation), which are only measurable in microsomal fractions, by CEs (measured in S9 fractions) as good alternatives for phase I reactions in PAH-exposed mussels. Some methodological considerations, such as the need of including commercial purified proteins in biomarker determinations for quality assurance, are evaluated.

The toxicological knowledge of mancozeb (MZ)-containing commercial formulations on non-target species is scarce and limited. Therefore, the objective of this work was to represent a realistic application scenario by evaluating the toxicity of environmental relevant and higher concentrations of a commercial formulation of MZ using zebrafish embryos. Following determination of the 96-h LC₅₀ value, the embryos at the blastula stage (~ 2 h post-fertilisation, hpf) were exposed to 0.5, 5, and 50 μg L⁻¹ of the active ingredient (~ 40× lower than the 96-h LC₅₀). During the exposure period (96 h), lethal, sublethal, and teratogenic parameters, as well as behaviour analysis, at 120 hpf, were assayed. Biochemical parameters such as oxidative stress–linked enzymes (superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR)), reactive oxygen species (ROS) levels, and glutathione levels (GSH and GSSG), as well as the activity of degradation (glutathione S-transferase (GST) and carboxylesterase (CarE)), neurotransmission (acetylcholinesterase (AChE)), and anaerobic respiration (lactate dehydrogenase (LDH))–related enzymes, were analysed at the end of the exposure period. Exposed embryos showed a marked decrease in the hatching rate and many malformations (cardiac and yolk sac oedema and spinal torsions), with a higher prevalence at the highest concentration. A dose-dependent decreased locomotor activity and a response to an aversive stimulus, as well as a light-dark transition decline, were observed at environmental relevant concentrations. Furthermore, the activities of SOD and GR increased while the activity of GST, AChE, and MDA contents decreased. Taken together, the involvement of mancozeb metabolites and the generation of ROS are suggested as responsible for the developmental phenotypes. While further studies are needed to fully support the hypothesis presented, the potential cumulative effects of mancozeb-containing formulations and its metabolites could represent an environmental risk which should not be disregarded.

Itol A, an isoryanodane diterpene derived from Itoa orientalis Hemsl. (Flacourtiaceae), is a potential plant-based insecticide. However, the effect of itol A on the tobacco cutworm [Spodoptera litura (Fab.) (Lepidoptera: Noctuidae)], an important and widely distributed insect pest, remains unclear. In this study, the toxicity and inhibitory potency of itol A on S. litura were evaluated. The results indicated that itol A exhibited larvicidal activity against the third instar larvae in a concentration-dependent manner (LC₅₀ 875.48 mg/L at 96 h). Antifeedant activity also was observed, and the 24-h AFC₅₀ values were 562.05 and 81.47 mg/L in the no-choice and choice experiments, respectively. The insect growth was inhibited after treatment of itol A, as reflected by long developmental periods, low-quality pupae, and various abnormalities. Itol A exerted ovicidal effect on S. litura, with an estimated LC₅₀ of 759.30 mg/L. Itol A deterred oviposition in the choice experiment (ODI₅₀ 909.60 mg/L). Besides, the activities of α-amylase, general protease, superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) were inhibited after itol A treatment over time compared to controls, which may be a relevant mechanism underlying the toxicity of itol A toward S. litura. However, the activities of lipase, carboxylesterase (CarE), glutathione S-transferase (GST), and cytochrome P450 monooxygenase (P450) were increased. Taken together, these results suggest that itol A could be a good botanical pesticide to reduce the population of S. litura in integrated pest management programs.