Combined effects of cadmium and sulfamethoxazole on Eisenia fetida: Insights into accumulation, subcellular partitioning, biomarkers and toxicological responses.

Cadmium (Cd) and sulfamethoxazole (SMX) frequently coexist in farmlands, yet their synergistic toxicological impacts on terrestrial invertebrates remain unexplored. In this study, earthworms were exposed to artificial soils percolated with Cd (5 mg/kg), SMX (5 mg/kg) or combination of them for 7 days, followed by a 12-day elimination phase in uncontaminated soil. The uptake of Cd and SMX by the earthworms, along with their subcellular distribution, was meticulously analyzed. Additionally, a suite of biomarkers-including superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA), and weight loss-were evaluated to assess the health status of the earthworms and the toxicological effects of the Cd and SMX mixture. Notably, the cotreatment with Cd and SMX resulted in a significantly higher weight loss in Eisenia fetida (41.25 %) compared to exposure to Cd alone (26.84 %). Moreover, the cotreatment group exhibited substantially higher concentrations of Cd in the total internal body, fraction C (cytosol), and fraction E (tissue fragments and cell membranes) in Eisenia fetida compared to Cd alone counterparts. The combined exposure also significantly elevated the SMX levels in the total body and fraction C compared with the SMX-only treated earthworms. Additionally, Eisenia fetida subjected to the combined treatment showed markedly increased activities of SOD, CAT, and MDA compared to those treated with Cd alone. The effect addition indices (EAIs), ranging from 1.00 to 2.23, unequivocally demonstrated a synergistic effect of the combined treatments. Interestingly, relocating the earthworms to clean soil did not mitigate the observed adverse effects. These findings underscore the increased risk posed by the Cd-SMX complex to terrestrial invertebrates in agricultural areas.

Cyclodextrin-based ternary supramolecular deep eutectic solvents for efficient extraction and analysis of trace quinolones and sulfonamides in wastewater by adjusting pH.

BACKGROUND: Antibiotics residues can accelerate the growth of drug-resistant bacteria and harm the ecological environment. Under the effect of enrichment and biomagnification, the emergence of drug-resistant pathogenic bacteria may eventually lead to humans being ineffective to drugs in the face of bacterial or fungal disease infections in the future. It is urgent to develop an efficient separation medium and analytical method for simultaneous extraction and determination of antibiotics in the water environment. RESULTS: This work doped 2,6-Di-O-methyl-ß-cyclodextrin, randomly methyl-ß-cyclodextrin, 2-hydroxypropyl-ß-cyclodextrin with thymol:fatty acid respectively to construct non-covalent interaction-dominated pH-responsive ternary supramolecular deep eutectic solvents (SUPRADESs), which can undergo a hydrophilic/hydrophobic transition with aqueous phase to achieve an efficient microextraction. Semi-empirical method illustrated that SUPRADESs have a wide range of hydrogen bond receptor sites. We developed a SUPRADES-based analytical method combined with liquid chromatography-triple quadrupole mass spectrometry for the extraction and determination of trace quinolones and sulfonamides in wastewater. The overall limits of detection of the method were 0.0021-0.0334 ng mL-1 and the limits of quantification were 0.0073-0.1114 ng mL-1. The linearity maintained good in the spiked level of 0.01-100 ng mL-1 (R2 > 0.99). The overall enrichment factors of the method were 157-201 with lower standard deviations (≤8.7). SIGNIFICANCE: The method gave an extraction recovery of 70.1-115.3 % for 28 antibiotics in livestock farming wastewater samples from Zhejiang, China, at trace levels (minimum 0.5 ng mL-1). The results demonstrated that inducing the phase transition between SUPRADES and aqueous phase by adjusting pH for extraction is a novel and efficient pretreatment strategy. To our knowledge, this is the first application of cyclodextrin-based ternary SUPRADESs with pH-responsive reversible hydrophobicity-hydrophilicity transition behavior in wastewater analysis.

Stereoselective bioaccumulation and degradation of chiral pesticide hexythiazox in earthworm-soil microcosm.

The chiral pesticide hexythiazox was extensively employed in agricultural activities and has garnered growing concern for its harmful impact on the ecosystem. This study investigates the toxicodynamic earthworm at the enantiomeric level of hexythiazox. Earthworms exhibited notable enantioselectivity during the accumulation stage. Furthermore, the presence of earthworms can impact the rate of degradation and enantioselectivity of hexythiazox in soil. The accumulation of the two hexythiazox enantiomers in the earthworm adhered to the one-compartment model, whereas the elimination phase was governed by the first-order kinetics equation. Furthermore, it was discovered that there was no notable enantioselectivity observed during the elimination phase.

Impacts of prothioconazole and prothioconazole-desthio on bile acid and glucolipid metabolism: Upregulation of CYP7A1 expression in HepG2 cells.

As an efficient triazole fungicide, prothioconazole (PTC) is widely used for the prevention and control of plant fungal pathogens. It was reported that the residues of PTC and prothioconazole-desthio (PTC-d) have been detected in the environment and crops, and the effects of PTC-d may be higher than that of PTC. Currently, PTC and PTC-d have been proven to induce hepatic metabolic disorders. However, their toxic effects on cellular bile acid (BA) and glucolipid metabolism remain unknown. In this study, HepG2 cells were exposed to 1-500 µM of PTC or PTC-d. High concentrations of PTC and PTC-d were found to induce cytotoxicity; thus, subsequent experimental exposure was conducted at concentrations of 10-50 µM. The expression levels of CYP7A1 and TG synthesis-related genes and levels of TG and total BA were observed to increase in HepG2 cells. Molecular docking analysis revealed direct interactions between PTC or PTC-d and CYP7A1 protein. To further investigate the underlying mechanisms, PTC and PTC-d were treated to HepG2 cells in which CYP7A1 expression was knocked down using siCYP7A1. It was observed that PTC and PTC-d affected the BA metabolism process and regulated the glycolipid metabolism process by promoting the expression of CYP7A1. In summary, we comprehensively analyzed the effects and mechanisms of PTC and PTC-d on cellular metabolism in HepG2 cells, providing theoretical data for evaluating the safety and potential risks associated with these substances.

Mefentrifluconazole exposure disrupted hepatic lipid metabolism disorder tightly associated with gut barrier function abnormal in mice.

Mefentrifluconazole (MFZ) is an azole fungicide that is placed in agriculture for the control of fungal hazards. However, due to their non-biodegradability, azole fungicides can accumulate in plants, animals, and the environment, thus becoming a major health concern worldwide. In this study, we exposed 7-week-old C57BL/6 mice to 10, 30, and 100 mg/kg of MFZ for 28 d to assess the toxic effects of MFZ on the liver and gut tissues of the mice. Histopathological, biochemical indexes, and transcriptomic analyses revealed that MFZ exposure disrupted the liver structure and hepatic lipid metabolism as well as damaged gut barrier function and promoted inflammation in mice. Moreover, 16S rRNA sequencing demonstrated that MFZ exposure significantly increased the abundance of patescibacteria at the generic level. Also, MFZ exposure increased the abundance of bacterial genera associated with inhibition of glycolipid metabolism. These results suggested that the disruption of liver lipid metabolism caused by MFZ exposure may be caused by changes in gut microbiota function. This study provided a new disease occurrence study for risk assessment of MFZ and strengthened the focus on some novel fungicides.

The Dissipation Behavior and Risk Assessment of Carbendazim Under Individual and Joint Applications on Peach (Amygdalus persica L.).

Dissipation, residue levels, and ingestion risks of carbendazim in peach (Amygdalus persica L.) were investigated with individual and joint applications in the present study. The dissipation kinetics of carbendazim, chlorpyrifos, prochloraz, and imidacloprid were evaluated by the first-order kinetics. When carbendazim was individually applied, the final residual concentration was 2.97 mg kg-1 and the half-life was 17.4 d. In the joint application of carbendazim with chlorpyrifos, prochloraz, and imidacloprid, the residual concentrations at 35 d after spraying were 7.16, 7.50, and 4.26 mg kg-1 and the half-lives were 30.8, 23.7, and 23.2 d, respectively, which showed an increase of 1.3-1.8 times compared with the single application of carbendazim. In addition, the effects of household processing of rinsing and peeling were investigated, and a high removal rate of 54.6% and 76.5% were found. Furthermore, the carbendazim ingestion risk assessment was conducted, which indicated that the acute health risk (aHI) and hazard quotient (HQ) of carbendazim were all within acceptable levels ranging from 21.7% to 40.9%. However, a higher ingestion risk of carbendazim was found under the joint application. This study provides some preliminary guidance for the joint application and risk assessment of carbendazim in peach, which is worth further investigation.

Exposure to the fungicide prothioconazole and its metabolite prothioconazole-desthio induced hepatic metabolism disorder and oxidative stress in mice.

Prothioconazole (PTC), as a popular triazole fungicide, with its main metabolite prothioconazole desthio (PTC-d), have attracted widespread concern due to their widely use and toxicological effects on non-target organisms. However, toxic effects of study analyzed PTC and PTC-d on the hepatic metabolism of mammalian still remains unclear. In this study, we conducted the study of the C57BL/6 mice which oral exposure to 30 mg/kg PTC and PTC-d via metabolomic analysis. In the liver, the metabolomics profile unveiled that exposure to 30 mg/kg PTC and PTC-d led to significantly altered 13 and 28 metabolites respectively, with 6 metabolites in common including significant decreased d-Fructose, Glutathione, showing the change of carbohydrate, lipid and amino acid metabolism. Via the further exploration of genes related to hepatic glycolipid metabolism and the biomarkers of oxidative stress, we found that liver was potentially damaged after exposure to 5 and 30 mg/kg PTC and PTC-d. Particularly, it was proved that PTC-d caused more adverse effect than its parent compound PTC on hepatotoxicity, and high concentration PTC or PTC-d exposure is more harmful than low concentration exposure.

In-syringe dispersive solid phase filter extraction cleanup followed by liquid chromatography-triple quadrupole mass spectrometry for fast determination of colchicine in plasma/urine.

As an effective treatment for acute gouty arthritis and cardiovascular disease, colchicine is also a toxic alkaloid and may cause poisoning or even death in overdose. The study of colchicine elimination and the diagnosis of poisoning etiology need the rapid and accurate quantitative analysis method in biological matrix. An analytical method was developed for colchicine in plasma and urine by in-syringe dispersive solid phase extraction (DSPE) followed by liquid chromatography-triple quadrupole mass spectrometry (LC-MS/MS). Sample extraction and protein precipitation were proceeded with acetonitrile. The extract was cleaned by in-syringe DSPE. An XBridge™ BEH C18 column（100 mm × 2.1 mm, 2.5 µm）was used to separate colchicine by gradient elution with mobile phase of 0.01% (v/v) ammonia-methanol. The amount and filling sequence of magnesium sulfate (MgSO4) and primary secondary amine (PSA) suitable for in-syringe DSPE were studied. Scopolamine was screened as the quantitative internal standard (IS) for colchicine analysis according to the consistency of recovery rate, chromatographic retention time and matrix effects. The limits of detection for colchicine in plasma and urine were both 0.06 ng mL-1 and the limits of quantitation were both 0.2 ng mL-1. The linear range was 0.04 - 20 ng mL-1 (Equivalent to 0.2-100 ng mL-1 in plasma or urine) with a correlation coefficient r > 0.999. By IS calibration, the average recoveries at three spiking levels in plasma and urine were 95.3-102.68% and 93.9-94.8% with the relative standard deviations (RSDs) of 2.9-5.7% and 2.3-3.4%, respectively. The matrix effects, stability, dilution effects and carryover for determination of colchicine in plasma and urine were also evaluated. The elimination of colchicine within 72-384 h post-ingestion was studied for a poisoning patient with the doses of 1 mg d-1 for 39 days and then 3 mg d-1 for 15 days).

Biodegradation of sulfametoxydiazine by Alcaligenes aquatillis FA: Performance, degradation pathways, and mechanisms.

This study reports the isolation and characterization of a novel bacterial strain Alcaligenes aquatillis FA with the ability to degrade sulfametoxydiazine (SMD), a commonly used sulfonamide antibiotic (SA) in livestock and poultry production. The biodegradation kinetics, pathways, and genomic background of SMD by FA were investigated. The results showed that strain FA had high specificity to degrade SMD, and was unable to effectively degrade its isomer, sulfamonomethoxine. The SMD biodegradation followed a first-order kinetic model with a rate constant of 27.39 mg·L-1·day-1 and a half-life of 5.98 days. The biodegradation pathways and detoxification processes of SMD were proposed based on the identification of its biodegradation byproducts and the biotoxicity assessment using both the ecological structure-activity relationship (ECOSAR) model and biological indicator. The involvement of novel degrading enzymes, such as dimethyllsulfone monooxygenase, 4-carboxymuconolactone decarboxylase, and 1,4-benzoquinone reductase, was inferred in the SMD biodegradation process. The presence of sul2 and dfrA genes in strain FA, which were constitutively expressed in its cells, suggests that multiple mechanisms were employed by the strain to resist SMD. This study provides new insights into the biodegradation of sulfonamide antibiotics (SAs) as it is the first to describe an SMD-degrading bacterium and its genetic information.

Pharmacokinetics, withdrawal time, and dietary risk assessment of enrofloxacin and its metabolite ciprofloxacin, and sulfachloropyridazine-trimethoprim in Taihe black-boned silky fowls.

Enrofloxacin (ENR) and sulfachloropyridazine combined with trimethoprim (TMP) were commonly used in poultries to treat bacterial infections. In this study, the pharmacokinetics of these antibiotics in four tissues of Taihe black-boned silky fowls was studied. The results showed that these drugs were absorbed and distributed rapidly, with the highest concentration showing in skin. Meanwhile, ENR and its metabolite ciprofloxacin (CIP) and TMP were depleted slowly, particularly in skin with the elimination half-lives being 37.1, 36.9, and 72.7 days, respectively. It may be attributed to the abundance of melanin in skin. The dietary risk assessment suggested that the long-term dietary intakes of ENR, CIP, and TMP showed a considerable threat to human health. Based on the experiment, the withdrawal times of 284 days for ENR + CIP and 159 days for TMP were acquired, which showed that these drugs are not appropriate for the application in Taihe black-boned silky fowls.

Effects of maternal exposure to procymidone on hepatic metabolism in the offspring of mice.

As an effective fungicide widely used in agricultural production, the excessive procymidone (PRO) residue has been detected in the environment and food. Our previous study demonstrated that PRO could destroy the intestinal barrier in mice and has a joint toxic effect. To explore the cross-generational impact of maternal exposure, 10-week-old C57BL/6 female mice were orally administrated to 10 and 100 mg/kg body weight/day of PRO during pregnancy and lactation. The offspring obtained nutrients from the maternal through the placenta and breast milk, and PRO residues were detected in the liver, intestine, and feces of F1 generation. Fecal examination found that the residual PRO had been completely metabolized when the offspring mice grew to 35 days. The drug residue of F1 generation male mice was higher than that of female mice. We attributed this result to the difference in cytochrome P450 (CYP450) enzyme expression between male and female mice. The transcriptional levels of CYP1A1, CYP1A2, CYP2D9, and CYP3A4, and CYP450 protein expression levels, were higher in female mice. Furthermore, targeted MS of plasma revealed abnormal amino acid levels. In addition, PRO-induced hepatic metabolite changes in F0 and F1-7w mice. KEGG pathway analysis further showed that PRO jointly changed the amino acid biosynthesis pathway of the maternal and offspring. In summary, these results indicated that maternal exposure to PRO during a special period would interfere with self metabolism, and offspring will also have metabolic disorders.

pH-responsive switchable deep eutectic solvents to mediate pretreatment method for trace analysis of triazole fungicides in peel wastes.

The existing QuEChERS-combined analytical pretreatment methods are limited by large reagent consumption, high environmental burden, and mediocre effects. To provide an efficient and green pretreatment method, this study developed pH-responsive switchable deep eutectic solvents (SDESs) to extract triazole fungicides (TFs) from fruit peel wastes, which could enhance the preconcentration effect of target compounds in food waste samples with complex matrices. The mechanisms of pH-induced phase transition and hydrophobicity-hydrophilicity conversion of pH-responsive SDESs were investigated by pH phase diagrams and chemical characterization techniques, respectively. We validated the established method by high-performance liquid chromatography-diode array detector (HPLC-DAD), and lower LOD (0.089-0.351 ng mL-1), LOQ (0.297-1.172 ng mL-1), RSD (≤8.8 %) and satisfactory recoveries (90.6 %-110.9 %) and preconcentration factors (389-512) were obtained in rotting grape peel, watermelon peel, and orange peel samples. Our findings highlight the potential of pH-responsive SDESs in the extraction and analysis of various natural food products.

The fungicide prothioconazole and its metabolite prothioconazole-desthio disturbed the liver-gut axis in mice.

The triazole fungicide prothioconazole (PTC) can cause adverse effects in animals, and its main metabolite prothioconazole-desthio (PTC-d) is even much more harmful. However, the toxic effects of PTC and PTC-d on the liver-gut axis of mice are still unknown. In the present experiment, we found that oral exposure to PTC and PTC-d increased total bile acids (TBAs) levels in the serum, liver, and feces. Correspondingly, the transcription of genes involved in bile acids (BAs) disposition was significantly influenced by PTC or PTC-d exposure. Furthermore, the BAs composition of serum BAs was analyzed by LC-MS, and the results indicated that PTC and PTC-d exposure changed the BAs composition, lowered the ratio of conjugated/unconjugated BAs, elevated the ratio of CA/b-MCA, and enhanced the hydrophobicity of BAs pool. 16s RNA gene sequencing of the DNA from colonic contents uncovered that PTC and PTC-d exposure altered the relative abundance and constitution of intestinal microbiota, increasing the relative level of Lactobacillus with bile salt hydrolase (BSH) activity. Furthermore, PTC and PTC-d exposure impaired the gut barrier function, causing an increase in mucus secretion. In particular, the effects of PTC-d on some endpoints in the BAs metabolism and gut barrier function had been proven to be more significant than the parent compound PTC. All these findings draw attention to the health risk of PTC and PTC-d exposure in regulating BAs metabolism, which might lead to some metabolic disorders and occur of related diseases in animals.

Atrazine Exposure Induces Hepatic Metabolism Disorder in Male Adult Zebrafish.

Atrazine (ATZ) is a herbicide used in agricultural production and has been detected in surface water due to its widespread use worldwide. This may pose a threat to the health of aquatic animals. To explore the ATZ−induced hepatic metabolism disorder, male zebrafish were exposed to 300 and 1000 µg/L ATZ for 21 days, respectively. The results revealed that ATZ exposure significantly reduced hepatic triglyceride (TG) levels, while significantly (p < 0.05) increased pyruvate (PYR) and total cholesterol (TC) levels. In addition, the liver sample from the 1000 µg/L ATZ−treated group was used for GC/MS metabolomic analysis. The principal component analysis (PCA) model showed significant separation of the 1000 µg/L ATZ group from the control group, indicating that ATZ exposure altered hepatic metabolism in male adult zebrafish. A total of 29 significantly (p < 0.05) different metabolites were observed and identified in the ATZ−treated group. Moreover, the most disturbed pathways by ATZ were the arginine and proline metabolic pathways, followed by the glutathione metabolic pathway. Three and two metabolites were significantly altered in the arginine and proline metabolic pathways and glutathione metabolic pathway, respectively. Based on these results, we suggested that ATZ was capable of altering liver metabolism in zebrafish and that its ecological risk to aquatic organisms cannot be ignored.

Maternal Sodium p-Perfluorous Nonenoxybenzene Sulfonate Exposure Disturbed Lipid Metabolism and Induced an Imbalance in Tyrosine Metabolism in the F1 Generation of Mice.

The toxicity of perfluorinated compounds (PFCs) to mammals has recently received increasing attention. However, the effects of maternal sodium p-perfluorous nonenoxybenzene sulfonate (OBS) exposure during pregnancy and lactation on the liver function of dams (F0) and offspring (F1) mice are still unknown. The results demonstrated that maternal OBS treatment could not only induce lipid metabolism dysfunction but also disrupt amino acid metabolism in the liver of F0 and F1 generations. OBS had marked accumulation in the liver, and the serum and liver triglyceride (TG) levels increased in the F0 and F1 generations after maternal OBS exposure. Moreover, maternal OBS exposure changed the transcriptional levels of genes related to lipid metabolism (fatty acid (FA) synthesis, TG synthesis, and transport) and induced changes in the amino acid level in dams and 20-day-old mice offspring (F1-20 d). Additionally, the regulation of lipid metabolism by OBS was mainly dependent on the activation of peroxisome proliferator-activated receptor γ (PPARγ) and cluster of differentiation 36 (CD36). Interestingly, OBS could also disturb tyrosine (TYR) metabolism by increasing the TYR level and downregulating fumarate acetoacetate hydrolase (FAH). Together, these results indicated that the liver can be perceived as the major target tissue of OBS, which strongly affected metabolic function and ultimately led to an imbalance in the metabolism of lipids and TYR. In summary, maternal OBS exposure during pregnancy and lactation has toxic effects on the hepatic metabolism of dams and offspring, indicating that the toxic effects could obviously cross generations of mice, and we should pay more attention to understanding the health risk to both dams and offspring.

Sub-Chronic Difenoconazole Exposure Induced Gut Microbiota Dysbiosis in Mice.

Difenoconazole (DIF) is a widely separated triazole fungicide in many countries. The excessive usage of DIF increases the high volume of residues in agriculture production and water bodies. Some previous studies demonstrated the toxic effects of DIF on non-target animals, however, there were still some gaps in the knowledge of the potential hazards of DIF to mammals and human health. Herein, 7-week-old male mice were exposed to 30 and 100 mg/kg/day DIF for 14 and 56 days. We observed that 56 days of DIF exposure decreased the colonic mucus expression of alcin blue-periodic acid-schiff (AB-PAS) stain and the immunochemical stain of muc2 protein. The transcript levels of mucin protein (muc1, muc2 and muc3) decreased significantly in the gut of mice followed 56 days of 100 mg/kg/day DIF exposure. In addition, the gut microbiota composition was also affected after 14 or 56 days of DIF exposure. Although the mucus expression after 14 days of DIF exposure only decreased slightly, the gut microbiota composition compared with the control group was changed significantly. Moreover, the DIF-30 and DIF-100 caused respectively different changes on the gut microbiota. The relative abundance of Bacteroidetes decreased significantly after 14 days and 56 days of DIF exposure. After 14 days of DIF exposure, there were 35 and 18 differential genera in the DIF-30 and DIF-100 group, respectively. There were 25 and 32 differential genera in the DIF-30 and DIF-100 group after 56 days of exposure, respectively. Meanwhile, the alpha diversity indexes, including observed species, Shannon, Simpson, Chao1 and ACE, in gut microbiota decreased significantly after 56 days of DIF exposure. Interestingly, the relative abundance of Akkermansia increased significantly after 56 days of 100 mg/kg/d DIF exposure. Although Akkermansia was considered as one probiotic, the phenomenon of dramatic Akkermansia increase with the decrease in gut microbiota diversity needed further discussion. These results provided some new insights on how DIF exposure impacts the mucus barrier and induces gut microbiota dysbiosis.

A magnetic solid phase extraction based on UiO-67@GO@Fe3O4 coupled with UPLC-MS/MS for the determination of nitroimidazoles and benzimidazoles in honey.

A novel composite material consisting of zirconiumbiphenyldicarboxylate metal-organic framework, graphite oxide and ferroferric oxide was fabricated by a facile one-step method and served as a magnetic solid phase extraction sorbent for the simultaneous determination of nitroimidazoles and benzimidazoles in honey. The amount of graphite oxide for the synthesis of composite material and analysis parameters were optimized. The optimum parameters were: dipotassium hydrogen phosphate buffer solution (pH 6) as diluent solvent, adsorption time 10 min, desorption time 5 min, methanol/acetonitrile (1:1, V/V) as desorption solvent. The targets were detected by ultra-high performance liquid chromatography tandem mass spectrometry. The recoveries of twelve analytes ranged in 70.5%-103.4% with relative standard deviations lower than 12.9% (n = 3). The quantification limits were 0.2-0.6 µg/kg. Using the composite material as sorbent, a simple, rapid and environmental-friendly method based on magnetic solid phase extraction was successfully developed for determination of seven nitroimidazoles and five benzimidazoles in honey.

Insights into the effects of difenoconazole on the livers in male mice at the biochemical and transcriptomic levels.

Difenoconazole (DFZ) is a broad-spectrum triazole fungicide, that is extensively used in agriculture. Studies have shown that residues of DFZ and other fungicides have toxic effects on nontarget organisms. However, its hepatoxicity in mammals remains unclear. Here, we characterized the toxic hepatic effects in male C57BL/6 mice exposed to 30 and 100 mg/kg bw DFZ for 14 and 56 days, respectively. The results revealed that DFZ could increase the relative liver weights, however, the relative fat and spleen weights decreased. More importantly, DFZ exposure changed the hepatic morphology and induced hepatic oxidative stress. Gene expression analysis suggested that DFZ could induce a glycolipid metabolism disorder. Moreover, hepatic transcriptomic analysis revealed the effects of DFZ exposure on the transcriptional levels of various genes, and enrichment analysis of differentially expressed genes (DEGs) showed that energy metabolism and immune-associated pathways were mainly affected. We validated the results from transcriptomic analysis and found that some key genes related to energy metabolism were affected. In addition, flow cytometry showed that the CD3+/CD4+ and CD3+ /CD8+ levels declined in the spleen of mice. Taken together, these findings combined with transcriptome analysis highlighted that DFZ caused different endpoints in the liver, which could provide more evidence for investigating the toxic effects of DFZ in mammals.

Stereoselective effects of fungicide difenoconazole and its four stereoisomers on gut barrier, microbiota, and glucolipid metabolism in male mice.

Difenoconazole is a commonly used triazole fungicide that consists of four stereoisomers [(2S,4S)-, (2S,4R)-, (2R,4R)-, and (2R,4S)-isomers] with different bioactivity. For example, the toxicity of the (2R,4S)-isomer to fish is approximately seven times higher than that of the (2S,4S)-isomer. However, the stereoselective toxic effects of difenoconazole stereoisomers on mammals have received little attention. In the present study, adult male mice were orally treated with a mixture of the four stereoisomers or each stereoisomer individually (0, 30, or 100 mg/kg/d) by gavage for 28 days. Pathological staining of the liver sections showed that the (2R,4R)-isomer caused lipid droplet accumulation. The mixture or each individual stereoisomers decreased the levels of amino acids and acyl-carnitine in serum. Moreover, the (2S,4R)-, (2R,4R)-, and (2R,4S)-isomers affected intestinal permeability, causing decreases in mucus secretion and tight junction protein expression in colon. Analysis of the gut microbiota composition showed that the stereoisomers caused decreases of OTU numbers and observed species at different levels. Interestingly, difenoconazole and its four stereoisomers reduced the relative abundance of Bacteroidetes at the phylum level and some short-chain fatty acid (SCFA)-producing bacteria. Taking the findings together, 2R-difenoconazole with strong bioactivity against pathogenic fungi also had significant effects in mammals, disrupting hepatic lipid metabolism, intestinal permeability, and gut microbiota. It is concluded that the health risks of the four difenoconazole stereoisomers to mammals should not be overlooked.

Determining a wide range of antibiotics and pesticides in poultry feathers using selective accelerated solvent extraction-liquid chromatography-mass spectrometry.

This study established a detection method based on accelerated solvent extraction-liquid chromatography-mass spectrometry for determining residues of 3 chloramphenicols, 8 macrolides, 18 sulfonamides, 4 nitroimidazoles, 15 insecticides, and 22 fungicides in poultry feathers. The extraction solvent, methanol, was used for a static extraction time of 5 min, and repeated three times. Fifty milligrams of adsorbents C18/PSA (1 : 1, W/W) were added to the extraction cell to achieve simultaneous extraction and purification. The extraction efficiency of three solvents, methanol, acetonitrile and ethyl acetate, was investigated. An orthogonal experimental design was used to explore the optimal combination of extraction temperature, static extraction time, number of extraction cycles, and adsorbent ratio for accelerated solvent extraction. After the optimal ratio was determined, the dosage of adsorbents was optimized. The extracted sample solution was concentrated by blowing nitrogen, redissolved, passed through a 0.22 µm PTFE membrane filter, then injected for instrumental analysis. The validation results showed that the recovery of the proposed method was 60.4-107.6%, the limit of detection 0.2-3.0 µg kg-1, and the limit of quantification 0.5-8.3 µg kg-1. This quantitative multi-residue detection method was able to determine the residues of 70 target compounds in poultry feathers.