Association of pyrethroids exposure with asthma in US children and adolescents: a nationally representative cross-sectional study.

Pyrethroids (PYR) are among the most widely used insecticides in households, leading to substantial exposure. Children and adolescents, especially during growth spurts, have a reduced capacity to effectively metabolize these insecticides. The relationship between PYR exposure and asthma in these age groups remains poorly understood, highlighting the need for further research.We used data from the 2007-2014 National Health and Nutrition Examination Survey, which included 1181 children aged 6-11 years and 1258 adolescents aged 12-19 years. The concentration of the PYR metabolite 3-phenoxybenzoic acid (3-PBA) in urine was quantified using solid-phase extraction-high-performance liquid chromatography-heated electrospray ionization tandem mass spectrometry. Asthma was defined based on self-reported doctor diagnoses from the questionnaire. PYR exposure was measured using urine samples collected simultaneously with the questionnaire. We explored the association between PYR exposure and asthma using multiple logistic regression analyses, adjusting for potential confounders.Multiple logistic regression analyses revealed no significant association between PYR exposure and asthma in children and adolescent boys (all P > 0.05). In contrast, PYR exposure was significantly associated with asthma in adolescent girls aged 12-19 years. Specifically, for "ever asthma," the odds ratios (ORs) were 2.49 (95% CI = 1.03-5.97) in the second quartile of PYR exposure and 2.48 (95% CI = 1.04-5.91) in the third quartile, each in comparison to the first quartile. For "current asthma," in comparison to the first quartile, the ORs were 3.99 (95% CI = 1.55-10.26) in the second quartile of PYR exposure, 3.39 (95% CI = 1.32-8.70) in the third quartile, and 2.93 (95% CI = 1.24-6.90) in the fourth quartile.Conclusions:Our study found a significant association between PYR exposure and asthma in adolescent girls, whereas no significant association was observed in children and adolescent boys. These findings suggest potential sex and age differences in susceptibility to PYR exposure. Further research is warranted to confirm these results and elucidate the underlying mechanisms.

An in silico investigation of the toxicological effects and biological activities of 3-phenoxybenzoic acid and its metabolite products.

We aimed to elucidate the toxic effects and biological activities of 3-phenoxybenzoic acid (3PBA) and its metabolite products.Numerous in silico methods were used to identify the toxic effects and biological activities of 3PBA, including PASS online, molecular docking, ADMETlab 2.0, ADMESWISS, MetaTox, and molecular dynamic simulation.Ten metabolite products were identified via Phase II reactions (O-glucuronidation, O-sulfation, and methylation).All of the investigated compounds were followed by Lipinski's rule, indicating that they were stimulants or inducers of hazardous processes.Because of their high gastrointestinal absorption and ability to reach the blood-brain barrier, the studied compounds' physicochemical and pharmacokinetic properties matched existing evidence of harmful effects, including haematemesis, reproductive dysfunction, allergic dermatitis, toxic respiration, and neurotoxicity.The studied compounds have been linked to the apoptotic pathway, the reproductivity system, neuroendocrine disruptors, phospholipid-translocating ATPase inhibitors, and JAK2 expression.An O-glucuronidation metabolite product demonstrated higher binding affinity and interaction with CYP2C9, CYP3A4, caspase 3, and caspase 8 than 3PBA and other metabolite products, whereas metabolite products from methylation were predominant and more toxic.Our in silico findings partly meet the 3Rs principle by minimizing animal testing before more study is needed to identify the detrimental effects of 3PBA on other organs (liver, kidneys).Future research directions may involve experimental validation of in silico predictions, elucidation of molecular mechanisms, and exploration of therapeutic interventions.These findings contribute to our understanding of the toxicological profile of 3PBA and its metabolites, which has implications for risk assessment and regulatory decisions.

Key properties & pharmacokinetics of 3PBA & its metabolites were reportedMetabolite products from methylation were predominant and more toxicMain toxics: haematemesis, reproductive dysfunction, toxic respiration, dermatitis.

Exposure to biocides and its association with atopic dermatitis among children and adolescents: A population-based cross-sectional study in South Korea.

BACKGROUND: Biocides have emerged as a contributor to the rising cases of atopic dermatitis among children and adolescents. Previous animal studies suggested that phenols, parabens, and pyrethroid insecticides present in these products might play a role in atopic dermatitis. However, there's limited epidemiological evidence confirming the individual or combined effects of exposure to these chemicals on atopic dermatitis in young populations. This study aimed to investigate the association between phenol, paraben, and pyrethroid metabolite levels in urine and atopic dermatitis among Korean children and adolescents METHODS: We analyzed 556 preschool children (3-5 years), 701 schoolchildren (6-11 years), and 731 adolescents (12-17 years) enrolled in the 4th Korean National Environmental Health Survey (KoNEHS) (2018-2020). We used logistic regression and Bayesian kernel machine regression to evaluate the association between atopic dermatitis and individual or mixed exposure to urinary triclosan (TCS), parabens (methylparaben, ethylparaben, propylparaben, and butylparaben), and 3-phenoxybenzoic acid (3-PBA) levels. RESULTS: Urinary TCS levels were positively associated with atopic dermatitis in schoolchildren. When stratified by sex, male schoolchildren exhibited an increasing prevalence of atopic dermatitis as their urinary TCS and 3-PBA levels increased. The combined effect of biocide mixtures on atopic dermatitis was also significantly increased in male schoolchildren, with TCS as the main contributor. CONCLUSIONS: These study findings suggest that biocides at levels found in Korean children and adolescents affect atopic dermatitis.

Synthesis, docking study, and antitumor evaluation of benzamides and oxadiazole derivatives of 3-phenoxybenzoic acid as VEGFR-2 inhibitors.

Current chemotherapeutic agents have several limitations, including lack of selectivity, the development of undesirable side effects, and chemoresistance. As a result, there is an unmet need for the development of novel small molecules with minimal side effects and the ability to specifically target tumor cells. A new series of 3-phenoxybenzoic acid derivatives, including 1,3,4-oxadiazole derivatives (4a-d) and benzamides derivatives (5a-e) were synthesized; their chemical structures were confirmed by Fourier-transform infrared spectroscopy, 1H nuclear magnetic resonance (NMR), 13C NMR, and mass spectra; and various physicochemical properties were determined. The antiproliferative activities of the new derivatives were evaluated by means of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Three compounds (4b, 4c, and 4d) exhibited cytotoxicity against two of the three cell lines tested, five compounds (3, 4a, 5a, 5b, and 5e) were toxic to one cell line, while two compounds (5c and 5d) were not cytotoxic to any of the three cell lines tested in the current study. Based on docking scores, MTT assay findings, and vascular endothelial growth factor receptor 2 (VEGFR-2) kinase activity data, Compound 4d was selected for further biological investigation. Flow cytometry was used to determine the mode of cell death (apoptosis vs. necrosis) and the effect on cell cycle progression. Compound 4d arrested HepG2 hepatocellular carcinoma cells in the G2/M phase and activated both the intrinsic and extrinsic apoptosis pathways. In conclusion, Compound 4d has shown promising results for future research as a potent VEGFR-2 tyrosine kinase inhibitor.

Biodegradation of the pyrethroid cypermethrin by bacterial consortia collected from orange crops.

Pyrethroids, such as cypermethrin (CYP), are widely employed in agriculture, promoting environmental pollution and the need for efficient decontamination methods. In this study, bacteria from orange crops were explored for CYP biodegradation. Among 40 tested bacterial strains, 20 grew in the presence of CYP and 19 performed statistically significant CYP biodegradation in 5 days (20.5%-97.8%). In addition, 3-phenoxybenzoic acid, the main metabolite from CYP, was quantified ranging from 1.1 mg.L-1 to 32.1 mg.L-1. The five most efficient strains, and consortia composed of 5, 10 and 20 bacteria biodegraded the CYP formulation as sole carbon source in phosphate buffer and in minimum mineral medium. Under optimized conditions determined employing Response Surface Methodology, Bacillus sp. CSA-1 and the consortium composed of 10 strains biodegraded 71.0% and 71.6% CYP in 24 h, respectively. Moreover, metabolite identification enabled the proposal of an extended biodegradation pathway with 29 identified compounds, including different new amide and amine derivatives that expanded the knowledge about the fate of this compound in the environment. Experiments of bioaugmentation in soil using Bacillus sp. CSA-1 and the consortium of 10 bacterial strains resulted in faster CYP biodegradation than natural attenuation, showing that the selection of efficient strains for composing a consortium is an interesting approach for bioremediation of pyrethroids.

Synthetic pyrethroids common metabolite 3-phenoxybenzoic acid induces caspase-3 and Bcl-2 mediated apoptosis in human hepatocyte cells.

Synthetic pyrethroids are a group of insecticides frequently used in public health and agriculture, and 3-PBA is a common metabolite of them. Although the liver is the primary organ responsible for metabolizing many compounds including pesticides, to the authors' knowledge there have been no studies on the direct hepatotoxic effects of 3-PBA. Therefore, this study aimed to investigate the possible hepatotoxic effects of 3-PBA on a Human Hepatoma Cell Line (HepG2) and the underlying apoptotic mechanisms. Firstly, an LC50 of 1041.242 µM was calculated for 3-PBA by using the WST-1 test with concentrations ranging between 1 µM and 10 mM. Following that, the HepG2 cells in the experimental group were exposed to 3 different concentrations of 3-PBA (1/5 LC50, 1/10 LC50 and 1/20 LC50) for 24 hours. The apoptotic mechanism was evaluated by using flow cytometry, and immunofluorescence assays for Caspase 3 and Bcl-2. In the flow cytometry assay, the total number of apoptotic cells increased in a dose dependent manner (p < 0.05). In the immunofluorescence assay, the Caspase 3 protein showed strong immunoreactivity in the experimental groups, while the reaction to the Bcl-2 protein was minimal. These results demonstrated that 3-PBA has a significant hepatotoxic effect on HepG2 cells and induces apoptosis via the regulation of Caspase-3 and Bcl-2. Furthermore, our results could further the understanding of the fundamental molecular mechanisms of 3-PBA hepatotoxicity. More studies are needed to determine the effects of long-term exposure to 3-PBA and also the molecular mechanisms underlying hepatotoxicity.

Multivalent nanobody as capture antibody-based enzyme linked immunosorbent assay for detection of 3-phenoxybenzoic acid in urine.

Nanobodies (Nbs) as capture antibodies in enzyme-linked immunosorbent assays (ELISAs) is greatly hampered by their poor performance after attaching onto polystyrene microplates. Reasons behind those phenomena remain unknown. One of possible explanation is that Nbs with a single domain might lose their accessibility of paratope when adsorbed on the plates. Increasing their binding sites might improve performance in capture Nbs-based ELISA. In this study, anti-3-phenoxybenzoic acid (3-PBA) Nbs was assembled to trivalent form (Nb3) in tandem with flexible linkers (G4S)3. Direct competitive ELISA on the basis of Nb3 and 3-PBA-horseradish peroxidase was developed for detection of 3-PBA in livestock urine. The ELISA had a half-maximum (IC50) inhibition concentration of 0.51 ng/mL, with a limit of detection of 0.02 ng/mL, which was more sensitive than that of the parental Nb with a IC50 of 2.39 ng/mL. The average recoveries of 3-PBA spiked in swine, sheep and dairy cow urine samples by the assay ranged from 89.52% to 114.25% and agreed well with those of liquid chromatography mass spectrometry (LC-MS). The above results indicated that multivalent Nbs could be treated as the capture antibody in ELISA for routine screening analysis of 3-PBA residues in urine.

Residential pyrethroid insecticide use, urinary 3-phenoxybenzoic acid levels, and attention-deficit/hyperactivity disorder-like symptoms in preschool-age children: The Environment and Development of Children study.

Previous animal studies have reported that pyrethroids can cause dopamine system abnormalities and attention-deficit/hyperactivity disorder (ADHD) phenotypes. However, epidemiological studies investigating the associations between pyrethroid exposure and ADHD are limited. We aimed to investigate the association between pyrethroid exposure and ADHD-like symptoms among preschool-age children. We used data from 385 children at 4 years of age participating in the Environment and Development of Children (EDC) study. We evaluated pyrethroid exposure through questionnaires and urinary 3-phenoxybenzoic acid (3-PBA) concentrations. We assessed ADHD-like symptoms using the Korean ADHD rating scale (K-ARS). We conducted negative binomial regressions to evaluate the associations between pyrethroid exposure and ADHD-like symptoms. Residential use of insecticide adhesive (ß = 0.42, 95% CI: 0.11, 0.74) and insecticide spray (ß = 0.33, 95% CI: 0.08, 0.59) was associated with an increase in log-transformed creatinine-adjusted urinary 3-PBA concentrations. Residential insecticide adhesive use was associated with a 51.6% increase in K-ARS scores (95% confidence interval [CI]: 6.3, 116.1) among boys, when compared with non-users. When compared with creatinine-adjusted 3-PBA levels <0.50 µg/g creatinine, creatinine-adjusted 3-PBA levels ≥3.80 µg/g creatinine were associated with a 58% increase in K-ARS scores (95% CI: 0.1, 150.5) among boys. We found associations of residential pyrethroid insecticide use and urinary 3-PBA concentrations with K-ARS scores among preschool-age boys. Since the present study explored cross-sectional associations in preschool-age children, the possibility of reverse causality cannot be dismissed. Further studies implementing a cohort study design are warranted.

Association between pyrethroid pesticide exposure and hearing loss in adolescents.

BACKGROUND: Hearing loss in adolescents is a serious public health problem with a high prevalence. Pyrethroids are one of the most widely applied insecticides that have been linked to neurotoxicity. However, there is no study about the effect of pyrethroid insecticide exposure on the auditory system in the general population. OBJECTIVE: To investigate the association between pyrethroid pesticide exposure and hearing loss in adolescents in the United States. METHODS: A total of 720 adolescents aged 12-19 years who participated in the National Health and Nutrition Examination Survey (NHANES, 2007-2010) were considered. 3-phenoxybenzoic acid (3-PBA), a urinary metabolite, was applied as a biomarker to assess pyrethroid exposure. Hearing loss in adolescents was defined as a pure-tone average (PTA) > 15 dB in either ear. Multivariate linear and logistic regression analyses were conducted to examine the associations of urinary 3-PBA with PTA hearing thresholds and risk of hearing loss, respectively. RESULTS: The weighted geometric mean of 3-PBA levels in urine was 0.32 µg/g creatinine, and 7.62% of adolescents had hearing loss. After adjusting for age, sex, race/ethnicity, BMI, serum cotinine, annual family income and exposure to loud noise/music, linear regression analyses found that Ln-transformed 3-PBA was positively correlated with increase of hearing thresholds in either left (ß = 0.61, 95% CI: 0.20-1.01) or right ear (ß = 0.52, 95% CI: 0.16-0.89). Logistic regression analyses showed that adjusted odds ratio (OR) for hearing loss in adolescents with the highest tertile (≥0.52 µg/g creatinine) of 3-PBA were 3.12 (95% CI: 1.42-6.83) compared with the lowest tertile (<0.18 µg/g creatinine), with significant linear trends across tertiles. CONCLUSION: Pyrethroid pesticide exposure was positively associated with hearing loss in U.S. adolescents. This study provides new evidence for the association between pyrethroid exposure and auditory function.

Construction and characterization of a class-specific single-chain variable fragment against pyrethroid metabolites.

Pyrethroids are insecticides that are widely used in rural and urban areas worldwide. After entering the environment, pyrethroids are rapidly metabolized or degraded by various biological or abiotic methods. In this study, a single-chain variable fragment (scFv) which could simultaneously detect three pyrethroid metabolites was constructed based on a hybridoma raised against 3-phenoxybenzoic acid (3-PBA). By molecular docking, it showed that there were hydrogen bonds, hydrophobic interactions, CH-π interaction, and cation-π interaction between 3-PBA and its scFv. All the contact residues contributing to hydrogen bonds are located in VH-CDR2 or its neighboring region, and two of them were mutants of the closest germline sequence. Based on competitive ELISA, the half maximal inhibitory concentration (IC50) of the scFv for 3-PBA, 3-phenoxybenzaldehyde (PBAld), and 3-phenoxybenzyl alcohol (PBAlc) were calculated to be 0.55, 0.59, and 0.63 µgmL-1, respectively. The scFv also showed 23.91%, 13.41%, 1.15%, 1.00%, and 0.56% cross-reactivity with phenothrin, deltamethrin, fenvalerate, beta-cypermethrin, and fenpropathrin. The broad specificity of the scFv may be due to its hapten design. The scFv could be employed in class-specific immunoassays for pyrethroid metabolites with phenoxybenzyl (PB) group. It is also potentially used for characterizing degradation of pyrethroids or detecting PBAlc (PBAld) alone, and the detection results should be confirmed by other selective methods. KEY POINTS: • A scFv which can simultaneously detect 3-PBA, PBAlc, and PBAld was constructed. • Antibody informatics and binding mode of the scFv were obtained. • The reason for its broad specificity was discussed. • It could be used to monitor single or multi-pyrethroid metabolites with PB group.

Co-metabolic enzymes and pathways of 3-phenoxybenzoic acid degradation by Aspergillus oryzae M-4.

As an intermediate metabolite of pyrethroids, 3-phenoxybenzoic acid (3-PBA) is more toxic than its parent compounds and has been detected in milk, soil, and human urine. 3-PBA can be metabolized through microbial degradation, but the microbial co-metabolic enzymes and pathways involved in 3-PBA degradation are unclear. This study investigated the enzymes types and possible pathways in the co-metabolic degradation of 3-PBA by Aspergillus oryzae M-4. The enzymes involved in co-metabolic degradation of 3-PBA and its intermediate metabolites were induced, and existed extracellularly and intracellularly except the catechol-degrading enzyme. Inhibitors and inducers of these oxidases were used to examine the enzymes required for co-metabolic degradation of 3-PBA and its intermediate metabolites. 3-PBA is hydroxylated to produce 3-hydroxy-5-phenoxy benzoic acid through the catalytic actions of lignin peroxidase (LiP). Phenol and gallic acid, the metabolites of 3-PBA, are produced via cleavage of an ether bond under the catalytic actions of cytochrome P450 (CYP450) and LiP. Phenol can be converted to catechol by LiP; catechol and gallic acid are cleaved to form long-chain olefin acid or olefin aldehyde by dioxygenase and LiP. In corn flour, some of these enzyme activators such as FeCl3, 4-cumaric acid, veratryl alcohol and sodium periodate appeared to improve 3-PBA degradation. The results provide a reliable pathway and characteristics for co-metabolic microbial degradation of 3-PBA in food and the environment.

Cohort-friendly protocol for the determination of two urinary biomarkers of exposure to pyrethroids and neonicotinoids using gas chromatography-triple quadrupole mass spectrometry.

Neonicotinoids (NEOs) and synthetic pyrethroids (PYRs) are active ingredients of commercial pesticides and/or insecticides with extensive indoor and outdoor applications, worldwide. Improved exposure metrics are warranted for NEOs and PYRs, if we are to better understand their human health effects. A cohort-friendly protocol for determining non-specific biomarkers of exposure to NEOs and PYRs, e.g. 6-chloronicotinic acid (6-CN) and 3-phenoxybenzoic acid (3-PBA), respectively, in human urine voids was proposed. A series of optimization experiments were conducted to validate the bioanalytical protocol using gas chromatography coupled with triple quadrupole mass spectrometry (GC-QqQ-MS/MS) in MRM mode. The method reached low detection limits for both analytes (0.075 µg L-1 for 6-CN and 0.050 µg L-1 for 3-PBA) in a short preparation and analysis time. The method used small initial urine sample volume (2 mL), short extraction time (≈ 240 min for the batches of 32 samples) and instrumental analysis time (≈ 14 min) for both pesticide metabolites in a single run. This protocol could facilitate the assessment of population exposure metrics for these pesticides and their inclusion in health risk assessment. Graphical abstract.

The relationship of urinary 3-phenoxybenzoic acid concentrations in utero and during childhood with adiposity in 4-year-old children.

BACKGROUND: Pyrethroid pesticides are reported to be the most commonly used residential insecticides worldwide. We aimed to investigate the relationship between prenatal and postnatal 3-phenoxybenzoic acid (3-PBA) concentrations, and growth and adiposity parameters in 4-year-old children. METHOD: We obtained data from 578 children who participated in the prospective Environment and Development of Children (EDC) study at around 4 years of age (45-55 months) between August 2008 and July 2011. Anthropometric measurements were obtained at age 4 years. Prenatal and postnatal urinary 3-PBA concentration was measured in maternal urine samples at around 20 weeks of gestation, and in the 4-year-old children, respectively. RESULT: The detection frequency of urinary 3-PBA (geometric mean concentration) was 98-99% (0.98 µg/g Cr) in maternal urine, and almost 99-100% (1.34 µg/g Cr) in 4-year-old children. Prenatal urinary3-PBA concentration was not associated with height, weight, or body mass index (BMI) z-scores at 4 years of age, regardless of sex. Postnatal urinary3-PBA concentration was not related to height z-scores, but was positively associated with weight z-scores with marginal significance among only girls (p = 0.058). Analyzed by sex, there was a significant relationship between postnatal urinary 3-PBA concentration and BMI z-scores (p = 0.015) among girls, after adjusting for covariates. CONCLUSION: Childhood urinary 3-PBA concentration measured at 4 years of age was positively associated with BMI z-scores in 4-year-old girls, but prenatal urinary 3-PBA concentration at midterm pregnancy exhibited no association.

Efficient biodegradation of 3-phenoxybenzoic acid and pyrethroid pesticides by the novel strain Klebsiella pneumoniae BPBA052.

A novel Klebsiella pneumoniae strain (BPBA052) capable of degrading 3-phenoxybenzoic acid (3-PBA) was isolated from soybean rhizosphere soil. The strain was obtained by screening after enrichment, isolation, and purification using 3-PBA as the sole carbon and energy source. It could degrade 96.37% of 3-PBA (100 mg/L) within 72 h, and its growth and 3-PBA degradation followed kinetics models of logistic growth (XBPBA052 = 0.0883 × e0.0947t / [1 - 0.0792 × (1 - 0.0883 × e0.0947t)]; µm = 0.0947 h-1, X0 = 0.0883, and Xm = 1.1145) and first-order degradation (CBPBA052 = 101.8194 × e-0.0403t, k = 0.0403, t1/2 = 17.22 h), respectively. Based on Box-Behnken response surface analysis, the optimal temperature, pH, and 3-PBA concentration for K. pneumoniae BPBA052 were 35.01 °C, 7.77, and 150 mg/L, respectively. Moreover, pyrethroid pesticides (PPs) (such as ß-cypermethrin, permethrin, bifenthrin, deltamethrin, and fenvalerate) and 3-PBA metabolites (including phenol, catechol, and protocatechuate) were efficiently utilized by BPBA052. We propose a novel microbial metabolic pathway for 3-PBA, based on metabolite identification; enzyme-degrading activity; and cloning of the phenol hydroxylase, catechol 1,2-dioxygenase, and protocatechuate 3,4-dioxygenase genes. This study provides a fundamental platform for further studies to reveal the mechanism of biodegradation of 3-BPA and show K. pneumoniae BPBA052 as a potential microbial resource for bioremediation of environments polluted with 3-PBA or PPs.

Developmental toxicity of deltamethrin and 3-phenoxybenzoic acid in embryo-larval stages of zebrafish (Danio rerio).

Synthetic pyrethroids are the major insecticides used widely in agriculture and household pest control. Deltamethrin (DM), a widely used type II pyrethroid insecticide, is a relatively potent neurotoxicant. 3-Phenoxybenzoic acid (PBA) is the major metabolite formed due to metabolism of DM. In order to illustrate the toxic response of zebrafish embryos/larvae to DM and PBA the present research was carried out. For this 4hpf embryos were treated with two concentrations of DM (100 and 200 µg/L) for 48 h and PBA (1000 and 2000 µg/L) for 96 h or 99.9% ethanol (solvent control). Early life stage parameters were observed at specified time points. DM-treated embryo/larvae exhibited increased mortality, delay in hatching time, decrease in percentage of hatched embryos, increase of heartbeat rate and decrease in blood flow; lightening of body and eye pigmentation in a dose dependent manner. Pericardial and yolk sac edema along with were also caused by DM. Along with these crooked notochord, tail deformation was noticed in hatched and unhatched embryos. In case of PBA treated embryos and larvae, increased embryos/larvae length and yolk sac size were observed. Other abnormalities like edema (yolk sac and pericardial), decreased eye and body pigmentation were also observed but in some embryos only. These were not as severe as observed in parental compound indicating that DM is more toxic than its metabolite PBA. The data contributes to a better understanding of the potential consequences of fish exposed to DM and PBA.

ß-Cypermethrin and its metabolite 3-phenoxybenzoic acid exhibit immunotoxicity in murine macrophages.

ß-Cypermethrin (ß-CYP), one of most important pyrethroids, is widely used to control insects, and has been detected in organisms, including human. Pyrethroids have been shown to pose neurotoxicity, hepatotoxicity, endocrine disruption and reproductive risks in mammals. However, research in immunotoxicity of pyrethroids, especially their metabolites, is limited. A common metabolite of pyrethroids is 3-phenoxybenzoic acid (3-PBA) in mammals. Thus, in this study, we evaluated the immunotoxicity of ß-CYP and 3-PBA in mouse macrophages, RAW 264.7 cells. MTT assays showed that both ß-CYP and 3-PBA reduced cell viability in a concentration- and time-dependent manner. Flow cytometry with Annexin-V/PI staining demonstrated that both ß-CYP and 3-PBA induced RAW 264.7 cell apoptosis. Furthermore, our results also showed that N-acetylcysteine partially blocked ß-CYP- and 3-PBA-induced cytotoxicity and apoptosis. Intrinsic apoptotic pathway was stimulated by both ß-CYP and 3-PBA exposure. In addition, we found that ß-CYP and 3-PBA inhibited mRNA levels of pro-inflammatory cytokines with or without LPS stimulation. Phagocytosis assay showed that both ß-CYP and 3-PBA inhibited phagocytic ability of macrophages. Moreover, it was also found that both ß-CYP and 3-PBA increased reactive oxygen species (ROS) levels in RAW 264.7 cells. Accordingly, both ß-CYP and 3-PBA were found to regulate the mRNA levels of oxidative stress-related genes in RAW 264.7 cells. Taken together, the results obtained in this study demonstrated that ß-CYP and 3-PBA may have immunotoxic effect on macrophages and that elevated ROS may underlie the mechanism. The present study will help to understand the health risks caused by ß-CYP and other pyrethroids.

PBPK modeling of the cis- and trans-permethrin isomers and their major urinary metabolites in rats.

Permethrin, a pyrethroid insecticide, is suspected to induce neuronal and hormonal disturbances in humans. The widespread exposure of the populations has been confirmed by the detection of the urinary metabolites of permethrin in biomonitoring studies. Permethrin is a chiral molecule presenting two forms, the cis and the trans isomers. Because in vitro studies indicated a metabolic interaction between the trans and cis isomers of permethrin, we adapted and calibrated a PBPK model for trans- and cis-permethrin separately in rats. The model also describes the toxicokinetics of three urinary metabolites, cis- and trans-3-(2,2 dichlorovinyl)-2,2-dimethyl-(1-cyclopropane) carboxylic acid (cis- and trans-DCCA), 3-phenoxybenzoic acid (3-PBA) and 4'OH-phenoxybenzoic acid (4'-OH-PBA). In vivo experiments performed in Sprague-Dawley rats were used to calibrate the PBPK model in a Bayesian framework. The model captured well the toxicokinetics of permethrin isomers and their metabolites including the rapid absorption, the accumulation in fat, the extensive metabolism of the parent compounds, and the rapid elimination of metabolites in urine. Average hepatic clearances in rats were estimated to be 2.4 and 5.7 L/h/kg for cis- and trans-permethrin, respectively. High concentrations of the metabolite 4'-OH-PBA were measured in urine compared to cis- and trans-DCCA and 3-PBA. The confidence in the extended PBPK model was then confirmed by good predictions of published experimental data obtained using the isomers mixture. The extended PBPK model could be extrapolated to humans to predict the internal dose of exposure to permethrin from biomonitoring data in urine.

Degradation of 3-phenoxybenzoic acid by a filamentous fungus Aspergillus oryzae M-4 strain with self-protection transformation.

A novel filamentous fungus M-4 strain was isolated from soy sauce koji and identified as Aspergillus oryzae (Collection number: CGMCC 11645) on the basis of morphological characteristics and internal transcribed spacer sequence. M-4 could degrade 80.62 % of 3-phenoxybenzoic acid (3-PBA; 100 mg L-1) within 5 days. 3-PBA degradation occurred in accordance with first-order kinetics. The degradation metabolites of 3-PBA were identified through high-performance liquid chromatography-mass spectrometry (HPLC-MS). Relevant enzymatic activities and substrate utilization were also investigated, which indicated that M-4 could effectively degrade the intermediates of 3-PBA. Base on analysis of these metabolites, a novel biochemical pathway for the degradation of 3-PBA was proposed. There exists a mutual transformation between 3-phenoxy-benzyl alcohol and 3-PBA, which was firstly reported about the degradation of 3-PBA and may be attributed to self-protection transformation of M-4; subsequently, 3-PBA was gradually transformed into phenol, 3-hydroxy-5-phenoxy benzoic acid, protocatechuic acid and gallic acid. The safety of M-4 was evaluated via an acute toxicity test in vivo. The biodegradation ability of M-4 without toxic effects reveals that this fungus may be likely to be used for eliminating 3-PBA from contaminated environment or fermented foods.

Detection of 3-phenoxybenzoic acid in river water with a colloidal gold-based lateral flow immunoassay.

3-Phenoxybenzoic acid (3-PBA) is a general metabolite of synthetic pyrethroids. It could be used as a generic biomarker for multiple pyrethroids exposure for human or pyrethroid residues in the environment. In this study, monoclonal antibodies (mAbs) against 3-PBA were developed by using PBA-bovine serum albumin (BSA) as an immunogen. In the competitive enzyme-linked immunosorbent assay (ELISA) format, the I50 and I10 values of purified mAbs were 0.63 and 0.13 µg/ml, respectively, with a dynamic range between 0.19 and 2.04 µg/ml. Then, the colloidal gold (CG)-based lateral flow immunoassay was established based on the mAbs. The working concentration of coating antigen and CG-labeled antibodies and the blocking effects were investigated to get optimal assay performance. The cutoff value for the assay was 1 µg/ml 3-PBA, and the detection time was within 10 min. A total of 40 river water samples were spiked with 3-PBA at different levels and determined by the lateral flow immunoassay without any sample pretreatments. The negative false rate was 2.5%, and no positive false results were observed at these levels. This lateral flow immunoassay has the potential to be an on-site screening method for monitoring 3-PBA or pyrethroid residues in environmental samples.

New Carbothioamide and Carboxamide Derivatives of 3-Phenoxybenzoic Acid as Potent VEGFR-2 Inhibitors: Synthesis, Molecular Docking, and Cytotoxicity Assessment.

INTRODUCTION/BACKGROUND: Because of the well-established link between angiogenesis and tumor development, the use of antiangiogenic therapeutics, such as those targeting VEGFR-2, presents a promising approach to cancer treatment. In the current study, a set of five hydrazine-1-- carbothioamide (compounds 3a-e) and three hydrazine-1-carboxamide derivatives (compounds 4a-c) were successfully synthesized from 3-phenoxybenzoic acid. These compounds were specially created as antiproliferative agents with the goal of targeting cancer cells by inhibiting VEGFR-2 tyrosine kinase. MATERIALS AND METHODS: The new derivatives were synthesized by conventional organic methods, and their structure was versified by IR, 1HNMR, 13CNMR, and mass spectroscopy. In silico investigation was carried out to identify the compounds' target, molecular similarity, ADMET, and toxicity profile. The cytotoxic activity of the prepared compounds was evaluated in vitro against three human cancer cell lines (DLD1 colorectal adenocarcinoma, HeLa cervical cancer, and HepG2 hepatocellular carcinoma). The effects of the leading compound on cell cycle progression and apoptosis induction were investigated by flow cytometry, and the specific apoptotic pathway triggered by the treatment was evaluated by RT-PCR and immunoblotting. Finally, the inhibitory activities of the new compounds against VEGFR-2 was measured. RESULTS: The designed derivatives exhibited comparable binding positions and interactions to the VEGFR-2 binding site to that of sorafenib (a standard VEGFR-2 tyrosine kinase inhibitor), as determined by molecular docking analysis. Compound 4b was the most cytotoxic compound, achieving the lowest IC50 against HeLa cells. Compound 4b, a strong representative of the synthesized series, induced cell cycle arrest at the G2/M phase, increased the proportion of necrotic and apoptotic HeLa cells, and activated caspase 3. The EC50 value of compound 4b against VEGFR-2 kinase activity was comparable to sorafenib's. CONCLUSION: Overall, the findings suggest that compound 4b has a promising future as a starting point for the development of new anticancer drugs.