RÉSUMÉ
Phenolic acids still gain significant attention due to their potential antimicrobial and cytotoxic properties. In this study, we have investigated the antimicrobial of six phenolic acids, namely chlorogenic, caffeic, p-coumaric, rosmarinic, gallic and tannic acids in the concentration range 0.5-500 µM, against Escherichia coli and Lactobacillus rhamnosus. The antimicrobial activity was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide colorimetric assay. Additionally, the cytotoxic effects of these phenolic acids on two cancer cell lines, the colorectal adenocarcinoma Caco-2 cell line and Dukes' type C colorectal adenocarcinoma DLD-1 cell line was examined. To further understand the molecular properties of these phenolic acids, quantum chemical calculations were performed using the Gaussian 09W program. Parameters such as ionization potential, electron affinity, electronegativity, chemical hardness, chemical softness, dipole moment, and electrophilicity index were obtained. The lipophilicity properties represented by logP parameter was also discussed. This study provides a comprehensive evaluation of the antimicrobial and cytotoxic activity of six phenolic acids, compounds deliberately selected due to their chemical structure. They are derivatives of benzoic or cinnamic acids with the increasing number of hydroxyl groups in the aromatic ring. The integration of experimental and computational methodologies provides a knowledge of the molecular characteristics of bioactive compounds and partial explanation of the relationship between the molecular structure and biological properties. This knowledge aids in guiding the development of bioactive components for use in dietary supplements, functional foods and pharmaceutical drugs.
Sujet(s)
Hydroxybenzoates , Humains , Hydroxybenzoates/composition chimique , Hydroxybenzoates/pharmacologie , Cellules Caco-2 , Lignée cellulaire tumorale , Escherichia coli/effets des médicaments et des substances chimiques , Anti-infectieux/pharmacologie , Anti-infectieux/composition chimique , Tests de sensibilité microbienne , Acide gallique/composition chimique , Acide gallique/pharmacologie , Cinnamates/composition chimique , Cinnamates/pharmacologie , Acides caféiques/composition chimique , Acides caféiques/pharmacologie , Acides coumariques/composition chimique , Acides coumariques/pharmacologieRÉSUMÉ
Mesotrione, as a widely used herbicide, is present in the environment in detectable amounts, causing serious damage. Here, we aimed to investigate the effect of mesotrione on Caco-2 cells and the possibility of its toxicity mitigation by cichoric acid. Therefore, we analyzed the cytotoxicity of both these compounds and the selected oxidative stress parameters, apoptosis and interaction of both the tested compounds with the cell membrane and their accumulation within the cells. In cytotoxicity studies, the stimulating activity of mesotrione was observed, and simultaneously, the inhibitory effect of cichoric acid was noticed. This effect was related to the results of oxidative stress analysis and apoptosis measurements. The activity level of key enzymes (glutathione peroxidase, catalase and superoxide dismutase) in Caco-2 cells exposed to cichoric acid was higher as compared to that of the control. The treatment with mesotrione did not induce apoptosis in the Caco-2 cells. The penetration of the studied compounds into the Caco-2 cells was measured by using an HPLC methodology, and the results indicate mesotrione's high penetration capacity. The distribution of charge on the surface of the cell membranes changed under the influence of both compounds. Considering the mutual interactions of beneficial and potentially toxic food ingredients, it should be noted that, despite the observed favorable trend, cichoric acid is not able to overcome the toxic and cancer-stimulating effects of this pesticide.
Sujet(s)
Apoptose , Acides caféiques , Cyclohexanones , Stress oxydatif , Humains , Cellules Caco-2 , Apoptose/effets des médicaments et des substances chimiques , Cyclohexanones/pharmacologie , Stress oxydatif/effets des médicaments et des substances chimiques , Acides caféiques/pharmacologie , Succinates/pharmacologie , Tumeurs colorectales/traitement médicamenteux , Tumeurs colorectales/métabolisme , Tumeurs colorectales/anatomopathologie , Adénocarcinome/traitement médicamenteux , Adénocarcinome/anatomopathologie , Adénocarcinome/métabolisme , Herbicides/toxicité , Superoxide dismutase/métabolisme , Survie cellulaire/effets des médicaments et des substances chimiques , Catalase/métabolisme , Glutathione peroxidase/métabolisme , Membrane cellulaire/effets des médicaments et des substances chimiques , Membrane cellulaire/métabolismeRÉSUMÉ
This paper presents the results of a study of the effects of the lanthanide ions Ce3+, Pr3+, Nd3+ and Sm3+ on the electronic structure and antioxidant and biological (antimicrobial and cytotoxic) properties of p-coumaric acid (p-CAH2). Structural studies were conducted via spectroscopic methods (FTIR, ATR, UV). Thermal degradation studies of the complexes were performed. The results are presented in the form of TG, DTG and DSC curves. Antioxidant properties were determined via activity tests against DPPH, ABTS and OH radicals. The reducing ability was tested via CUPRAC assays. Minimum inhibitory concentrations (MICs) of the ligand and lanthanide complexes were determined on E. coli, B. subtilis and C. albicans microorganisms. The antimicrobial activity was also determined using the MTT assay. The results were presented as the relative cell viability of C. albicans, P. aeruginosa, E. coli and S. aureus compared to controls and expressed as percentages. In the obtained complexes in the solid phase, lanthanide ions coordinate three ligands in a bidentate chelating coordination mode through the carboxyl group of the acid. Spectroscopic analysis showed that lanthanide ions increase the aromaticity of the pi electron system of the ligand. Thermal analysis showed that the complexes are hydrated and have a higher thermal stability than the ligand. The products of thermal decomposition of the complexes are lanthanide oxides. In the aqueous phase, the metal combines with the ligand in a 1:1 molar ratio. Antioxidant activity tests showed that the complexes have a similar ability to remove free radicals. ABTS and DPPH tests showed that the complexes have twice the ability to neutralise radicals than the ligand, and a much higher ability to remove the hydroxyl radical. The abilities of the complexes and the free ligand to reduce Cu2+ ions in the CUPRAC test are at a similar level. Lanthanide complexes of p-coumaric acid are characterised by a higher antimicrobial capacity than the free ligand against Escherichia coli bacteria, Bacillus subtilis and Candida albicans fungi.
RÉSUMÉ
An increasing level of pesticide exposition is being observed as a result of the consumption of large amounts of fruits, vegetables and grain products, which are key components of the vegetarian diet. Fungicides have been classified as endocrine-disrupting compounds, but their mechanisms of action have not yet been clarified. The effect of boscalid (B), cyprodinil (C) and iprodione (I) combined with Tamoxifen (T) and 17ß-estradiol (E2) on cell viability, cell proliferation, reporter gene expression, ROS content, the cell membrane's function, cell morphology and antioxidant enzymes gene expression in MCF-7 and T47D-KBluc cell lines were investigated. The cell lines were chosen due to their response to 17ß -estradiol. The selected fungicides are commonly used in Poland to protect crops against fungi. Our results revealed that the studied fungicides caused significant increases in cell viability and proliferation, and estrogenic activity was present in all studied compounds depending on their concentrations. Oxidative stress activated uncontrolled cancer cell proliferation by inducing ROS production and by inhibiting antioxidant defense. Our findings verify that the studied fungicides could possibly exhibit endocrine-disrupting properties and exposure should be avoided.
Sujet(s)
Oestrogènes , Fongicides industriels , Oestrogènes/pharmacologie , Fongicides industriels/pharmacologie , Antioxydants/pharmacologie , Espèces réactives de l'oxygène , Oestradiol/pharmacologie , OestroneRÉSUMÉ
Chlorpyrifos (CPF) was the most frequently used pesticide in food production in the European Union (EU) until 2020. Unfortunately, this compound is still being applied in other parts of the world. National monitoring of pesticides conducted in various countries indicates the presence of CPF in soil, food, and water, which may have toxic effects on consumers, farmers, and animal health. In addition, CPF may influence changes in the population of fungi, bacteria, and actinomycete in soil and can inhibit nitrogen mineralization. The mechanisms of CPF activity are based on the inhibition of acetylcholinesterase (AChE) activity. This compound also exhibits reproductive toxicity, neurotoxicity, and genotoxicity. The problem seems to be the discrepancy between the actual observations and the final conclusions drawn for the substance's approval in reports presenting the toxic impact of CPF on human health. Therefore, this influence is still a current and important issue that requires continuous monitoring despite its withdrawal from the market in the EU. This review traces the scientific reports describing the effects of CPF resulting in changes occurring in both the environment and at the cellular and tissue level in humans and animals. It also provides an insight into the hazards and risks to human health in food consumer products in which CPF has been detected.
Sujet(s)
Chlorpyriphos , Insecticides , Pesticides , Acetylcholinesterase , Animaux , Chlorpyriphos/toxicité , Humains , Azote , Sol , EauRÉSUMÉ
The carriers of nosocomial infections are the hands of medical personnel and inanimate surfaces. Both hands and surfaces may be contaminated as a result of contact with the patient, their body fluids, and touching contaminated surfaces in the patient's surroundings. Visually clean inanimate surfaces are an important source of pathogens. Microorganisms have properties thanks to which they can survive in unfavorable conditions, from a few days to several months. Bacteria, viruses and fungi are able to transmit from inanimate surfaces to the skin of the patient and the medical staff. These pathogens include SARS-CoV-2, which can survive on various types of inanimate surfaces, being a potential source of infection. By following the recommendations related to washing and disinfecting hands and surfaces, and using appropriate washing and disinfecting agents with a broad biocidal spectrum, high material compatibility and the shortest duration of action, we contribute to breaking the chain of nosocomial infections.
Sujet(s)
COVID-19 , Infection croisée , Virus , Bactéries , Infection croisée/épidémiologie , Infection croisée/microbiologie , Infection croisée/prévention et contrôle , Champignons , Humains , SARS-CoV-2RÉSUMÉ
The structural, spectral, thermal, and biological properties of hydroxyphenylacetic acid and lithium, sodium, potassium, rubidium, and cesium 2-hydroxyphenylacetates were analyzed by means of infrared spectroscopy FT-IR, electronic absorption spectroscopy UV-VIS, nuclear magnetic resonance 1H and 13C NMR, thermogravimetric analysis (TG/DSC), and quantum-chemical calculations at B3LYP/6-311++G** level. Moreover, the antioxidant (ABTS, FRAP, and CUPRAC assays), antibacterial (against E. coli, K. aerogenes, P. fluorescens, and B. subtilis) and antifungal (against C. albicans) properties of studied compounds were measured. The effect of alkali metal ions on the structure, thermal, and biological properties of 2-hydroxyphenylacetates was discussed.
RÉSUMÉ
Leachate from landfills is a product of complex biological and physicochemical processes occurring during waste storage. In the present study, the toxicity of landfill leachate (LL) to human and bacterial cells was investigated for better understanding of LL environmental toxicity. Studies regarding LL physicochemical properties and cytotoxicity analysis were conducted. In Escherichia coli, Pseudomonas fluorescens, Bacillus subtilis, fibroblasts and melanoma A-375 cells, cell viability assays were applied. For the determination of LL antibacterial activity, twofold dilution series of LL were prepared in the range from 50% to 0.1% (50%, 25%, 12.5%, 6.25%, 3.13%, 1.56%, 0.78%, 0.39%, 0.2%, 0.1%). Human cells viability was examined at LL concentrations of 0.1%, 0.5%, 1%, 1.5%, 2%, 2.5%, 5%, 10%, 15%, 20% and 30%. ROS (reactive oxygen species) content and apoptosis level were also measured in bacterial and human cells under the influence of LL. Unexpectedly obtained results indicate stimulation of bacterial viability by LL. Fibroblasts under the influence of LL showed decrease in their viability and increase in apoptosis level and A-375 melanoma cells showed an increase in relative viability and decrease in apoptosis. ROS level in bacterial cells was elevated in higher LL concentrations and decreased in lower LL concentrations. In human cells, ROS content was rather high in both tested cell lines. Presented results indicate cytotoxic potential of analyzed LL and the necessity of LL monitoring because it may pose a health hazard for exposed human populations and the whole human environment.
Sujet(s)
Nappe phréatique , Élimination des déchets , Polluants chimiques de l'eau , Surveillance de l'environnement , Humains , Installations d'élimination des déchets , Polluants chimiques de l'eau/analyse , Polluants chimiques de l'eau/toxicitéRÉSUMÉ
The interaction of different food ingredients is now a very important and often emerging topic of research. Pesticides and their breakdown products, which may be carcinogenic, are one of the frequently occurring food contaminants. Compounds like traumatic acid (TA), which originates from plants, are beneficial, antioxidant, and anticancer food ingredients. Previously obtained results from our research group indicated antioxidative in normal human fibroblasts and prooxidative in cancer cells activity of TA. Since the literature data show an undoubted connection between the presence of pesticides in food and the increased incidence of different types of cancers, we attempted to clarify whether TA can abolish the effect of mesotrione stimulating the growth of cancer cells. In order to study the influence of mesotrione on breast cancer cells, we decided to carry out cytotoxicity studies of environmentally significant herbicide concentrations. We also analyzed the cytotoxicity of TA and mixtures of these two compounds. After selecting the most effective concentrations of both components tested, we conducted analyses of oxidative stress parameters and apoptosis in ZR-75-1 cells. The obtained results allow us to conclude that traumatic acid by stimulating oxidative stress and apoptosis contributes to inhibiting the growth and development of cells of the ZR-75-1 line strengthened by mesotrione. This may mean that TA is a compound with pro-oxidative and proapoptotic effects in cancer cells whose development and proliferation are stimulated by the presence of mesotrione. The presented results may be helpful in answering the question of whether herbicides and their residues in edibles may constitute potential threat for people diagnosed with cancer and whether compounds with proven pro-oxidative effects on cancer cells can have potential cytoprotective functions.
Sujet(s)
Antinéoplasiques d'origine végétale , Antioxydants , Tumeurs du sein/anatomopathologie , Prolifération cellulaire/effets des médicaments et des substances chimiques , Cyclohexanones/effets indésirables , Cytoprotection/effets des médicaments et des substances chimiques , Diacides carboxyliques/pharmacologie , Contamination des aliments , Herbicides/effets indésirables , Apoptose/effets des médicaments et des substances chimiques , Lignée cellulaire tumorale , Cellules cultivées , Femelle , Humains , Stress oxydatif/effets des médicaments et des substances chimiquesRÉSUMÉ
Cichoric acid (CA) belongs to the group of polyphenols, which occurs in a variety of plant species and it is characterized by anticancer, antibacterial, and antiviral properties. Selected polyphenols have the ability to combine with metal ions to form chelate complexes that reveal greater biological activity than free compounds. In order to study possible antimicrobial and anticancer effect of CA and its complexes with copper(II)/zinc(II)/nickel(II)/cobalt(II) we decided to conduct cytotoxicity tests to estimate the most effective concentrations of tested compounds. The results of the presented study demonstrated, for the first time, that the treatment with newly synthesized CA-metal complexes has anticancer and antimicrobial effects, which were examined in seven different cell lines: MCF-7, MDA-MB-231, and ZR-75-1 breast cancer cell lines, A375 melanoma cell line, DLD-1 cell line, LN-229 cell line, FN cell line; five bacterial strains: Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, Proteus vulgaris, Lactobacillus rhamnosus, yeast Sacchcaromyces boulardii, and pathogenic yeast-like fungi Candida albicans. The presented study indicates that CA-metal complexes could be considered as a potential supplementary tool in anticancer therapy, however, because of their possible toxic activity on fibroblasts, they should be used with caution. Some of the tested complexes have also preservative properties and positive influence on normal non-pathogenic microorganisms, which was demonstrated in selected microbial strains, therefore they may serve as food preservatives of natural origin with cytoprotective properties.
Sujet(s)
Anti-infectieux/administration et posologie , Antinéoplasiques/administration et posologie , Bactéries/effets des médicaments et des substances chimiques , Acides caféiques/pharmacologie , Complexes de coordination/administration et posologie , Extraits de plantes/pharmacologie , Succinates/pharmacologie , Levures/effets des médicaments et des substances chimiques , Anti-infectieux/effets indésirables , Anti-infectieux/pharmacologie , Anti-infectieux/usage thérapeutique , Antinéoplasiques/effets indésirables , Antinéoplasiques/pharmacologie , Antinéoplasiques/usage thérapeutique , Acides caféiques/usage thérapeutique , Candida albicans/effets des médicaments et des substances chimiques , Lignée cellulaire tumorale , Complexes de coordination/effets indésirables , Complexes de coordination/pharmacologie , Complexes de coordination/usage thérapeutique , Fibroblastes/effets des médicaments et des substances chimiques , Humains , Ions , Métaux/pharmacologie , Métaux/usage thérapeutique , Tests de sensibilité microbienne , Tumeurs/traitement médicamenteux , Extraits de plantes/usage thérapeutique , Saccharomyces boulardii/effets des médicaments et des substances chimiques , Succinates/usage thérapeutiqueRÉSUMÉ
Bifenox and Dichlobenil belong to the commonly used in Poland in agriculture group of herbicides and their residues are often detected in the environment. They are poorly known regarding their possible carcinogenic and antibacterial effect at the cellular level. Therefore, we decided to study their activity in bacterial strains Aliivibrio fisheri,E. coli, P. aeruginosa, and C. albicans (yeast) and human cancer ZR-75-1 cells. Compounds under study exhibit stimulatory effect on analyzed bacterial strains. The study performed on mammalian cells better reflects the influence of environmental pollutants on human organism, therefore we evaluated the effect of herbicides on ZR-75-1 cells. Cells viability, apoptosis and selected oxidative stress parameters in ZR-75-1 cells were investigated. Both analyzed substances exhibit stimulatory effects on analyzed parameters, however they do not stimulate apoptosis which correlate positively with the induction of oxidative stress. Bifenox and Dichlobenil enhance oxidative stress parameters by the generation of high levels of ROS, which can lead to their adaptation and resistance to the standard treatment regimen.
Sujet(s)
Bactéries/effets des médicaments et des substances chimiques , Survie cellulaire/effets des médicaments et des substances chimiques , Herbicides/toxicité , Nitriles/toxicité , Éthers phényliques/toxicité , Lignée cellulaire tumorale , Polluants environnementaux/analyse , Humains , Stress oxydatifRÉSUMÉ
The main consequence of herbicides use is the presence of their residues in food of plant origin. A growing body of evidence indicates that herbicides cause detrimental effects upon human health while demonstrating a direct link of pesticides exposure with the occurrence of human chronic diseases, including cancer. There is a pressing need to develop our knowledge regarding interactions of food contaminants and food components both in vitro and in vivo. Pesticides are highly undesirable food contaminants, and traumatic acid (TA) is a very beneficial food ingredient, therefore we decided to study if TA may act as a compound that delays the stimulatory effect of pesticides on breast cancer cells. To analyze the potential effects that selected herbicides (MCPA, mesotrione, bifenox and dichlobenil) may have upon cancerous cells, we conducted studies of the cytotoxicity of physiological concentrations of four pesticides and the mix of TA with tested herbicides in three different breast cancer cell lines (MCF-7, ZR-75-1 and MDA-MB-231) and one normal healthy breast cell line MCF-12A. Based on the obtained results we conclude that TA in a concentration-dependent manner might influence selected effects of the studied herbicides for particular cancer cells lines.
Sujet(s)
Diacides carboxyliques/toxicité , Herbicides/toxicité , Tumeurs du sein , Lignée cellulaire tumorale , Survie cellulaire/effets des médicaments et des substances chimiques , Femelle , HumainsRÉSUMÉ
Traumatic acid (TA) - an oxidative derivative of unsaturated fatty acids, belongs to the cytokinins category - a group of plant hormones, which play an important role in growth and development. Previously we demonstrated its positive influence on oxidative stress parameters in normal human fibroblasts, therefore we decided to investigate its activity in cancer cells. MCF-7 breast cancer cell line was chosen as an experimental model because of proved association between the consumption of dietary fat and the incidence of breast cancer. TA cytotoxicity and its effects on MCF-7â¯cells proliferation, viability, apoptosis/necrosis, thiol group content, lipid peroxidation, reduced/oxidized glutathione (GSH/GSSG) and ROS (reactive oxygen species) content was examined. The results show a significant effect of TA on tested parameters. TA caused a decrease in cells proliferation and viability, GSH/GSSG ratio and thiol group content. It increases caspase 7 activity, membrane lipid peroxidation and ROS content, simultaneously reducing breast cancer cell growth through oxidative stress influence on apoptosis. The present findings reveal that TA exhibits multiple and complex activity in MCF-7 breast cancer cells and it exhibits potential anticancer properties and tumor preventive activity.
Sujet(s)
Antinéoplasiques/pharmacologie , Tumeurs du sein/traitement médicamenteux , Diacides carboxyliques/pharmacologie , Antinéoplasiques/effets indésirables , Apoptose/effets des médicaments et des substances chimiques , Tumeurs du sein/métabolisme , Tumeurs du sein/anatomopathologie , Prolifération cellulaire/effets des médicaments et des substances chimiques , Survie cellulaire/effets des médicaments et des substances chimiques , Diacides carboxyliques/effets indésirables , Relation dose-effet des médicaments , Tests de criblage d'agents antitumoraux , Femelle , Humains , Peroxydation lipidique/effets des médicaments et des substances chimiques , Cellules MCF-7 , Relation structure-activité , Cellules cancéreuses en cultureRÉSUMÉ
This paper analyses sources of sixteen PAHs - polycyclic aromatic hydrocarbons in urbanized areas by using selected diagnostic ratios. Simultaneously, an attempt was made to determine how sewage sludge changes PAHs content in urbanized areas soils. In the experiment three lawns along the main roads in Bialystok with different traffic intensity, three doses of sewage sludge and two years of study were considered. There was no effect of fertilization with sewage sludge on the sum of 16 PAHs in urban soil samples, nevertheless, the sum of 16 PAHs was reduced from 2.6 in 2011 to 2.3 mg/kg in 2012. Among 16 tested PAHs compounds, benzo[a]pyrene was the most dominant compound in samples collected in both years - about 15% of all PAHs. The results suggest that application of sludge into the soil did not influence the concentration of 2-3-ring, 4-ring and 5-6-ring PAHs. For the objects fertilized with a dose 150.0 Mg/ha, of sludge the total sum of potentially carcinogenic PAHs in the urban soil lowered by approximately 68% in comparison with the control plots. PAHs contamination of the urban soil samples resulted from the influence of coal, petroleum and biomass combustion. Moreover, PAHs can enter soil via at mospheric deposition.
Sujet(s)
Hydrocarbures aromatiques polycycliques/analyse , Polluants du sol/analyse , Élimination des déchets liquides/méthodes , Benzo[a]pyrène/analyse , Charbon/analyse , Engrais , Eaux d'égout/analyse , Sol/composition chimique , UrbanisationRÉSUMÉ
The present study aimed at evaluating the dissipation of S-metolachlor (S-MET) at three doses in maize growing on diverse physico-chemical properties of soil. The effect of herbicide on dehydrogenase (DHA) and acid phosphatase (ACP) activity was estimated. A modified QuEChERS method using LC-MS/MS has been developed. The limit of quantification (0.001 mg kg-1) and detection (0.0005 mg kg-1) were very low for soil and maize samples. The mean recoveries and RSDs for the six spiked levels (0.001-0.5 mg kg-1) were 91.3 and 5.8%. The biggest differences in concentration of S-MET in maize were observed between the 28th and 63rd days. The dissipation of S-MET in the alkaline soil was the slowest between the 2nd and 7th days, and in the acidic soil between the 5th and 11th days. DT50 of S-MET calculated according to the first-order kinetics model was 11.1-14.7 days (soil) and 9.6-13.9 days (maize). The enzymatic activity of soil was higher in the acidic environment. One observed the significant positive correlation of ACP with pH of soil and contents of potassium and magnesium and negative with contents of phosphorus and organic carbon. The results indicated that at harvest time, the residues of S-MET in maize were well below the safety limit for maize. The findings of this study will foster the research on main parameters influencing the dissipation in maize ecosystems.
Sujet(s)
Acétamides/analyse , Surveillance de l'environnement , Herbicides/analyse , Polluants du sol/analyse , Chromatographie en phase liquide , Écosystème , Cinétique , Sol/composition chimique , Microbiologie du sol , Spectrométrie de masse en tandem , Zea mays/composition chimiqueRÉSUMÉ
Pesticides cause serious environmental and health problems both to humans and animals. The aim of this review is to discuss selected herbicides and fungicides regarding their mode of action and their influence on basic oxidative stress parameters and endocrine disruption properties tested in selected cell cultures in vitro. Because of numerous difficulties which animal studies are subject to, cell cultures are an excellent experimental model reflecting human exposure to different pesticides through all relevant routes. This experimental model can be used to monitor aggregate and cumulative pesticide exposures.