Enzymes Ethoxyresorufin O-deethylase Research Articles

Behind conventional (micro)plastics: An ecotoxicological characterization of aqueous suspensions from End-of-Life Tire particles

Million tons of tires become waste every year, and the so-called End-of-Life Tires (ELTs) are ground into powder (ELT-dp; size < 0.8 mm) and granules (ELT-dg; 0.8 < size < 2.5 mm) for recycling. The aim of this study was to evaluate the sub-lethal effects of three different concentrations (0.1, 1, and 10 mg/L) of aqueous suspensions from ELT-dp and ELT-dg on Danio rerio (zebrafish) larvae exposed from 0 to 120 h post-fertilization (hpf). Chronic effects were assessed through biomarkers, real-time PCR, and proteomics. We observed a significant increase in swimming behavior and heart rate only in specimens exposed to ELT-dp suspensions at 1 and 10 mg/L, respectively. Conversely, the activities of detoxifying enzymes ethoxyresorufin-O-deethylase (EROD) and glutathione-S-transferase (GST) showed significant modulation only in specimens exposed to ELT-dg groups. Although no effects were observed through real-time PCR, proteomics highlighted alterations induced by the three ELT-dp concentrations in over 100 proteins involved in metabolic pathways of aromatic and nitrogen compounds. The results obtained suggest that the toxic mechanism of action (MoA) of ELT suspensions is mainly associated with the induction of effects by released chemicals in water, with a higher toxicity of ELT-dp compared to ELT-dg.

Chronic Effects of Carbamazepine, Progesterone and Their Mixtures at Environmentally Relevant Concentrations on Biochemical Markers of Zebrafish (Danio rerio).

The impact of pharmaceuticals on non-target organisms in the environment is of increasing concern and study. Pharmaceuticals and other pollutants are often present as mixtures in an environmental compartment. Studies on the toxicological implications of these drugs on fish, particularly as mixtures at environmentally relevant concentrations, are very limited. Thus, this study aimed to evaluate the chronic effects of the anticonvulsant drug carbamazepine (CBZ) and progesterone (P4) at environmentally relevant concentrations, individually and in binary mixtures, applying a suite of biomarkers at the molecular level in zebrafish (Danio rerio). The effects on biotransformation enzymes 7-ethoxyresorufin O-deethylase (EROD) and glutathione-S-transferase (GST), antioxidant enzymes catalase (CAT), superoxide dismutase (SOD), glutathione peroxidases (GPxSe and GPxTOT), and glutathione reductase (GR), and markers of damage, such as DNA strand breaks (DNAsb), lactate dehydrogenase (LDH), lipid peroxidation (LPO), and vitellogenin-like proteins (VTG), were evaluated. Analyses of the biochemical markers indicated that a synergistic dose-ratio-dependent effect of CBZ and P4 in zebrafish occurs after chronic exposure regarding VTG, biotransformation enzymes (EROD, GST), and oxidative stress marker (DNAsb). The results suggest a synergistic effect regarding VTG, thus indicating a high risk to the reproductive success of fish if these pharmaceuticals co-occur.

Synthesis and Antioxidant Properties of New Benzimidazole Derivatives

In this study, new benzimidazole derivatives were synthesized. After purity checks were made with Thin Layer Chromatography (TLC) and melting points, their structures were proved by mass spectroscopy, 1H, 13C NMR and elemental analysis technique. Antioxidant activities of the obtained materials were searched in terms of their effect on 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, lipid peroxidation inhibition and ethoxyresorufin O-deethylase (EROD) enzyme activity. It was observed the compounds were generally moderately active in terms of DPPH radical scavenging activity and lipid peroxidation inhibition compared to the standard compound and had a good effect on EROD enzyme activity.

In Vivo Effects on Stress Protein, Genotoxicity, and Oxidative Toxicity Parameters in Oreochromis niloticus Tissues Exposed to Thiamethoxam

This study aimed to observe the effect of toxicity of the pesticide thiamethoxam (TMX) at sublethal concentrations in the liver and brain of Oreochromis niloticus. In the experiment, fish were exposed to 50, 100, and, 150 mg/L with thiamethoxam for 48 h and 15 days. The superoxide dismutase (SOD), catalase (CAT), glutathione S-transferaz (GST), glutathione peroxidase (GPx), and ethoxyresorufin-O-deethylase (EROD) activities; and thiobarbituric acid reactive substance (TBARS), heat shock proteins 70 (HSP70), glutathione (GSH), and genotoxicity parameter 8-hydroxy-2′-deoxyguanosine (8-OHdG) were analyzed by spectrophotometric methods and ELISA techniques. Depending on time and dose in TMX exposure in liver tissue, a significant decrease in GSH level; an increase in SOD, GST, GPx, and EROD enzyme activities; and HSP70, TBARS, and 8-OHdG levels was determined. In brain tissue, SOD, GST, and EROD enzyme activities, an increase in HSP70, TBARS, and 8-OHdG levels, and a decrease in CAT enzyme activity and GSH levels were determined. In this study, TMX in the concentrations used showed that changes in oxidative stress biomarkers, genotoxicity parameter 8-OHdG levels, and HSP70 levels caused toxic effects in the model organism. As a result of the study, the changes and protective effects of the antioxidant system and stress proteins at the cellular level were determined in sublethal doses of toxic effects caused by TMX in the vital organs of the organism. In this toxicological study, TMX exposure resulted in toxicity to O. niloticus liver and brain tissues, in addition, responses of biomarkers to time and concentrations were determined. As a result of this study, the potential toxic effects of the commonly used pesticide TMX will reveal both the ecological risks of the aquatic organism and the basic data of the safety and risk assessments of O. niloticus consumed as food for human health.

Multiple biomarker responses in caged benthic gastropods Bellamya aeruginosa after in situ exposure to Taihu Lake in China

BackgroundFreshwater sediments have been recognized as a long-term sink and potential source for environmental pollutants released into the aquatic ecosystems. In this study, the sediment quality of Taihu Lake, which is susceptible to anthropogenic contamination, was assessed by a combination of chemical analytical and biological end points. Specifically, the snail Bellamya aeruginosa was caged in situ at two locations representing different pollution levels for different exposure times (7, 14 and 21 days). At each of these time points, biochemical parameters, i.e., phase I biotransformation enzymes ethoxyresorufin-O-deethylase (EROD), the antioxidant enzymes superoxide dismutase and catalase, reactive oxygen species, protein carbonyl content and lipid peroxidation, were evaluated in the hepatopancreas of snails. In addition, surface sediments were collected for analysis of contaminants of concern, including inorganic pollutants, organochlorine pesticides, polychlorinated biphenyls and polybrominated diphenyl ethers.ResultsChemical analyses revealed that sediments from Taihu Lake were contaminated with trace elements and organic pollutants. Concentrations of trace elements (Cu, Ni and As) and organochlorinated pesticides (4,4′-DDE) exceeded their corresponding threshold effect level according to the sediment quality assessment values for freshwater ecosystems in Canada, indicating that adverse biological effects may occur. All biomarkers, except EROD activity, were induced in snails during all exposure times. The integrated biomarker response index (IBR) indicated that during the initial exposure phase (7 days), B. aeruginosa were subjected to significant environmental stress, which diminished during later sampling time points.ConclusionsResults showed that IBR correlated well with the levels of environmental contaminants, demonstrating the applicability of this biomonitoring approach to complex environmental exposure scenarios.

Interactive effects of polystyrene microplastics and roxithromycin on bioaccumulation and biochemical status in the freshwater fish red tilapia (Oreochromis niloticus)

There are hundreds of thousands metric tons of microplastics (MPs) present in aquatic environments. The MPs coexist with other pollutants in water bodies, such as pharmaceuticals, and may carry and transfer them into aquatic organisms, consequently causing unpredictable ecological risks. The purpose of the present study was to evaluate the effect of the presence of polystyrene microplastics (PS-MPs) on the distribution and bioaccumulation of roxithromycin (ROX) in freshwater fish red tilapia (Oreochromis niloticus) as well as their interactive biochemical effects in red tilapia. PS-MPs were found to enhance the bioaccumulation of ROX in fish tissues compared to ROX-alone exposure. In the treatment of PS-MPs (100 μg L−1) combined with ROX (50 μg L−1), the highest concentrations of ROX reached 39,672.9 ± 6311.4, 1767.9 ± 277.8, 2907.5 ± 225.0, and 4307.1 ± 186.5 μg kg−1 in gut, gills, brain, and liver, respectively. Furthermore, compared to the ROX alone, the neurotoxicity caused by ROX was alleviated due to the presence of MPs after 14 d of exposure. The activities of cytochrome P450 (CYP) enzymes [7-ethoxyresorufin O-deethylase (EROD) and 7-benzyloxy-4-trifluoromethyl-coumarin O-dibenzyloxylase (BFCOD)] in fish livers exposed to all co-exposure treatments exhibited great variability compared to ROX alone after 14 d of exposure, suggesting that the presence of MPs may affect the metabolism of ROX in tilapia. Compared with ROX alone, the superoxide dismutase (SOD) activity increased significantly, and malondialdehyde (MDA) contents decreased in the co-exposure treatments, showing that oxidative damage in situations of co-exposure to MPs and ROX was mitigated in fish livers after 14 d of exposure. Collectively, the presence of MPs could affect the fate and toxicity of other organic pollutants in fish. The results emphasize the importance to study the interactions between MPs and other organic pollutants in aquatic environments.

Accumulation, tissue distribution, and biochemical effects of polystyrene microplastics in the freshwater fish red tilapia (Oreochromis niloticus)

While the presence of microplastics (MPs) in marine environments has been detected worldwide, the importance of MPs pollution in freshwater environments has also been emphasized in recent years. However, the body of knowledge regarding the biological effects of MPs on freshwater organisms is still much more limited than on marine organisms. The aim of the present study was to evaluate the accumulation and tissue distribution of MPs in the freshwater fish red tilapia (Oreochromis niloticus), as well as the biochemical effects of MPs on O. niloticus. During 14 days of exposure to 0.1 μm polystyrene-MPs at concentrations of 1, 10, and 100 μg L−1, the MPs concentrations in various tissues of O. niloticus generally increased over time following the order gut > gills > liver ≈ brain. Moreover, the acetylcholinesterase (AChE) activity in the fish brain was inhibited by MPs exposure, with a maximum inhibition rate of 37.7%, suggesting the potential neurotoxicity of MPs to freshwater fish. The activities of cytochrome P450 (CYP) enzymes [7-ethoxyresorufin O-deethylase (EROD) and 7-benzyloxy-4-trifluoromethyl-coumarin O-dibenzyloxylase (BFCOD)] in the fish liver exhibited clear temporal variabilities, with significant decreases followed by elevations compared to the control. The alterations of the EROD and BFCOD activities indicate the potential involvement of CYP enzymes for the metabolism of MPs. The activity of antioxidative enzyme superoxide dismutase (SOD) in the liver was significantly induced throughout the exposure period, while the malondialdehyde (MDA) content did not vary with MPs exposure, suggesting that the antioxidative enzymatic system in O. niloticus could prevent oxidative damage. These results highlight the ingestion and accumulation of MPs in different tissues of freshwater fish, which lead to perturbations in fish biological systems and should be considered in environmental risk assessment.

Evaluation of caffeine effects on biochemical and genotoxic biomarkers in the neotropical freshwater teleost Prochilodus lineatus

Caffeine is often found in aquatic environments, leading to concerns regarding its adverse consequences for aquatic biota. Biochemical and genotoxic biomarkers were analysed in juveniles of Prochilodus lineatus to evaluate the effects of caffeine. Fish were exposed to caffeine (0.3, 3 and 30 μg L−1) for either 24 h or 168 h. Longer exposure to caffeine resulted in a significant reduction in the activity of the phase I biotransformation enzyme ethoxyresorufin-O-deethylase (EROD) in the brain but a significant increase in the liver. Changes in glutathione content (GSH), glutathione S-transferase (GST) activity, and lipid peroxidation were not found in the liver and brain of fish exposed to caffeine. DNA damage in erythrocytes were also not found. These results show that caffeine may interfere with the biotransformation mechanism of P. lineatus after 168 h exposure, but it does not generate sufficient changes to trigger a state of oxidative stress.

A comparative approach using biomarkers in feral and caged Neotropical fish: Implications for biomonitoring freshwater ecosystems in agricultural areas

The aim of this study was to investigate the responses of biomarkers in feral and caged fish and the capacity of these biomarkers to discriminate contamination levels along a stream located in an agricultural area in Southern Brazil. Specimens of the Neotropical fish, Astyanax altiparanae, were confined for 168h in three lakes along the stream. Additionally, during the weeks of in situ exposure, wild specimens of this species were collected from the same sites. Biochemical biomarkers were analyzed, such as phase I biotransformation enzyme 7-ethoxyresorufin-O-deethylase (EROD) and phase II biotransformation enzyme glutathione S-transferase, and we also determined hepatic and branchial levels of non-protein thiols (NPSH), oxidative damage such as lipid peroxidation (LPO), and acetylcholinesterase (AChE) activity in muscle and brain. Genetic biomarkers such as DNA breaks (comet assay), frequency of micronuclei (MN) and erythrocytic nuclear abnormalities (ENA) were also examined. The results indicate that the most sensitive biomarkers for discriminating contamination levels are DNA breaks, LPO and AChE activity. Similar results were obtained for both caged and feral fish. The biomarkers that reflect the results of cumulative events, such as ENA, were more discriminative for chronically exposed specimens (feral fishes). Analyzing biomarkers using an integrated response index showed that both approaches (using feral and caged A. altiparanae) were effective for discriminating contamination levels along the stream, corroborating the results of chemical analyses for selected pesticides. Taken together, these results highlight the importance of biomarker selection and show that both approaches (caged and feral fish) are satisfactory for evaluating water quality in streams impacted by agricultural activities.

Modulation of erythromycin-induced biochemical responses in crucian carp by ketoconazole.

The individual and combined biochemical responses of erythromycin and ketoconazole have been examined in an organism representative of the aquatic environment, crucian carp (Carassius auratus). The possible interactions between erythromycin and ketoconazole were investigated on the bioaccumulation and the expression of biotransformation enzymes 7-ethoxyresorufin-O-deethylase (EROD) and glutathione S-transferase (GST), and an antioxidant defense enzyme superoxide dismutase (SOD) in fish tissues. After 14days of combined exposure (erythromycin+ketoconazole), the addition of ketoconazole at nominal concentrations of 0.2, 2, and 20μg/L significantly increased the accumulation of erythromycin in fish bile; however, elevated erythromycin accumulation levels were not observed in the other test tissues. The inductions of EROD and SOD activity to erythromycin were inhibited by the combined exposure of ketoconazole in most cases; however, the GST activity returned to normal with exposure time and concentration of combined administration. From the tested pharmaceutical mixtures, it indicated that certain specific combinations may pose some perturbations in biochemical responses in fish and also provide a better understanding of the effects of toxic mixtures.

Biomarker responses in eelpouts from four coastal areas in Sweden, Denmark and Germany

To increase our understanding of possible chemical impacts on coastal fish populations in the Baltic Sea, Kattegat and Skagerrak, the viviparous eelpout (Zoarces viviparus) was used as sentinel species in two major sampling campaigns (spring and autumn) in 16 different coastal sites. Condition factor (CF), liver somatic index (LSI), gonad somatic index (GSI) were measured and the activity of the hepatic enzymes ethoxyresorufin-O-deethylase (EROD), glutathione reductase GR), glutathione S-transferase (GST), catalase (CAT) and muscular activity of acetylcholinesterase (AChE) were assessed. PAH metabolites in bile were also analyzed. The most notable finding in the data set was the low EROD activity in eelpouts collected at the relatively polluted region in Germany compared to the other regions, which could be due to an inhibition of the CYP1A-system or to adaptation to chronic exposure of pollutants in this area. Additionally, low AChE activity was noted in the German region in the autumn campaign and low AChE activity detected in the Danish region in the spring campaign. These differences suggest possible season-specific differences in the use and release of AChE-inhibiting chemicals in the Danish and German regions. Clustering of biomarkers on site level indicated a relationship between CF and GSI and suggested that sites with a high CF contained eelpout that put a larger effort into their larvae development. Clustering of the oxidative stress markers GR, GST and CAT on the individual level reflected a possible coordinated regulation of these enzymes. Overall, the results support the importance of taking into account general regional differences and seasonal variation in biomarker activity when monitoring and assessing the effects of pollution. Despite the expected seasonal variation for most of the measured endpoint, several markers (GSI, EROD and CF) vary similarly between all selected sites in both spring and autumn. This suggests that the differences between sites for these endpoints are independent of season.

Multiple biomarker responses in Prochilodus lineatus subjected to short-term in situ exposure to streams from agricultural areas in Southern Brazil

In order to assess the quality of streams susceptible to contamination by pesticides we apply biochemical and genotoxic biomarkers in the Neotropical fish Prochilodus lineatus submitted to in situ tests. Fish were caged, for 96h, in two streams located in areas with intensive use of pesticides, the Apertados (AP) and the Jacutinga (JC), and in a small stream (Godoy stream — GD) found inside a forest fragment adjacent to a State Park. Biochemical parameters, such as biotransformation enzymes 7-ethoxyresorufin-O-deethylase (EROD) and glutathione-S-transferase (GST), non-protein thiols (NPSH), lipoperoxidation (LPO), protein carbonylation (PCO) and acetylcholinesterase (AChE) were evaluated in various fish organs, as well as genotoxic biomarkers (damage to DNA and occurrence of micronuclei and erythrocyte nuclear abnormalities). Samples of water and sediment were collected for analysis of metals (Cu, Cr, Pb, Ni, Mn, Cd and Zn), organochloride pesticides, and triazine and glyphosate herbicides. We observed an increase in liver GST activity in fish at AP and gill GST activity in fish at JC. An increase in liver LPO was also observed in fish exposed to AP and JC. The same animals also exhibited increased DNA damage and erythrocyte nuclear abnormalities (ENAs) compared to the fish kept in GD. A number of compounds showed concentrations higher than the permitted levels, in particular, dichlorodiphenyltrichloroethane (DDT), its metabolites dichlorodiphenyldichloroethylene (DDE) and dichlorodiphenyldichloroethane (DDD), hexachlorocyclohexanes (HCH), heptachloride, diclofluanid and aldrins. These pesticides were detected at higher concentrations in water and sediment samples from AP, followed by JC and GD. The Integrated Biomarker Response Index (IBR) indicated that AP and JC (AP: 21.7>JC: 18.5>GD: 12.6) have the worst environmental quality. Integrated biomarker analysis revealed that the alterations observed related well with the levels of environmental contaminants, demonstrating the effectiveness of this biomonitoring approach.

Bioconcentration, metabolism and alterations of thyroid hormones of Tris(1,3-dichloro-2-propyl) phosphate (TDCPP) in Zebrafish.

Tris(1,3-dichloro-2-propyl) phosphate (TDCPP) is an organophosphate widely used as a flame retardant, and has frequently been detected in the environment and biota. The present study investigates the bioconcentration and metabolism of TDCPP in fish. Zebrafish embryos (from 2h post-fertilization) were exposed to TDCPP (0, 4, 20 and 100μg/L) for six months. Exposure to TDCPP significantly induced phase I metabolic enzymes 7-ethoxyresorufin O-deethylase (EROD) and 7-methoxyresorufin O-demethylase (MROD) in fish. The mRNA expression of genes related to Phase I and II metabolic enzymes, such as cyp1a1, cyp1b1, cyp1c1 and ugt1ab were also significantly upregulated. Exposure to TDCPP significantly reduced plasma thyroxine (T4) and 3,5,3'-triiodothyronine (T3) levels in females. Chemicals analysis indicated significant levels of TDCPP and its metabolite, bis (1,3-dichloro-2-propyl) phosphate (BDCPP), in the liver. The present study reveals that exposure to low concentrations of TDCPP can cause bioconcentration in fish, and TDCPP can be readily metabolized in liver.

Biomarker responses in the bivalve Chlamys farreri to the water-soluble fraction of crude oil

To investigate the effect of the water soluble fraction of crude oil (WSF) on marine bivalves, the scallop Chlamys farreri was exposed to three WSF concentrations (0.18 mg/L, 0.32 mg/L, and 0.51 mg/L, respectively) in seawater. Petroleum hydrocarbon contents in scallops and a suite of enzymes [7-Ethoxyresorufin-O-deethylase (EROD), aryl hydrocarbon hydroxylase (AHH), glutathione S-transferase (GST), and glutathione peroxidase (GPx)] in gills and digestive glands were monitored over 10 days. The results revealed that WSF affected the activity of the four enzymes in the gills and digestive glands. EROD activity in the gills was significantly induced in most individuals of the three test groups, while in the digestive gland it was significantly induced in the low-concentration group within 4 days but was inhibited in the middle- and high-concentration groups on days 1, 4, and 10. AHH activity in the gills of all treatment groups was significantly induced on day 1. In the digestive gland, AHH activity was induced in most individuals from the treatment groups. In all treatment groups, GST activity was significantly inhibited from days 2 to 10 in the gills and was induced after day 4 in the digestive gland. GPx activity in the gills was significantly inhibited throughout the exposure period in all treatment groups. There was no overall significant difference in GPx activity in the digestive gland between the control and treatment groups. Our results also revealed that petroleum hydrocarbon concentrations in the tissues increased linearly with exposure time. EROD activity in the digestive gland and GST and GPx activity in the gill tissue were negatively correlated with petroleum hydrocarbon body burden. These enzymes play important roles in detoxification and can act as potential biomarkers for monitoring petroleum hydrocarbon contaminants in the marine environment.

E2 potentializes benzo(a)pyrene-induced hepatic cytochrome P450 enzyme activities in Nile tilapia at high concentrations.

In the aquatic environment, biotransformation enzymes are established biomarkers for assessing PAH exposure in fish, but little is known about the effect of 17β-estradiol (E2) on these enzymes during exposure to benzo(a)pyrene (BaP). In this study, Nile tilapia (Oreochromis niloticus) were exposed for 3, 5, and 10 days to BaP (300 μg L(-1)) and E2 (5 μg L(-1)). These substances were applied isolated or mixed. In the mixture experiment, fish were analyzed pre- and postexposure in order to better understand whether preexposure to the hormone masks the responses activated by PAH or vice versa. Phase I enzymes ethoxyresorufin-O-deethylase (EROD), pentoxyresorufin-O-depenthylase (PROD), and benzyloxyresorufin-O-debenzylase (BROD) activities as well as the phase II enzyme glutathione S-transferase (GST) were analyzed. Isolated E2 treatment decreased EROD activity after 3 days, but this enzyme activity returned to control values after 5 and 10 days of exposure. Isolated BaP treatment significantly induced EROD activity after 3 and 5 days, and the activity returned to control levels after ten exposure days. Combined treatment (E2 + Bap) significantly increased EROD activity, both in the pre- and postexposure. This increase was even higher than in the isolated BaP treatment, suggesting a synergism between these two compounds. When E2 and BaP were used singly, they did not change BROD and PROD activities. However, combined treatment (E2 + Bap) significantly increased PROD activity. Isolated BaP treatment increased GST activity after 10 days. However, this response was not observed in the mixture treatment, suggesting that E2 suppressed the GST induction modulated by BaP. The results put together indicated that E2 altered the biotransformation pathway regarding enzymes activated by BaP in Nile tilapia.

Assessment of domestic landfill leachate toxicity to the Asian clam Corbicula fluminea via biomarkers

In order to evaluate the effects of domestic landfill leachate to bivalves Corbicula fluminea, clams were exposed to different leachate concentrations (v/v): 2, 3, 6 and 10 percent, corresponding to dilutions observed along a stream that receives this effluent, or only to clean water for comparisons. After 5 and 15 days of exposure the activity of the biotransformation enzymes 7-ethoxyresorufin-O-deethylase (EROD) and glutathione S-transferase (GST), the multixenobiotic resistance mechanism (MXR) and lipid peroxidation (LPO) in gills and digestive gland and metallothionein (MT) content in gills were evaluated. Differences in biomarkers responses were observed between gills and digestive gland, except for MXR that decreased in both tissues of clams exposed to 6 percent for 5 days. EROD activity in gills was reduced in all leachate concentrations after 5 days and only in 2 percent after 15 days exposure, while an EROD increase was observed in digestive gland after 15 days exposure to 6 percent. GST activity increased only in the gills of clams exposed to 10 percent for 5 days. LPO varied between tissues and different conditions. A significant increase in LPO was observed in the gills, after 5 days exposure to 2 and 6 percent, and in digestive gland after 5 and 15 days exposure to 2 and 3 percent. MT content in the gills increased after 15 days exposure to 2 percent. In conclusion, different leachate concentrations tested here caused biochemical changes in C. fluminea, but due to the observed variability in biomarkers responses among leachate concentrations, it was difficult to determine patterns or thresholds concentrations.

Effects of atrazine and chlorpyrifos on cytochrome P450 in common carp liver

Atrazine (ATR) and chlorpyrifos (CPF), widely used in agriculture, have resulted in a series of toxicological and environmental problems. We investigated the activities of the biotransformation enzymes ethoxyresorufin-O-deethylase (EROD) and pentoxyresorufin-O-deethylase (PROD), total cytochrome P450 (CYP), CYP1A mRNA level and level of tissue ATR, CPF, and their metabolites in the liver of common carp (Cyprinus carpio L.) after a 40-d exposure to CPF and ATR, alone or in combination, and a 20-d recovery. In the present study, juvenile common carp was exposed to ATR (at concentrations of 4.28, 42.8 and 428μgL−1), CPF (1.16, 11.6 and 116μgL−1), and ATR/CPF mixture (at concentrations of 1.13, 11.3 and 113μgL−1). A general increasing trend for the activity of the biotransformation enzymes (EROD and PROD), CYP and CYP1A mRNA level was observed in the liver of common carp exposed to ATR, CPF and the ATR/CPF mixture. In addition, ATR, CPF, and their metabolites demonstrated a high accumulation in the liver. These results demonstrated that the CYP system in fish could be used as a biomarkers in evaluating the impact of ATR and CPF exposure on the common carp.

Detection of Multiple Cytochrome P450 in Hepatic Tissue of Heteropneustes fossilis (Bloch) Exposed to Cypermethrin

Cypermethrin (alpha-cyano-3-phenoxybenzyl ester of 2,2-dimethyl-3-(2,2-dichlorovinyl) cyclopropane carboxylic acid) is a synthetic pyrethroid. It is one of the most widely used pesticide in commercial agricultural applications because of its high effectiveness against target species. Beside its target toxicity it is also highly toxic to other non-target species like fish, bees and aquatic insects. The aim of this study was to detect the presence of cytochrome P450 (CYP 450) in the hepatic microsomes of Heteropneustes fossilis upon exposure to cypermethrin. The 96 h LC50 value for each exposure route was calculated and two groups were treated, with one group receiving a single IP (intraperitoneal) injection for 96 h (0.030 mg/kg body weight) and the other group with 1/3 sub-lethal concentration (1.2 μg/l) of the LC50 value in water for 15 days. Activities of the enzymes ethoxyresorufin-o-deethylase (EROD), N,N-dimethylaniline demethylase, aniline hydroxylase and erythromycin demethylase mediated respectively by the isozymes CYP1A, CYP2B, CYP2E1 and CYP3A4 were studied. The liver somatic index (LSI) was also calculated to determine the physiological status of the fish. Activities of CYP1A, CYP2B and CYP2E1 enzymes increased significantly while that of CYP3A4 enzyme inhibited as compared to control. Total CYP 450 content was also significantly induced in both the treated groups. The increase in activities of CYP P450 isozymes could be used as a biomarker to indicate the pollution of an aquatic environment by the pesticide.

Atrazine promotes biochemical changes and DNA damage in a Neotropical fish species

The effects of Atrazine, an herbicide used worldwide and considered as a potential contaminant in aquatic environments, were assessed on the Neotropical fish Prochilodus lineatus acutely (24 and 48h) exposed to 2 or 10μgL−1 of atrazine by using a set of biochemical and genetic biomarkers. The following parameters were measured in the liver: activity of the biotransformation enzymes ethoxyresorufin-O-deethylase (EROD) and glutathione S transferase (GST), antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), content of reduced glutathione (GSH), generation of reactive oxygen species (ROS) and occurrence of lipid peroxidation (LPO); in brain and muscle the activity of acetylcholinesterase (AChE) and DNA damage (comet assay) on erythrocytes, gills and liver cells. A general decreasing trend on the biotransformation and antioxidant enzymes was observed in the liver of P. lineatus exposed to atrazine; except for GR, all the other antioxidant enzymes (SOD, CAT and GPx) and biotransformation enzymes (EROD and GST) showed inhibited activity. Changes in muscle or brain AChE were not detected. DNA damage was observed in the different cell types of fish exposed to the herbicide, and it was probably not from oxidative origin, since no increase in ROS generation and LPO was detected in the liver. These results show that atrazine behaves as enzyme inhibitor, impairing hepatic metabolism, and produces genotoxic damage to different cell types of P. lineatus.

Effects of carbofuran on the sea bass ( Dicentrarchus labrax L.): Study of biomarkers and behaviour alterations

The objective of this study was to investigate the acute effects of the pesticide carbofuran on the sea bass ( Dicentrarchus labrax) using parameters at different levels of biological organisation (swimming behaviour and several biomarkers) and possible relationships between alterations found in different effect criteria. In a bioassay, sea bass juveniles were individually exposed to different doses of carbofuran (31, 63, 125 and 250 μg/L) for 96 h. At the end of the bioassay, the swimming performance and 11 biomarkers were determined. Biomarkers were: hepatosomatic index (HSI), lipid peroxidation (LPO), reduced glutathione and the activities of the enzymes ethoxyresorufin O-deethylase (EROD), glutathione S-transferases, superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, brain acetylcholinesterase (AChE) and muscle cholinesterases (ChE). After 96 h of exposure, carbofuran induced a decrease of the swimming velocity and inhibition of EROD activity at all concentrations tested, and inhibition of muscle ChE and brain AChE activities at 250 μg/L. No relevant alterations in any of the other tested parameters were found. These results show that carbofuran induced adverse effects on fish by interfering with neurofunction, capability of detoxication and swimming velocity. In addition, positive and significant correlations between the swimming velocity and (i) brain AChE activity, (ii) muscle ChE activity and (iii) EROD activity suggest that the inhibition of these enzymes may somehow be related to the behavioural changes observed. Since these functions are determinant for the survival and performance of the fish in the wild, the findings of the present study suggest that adverse effects may occur in populations exposed to carbofuran if a sufficient number of animals is affected.