Increase In EROD Activity Research Articles

Toxicity evaluation of water-accommodated fraction of heavy and light oils on the rainbow trout fish cell line RTL-W1.

Fish are currently used models for the toxicity assessment of chemicals, including polycyclic aromatic hydrocarbons (PAHs). Alternative methods including fish cell lines are currently used to provide fast and reliable results on the toxic properties of chemicals while respecting ethical concerns about animal testing. The Rainbow trout liver cell line RTLW1 was used to analyze the effects of two water-accommodated fractions from two crude oils: Arabian Light crude oil (LO) and refined oil from Erika (HO). Several toxicity endpoints were assessed in this study, including cytotoxicity, EROD activity, DNA damage (comet and micronucleus assays), and ROS production. RTL-W1 cells were exposed for 24h at two or three dilutions of WAF at 1000µg/L (0.1% (1μg/L), 1% (10μg/L), and 10% (100μg/L)) for cytotoxicity and EROD activity and 1% and 10% for ROS production and genotoxicity). Exposure of RTL-W1 cells to LO WAF induced a significant increase of EROD activity and ROS production and altered DNA integrity as revealed by both the comet assay and the micronucleus test for 10µg/L of LO. On the other hand, HO WAF exhibited limited toxic effects except for an EROD induction for 1% WAF dilution. These results confirmed the usefulness of RTL-W1 cells for in vitro toxicological assessment of chemical mixtures.

THE BIOCHEMICAL RESPONSES OF BIVALVE MOLLUSCS TO NEUROLEPTIC CHLORPROMAZINE ARE COMPARABLE WITH THE RESPONSES OF HIGHER VERTEBRATES

Pharmaceuticals such as chlorpromazine (Cpz) are emerging aquatic pollutants with potential effects on non-target organisms. However, its effects on aquatic organisms remain limited and inconclusive. The aim of this study was to compare the responses to Cpz in marine and freshwater bivalve molluscs. Methods. Mytilus galloprovincialis and Unio tumidus were exposed to pM and nM concentrations of Cpz for 14 days and analysed 16 parameters, including cytotoxicity, oxidative/reductive stress responses, metallothionein concentration and biotransformation enzymes in the digestive gland. Results. In both species, Cpz increased EROD activity, possibly leading to the formation of reactive metabolites. Marine mussels exposed to Cpz I and II showed increased GST activity, suggesting detoxification, whereas freshwater mussels showed decreased GST activity, suggesting higher Cpz toxicity. Activation of the GTPase dynamin in U. tumidus exposed to Cpz I confirmed the effect of Cpz on endocytosis. Oxidative stress responses were observed in both species, with changes in oxidative/reductive stress responses, suggesting toxic effects of Cpz. GSH and metallothionein concentrations increased in all exposures, while the NADH/NAD+ ratio increased significantly in U. tumidus. Loss of lysosomal membrane stability was observed in all exposed groups, with CtD efflux detected only in mussels. Differences in caspase-3 activity were also observed between species. Conclusion. Cpz showed adverse effects on vertebrates at μM concentrations, while mussels were affected at pM to nM concentrations, indicating concentration-dependent interspecies effects. Stress responses were similar between species, suggesting that marine molluscs are a potential model for assessing the adverse effects of Cpz on higher vertebrates.

Transient exposure to sublethal concentrations of a pesticide mixture (chlorpyrifos–difenoconazole) caused different responses in fish species from different trophic levels of the same community

The assessment of early effects caused in biota by sublethal exposure to pesticide mixtures should enhance the realism in the ecological risk assessment for agricultural landscapes. This study aimed to evaluate sub-individual responses in fish, which can be linked with outcomes at higher levels of biological organization and affect their trophic relationships. A multilevel biomarker approach was applied to assess the effects of a 48 h exposure of two freshwater mesoamerican fish species (Parachromis dovii and Poecilia gillii) to a mixture of sublethal concentrations of chlorpyrifos (5 μg/L) and difenoconazole (325 μg/L). Transcriptomic induction of cyp1A and the activities of 7-ethoxy-resorufin-O-distillase (EROD) and glutathione S-transferase (GST) were measured as biotransformation-related biomarkers; cholinesterase activity (ChE) was assessed as a neurotoxicity biomarker; resting metabolic rate (RMR) was measured as a physiological biomarker; and the movement of fish in a dark–light environment as a behavior biomarker. The exposure to the mixture had evident effects on P. gillii, with significant induction of cyp1A transcription, increased EROD activity, ChE inhibition in muscle, and increased permanence in the light side of the dark–light environment. Meanwhile, P. dovii only showed significant induction of cyp1A, without evidence of neurotoxicity or changes in behavior. This study demonstrates that the severity of the effects caused by the exposure to a mixture of pesticides can differ among species from the same trophic chain. The potential impairment of predator-prey relationships is a relevant effect that pesticide pollution can cause and it should be considered for the risk assessment of such contaminants.

Uptake and effects of graphene oxide nanomaterials alone and in combination with polycyclic aromatic hydrocarbons in zebrafish

Because of its surface characteristics, once in the aquatic environment, graphene could act as a carrier of pollutants, such as polycyclic aromatic hydrocarbons (PAHs), to aquatic organisms. In this study we aimed to (1) assess the capacity of graphene oxide (GO) to sorb PAHs and (2) to evaluate the toxicity of GO alone and in combination with PAHs on zebrafish embryos and adults. GO showed a high sorption capacity for benzo(a)pyrene (B(a)P) (98% of B(a)P sorbed from a nominal concentration of 100 μg/L) and for other PAHs of the water accommodated fraction (WAF) of a naphthenic North Sea crude oil, depending on their log Kow (95.7% of phenanthrene, 84.4% of fluorene and 51.5% of acenaphthene). In embryos exposed to different GO nanomaterials alone and with PAHs, no significant mortality was recorded for any treatment. Nevertheless, malformation rate increased significantly in embryos exposed to the highest concentrations (5 or 10 mg/L) of GO and reduced GO (rGO) alone and with sorbed B(a)P (GO-B(a)P). On the other hand, adults were exposed for 21 days to 2 mg/L of GO, GO-B(a)P and GO co-exposed with WAF (GO + WAF) and to 100 μg/L B(a)P. Fish exposed to GO presented GO in the intestine lumen and liver vacuolisation. Transcription level of genes related to cell cycle regulation and oxidative stress was not altered, but the slight up-regulation of cyp1a measured in fish exposed to B(a)P for 3 days resulted in a significantly increased EROD activity. Fish exposed to GO-B(a)P and to B(a)P for 3 days and to GO + WAF for 21 days showed significantly higher catalase activity in the gills than control fish. Significantly lower acetylcholinesterase activity, indicating neurotoxic effects, was also observed in all fish treated for 21 days. Results demonstrated the capacity of GO to carry PAHs and to exert sublethal effects in zebrafish.

PAH SORPTION TO NANOPLASTICS AND THE TROJAN HORSE EFFECT AS DRIVERS OF MITOCHONDRIAL TOXICITY AND PAH LOCALIZATION IN ZEBRAFISH.

Plastics are world-wide pollutants that pose a potential threat to wildlife and human health. Small plastic particles, such as microplastics and nanoplastics, are easily ingested, and can act as a Trojan Horse by carrying microorganisms and pollutants. This study investigated the potential role of the Trojan Horse effect in the toxicity of nanoplastics to the vertebrate model organism, zebrafish (Danio rerio). First, we investigated if this effect could affect the toxicity of nanoplastics. Second, we analyzed if it could contribute to the biodistribution of the associated contaminants. And third, we focused on its effect on the mitochondrial toxicity of nanoplastics. We incubated 44 nm polystyrene nanoparticles with a real-world mixture of polycyclic aromatic hydrocarbons (PAHs) for 7 days and removed the free PAHs by ultrafiltration. We dosed embryos with 1 ppm of nanoplastics (NanoPS) or PAH-sorbed nanoplastics (PAH-NanoPS). Neither type of plastic particle caused changes in embryonic and larval development. Fluorescence microscopy and increased EROD activity suggested the uptake of PAHs in larvae exposed to PAH-NanoPS. This coincided with higher concentrations in the yolk sac and the brain. However, PAH-only exposure leads to their accumulation in the yolk sac but not in the brain, suggesting that that the spatial distribution of bioaccumulated PAHs can differ depending on their source of exposure. Both nanoplastic particles affected mitochondrial energy metabolism but caused different adverse effects. While NanoPS decreased NADH production, PAH-NanoPS decreased mitochondrial coupling efficiency and spare respiratory capacity. In summary, the addition of PAHs to the surface of nanoplastics did not translate into increased developmental toxicity. Low levels of PAHs were accumulated in the organisms, and the transfer of PAHs seems to happen in tissues and possibly organelles where nanoplastics accumulate. Disruption of the energy metabolism in the mitochondria may be a key factor in the toxicity of nanoplastics, and the Trojan Horse effect may amplify this effect.

Ecological changes as a plausible explanation for differences in uptake of contaminants between European perch and eelpout in a coastal area of the Baltic Sea

Unexpected increasing trends in the concentration of contaminants in European perch (Perca fluviatilis) and in activity of ethoxyresorufin-O-deethylase (EROD) in European perch and eelpout (Zoarces viviparus) have been observed at a Swedish coastal reference site. This study uses data from different sources to investigate plausible explanations. The results showed that a change in diet and an improved overall condition coincide with an increase in mercury in European perch. Furthermore, an increase in several organic contaminants in European perch coincided with the introduction of an invasive deep-burrowing polychaete, which likely contributed to the release of contaminants through bioturbation. The increase in EROD-activity in both species seems to be related to contaminants that reach the fish through the water rather than the diet. The results show that for contaminants that are taken up via the diet, trends in contamination can be opposite for different species of fish in the same area.

Environmental samples of microplastics induce significant toxic effects in fish larvae

Microplastics (MPs) are present throughout aquatic ecosystems, and can be ingested by a wide variety of organisms. At present, the physical and chemical effects of environmental MPs on aquatic organisms are poorly documented. This study aims to examine the physiological and behavioral effects caused by fish consuming environmental microplastics at different life stages. MP samples were collected from beaches on three islands (Easter Island, Guam and Hawaii) located near the North and South gyres of the Pacific Ocean. Larvae and juveniles of Japanese Medaka were fed for 30days with three doses of MPs (0.01, 0.1 and 1% w/w in fish food) approximate to the concentrations measured in moderately and heavily contaminated ocean areas. Ingestion of MPs by medaka larvae caused (variously) death, decreased head/body ratios, increased EROD activity and DNA breaks and, alterations to swimming behavior. A diet of 0.1% MPs was the most toxic. Two-month-old juveniles fed with 0.01% MPs did not exhibit any symptoms except an increase in DNA breaks. Our results demonstrate ingestion and mainly sublethal effects of environmental MPs in early life stages of fish at realistic MP concentrations. The toxicity of microplastics varies from one sample to another, depending on polymer composition, weathering and pollutant content. This study examines the ecological consequences microplastic build-up in aquatic ecosystems, more particularly in coastal marine areas, which serve as breeding and growing grounds for a number of aquatic species.

Toxicity assessment of pollutants sorbed on environmental microplastics collected on beaches: Part II-adverse effects on Japanese medaka early life stages

While microplastics are present in great abundance across all seas and oceans, little is known about their effects on marine life. In the aquatic environment, they can accumulate a variety of chemicals and can be ingested by many marine organisms including fish, with chronic physical and chemical effects. The purpose of this paper is to evaluate the toxic effects of pollutants sorbed at the surface of environmental microplastics (MPs), collected on various beaches from three islands of the Pacific Ocean. Developmental toxicity of virgin MPs or artificially coated with B[a]P and environmental MPs from Easter Island, Guam and Hawaii was evaluated on embryos and prolarvae of Japanese medaka. Mortality, hatching success, biometry, malformations, EROD activity and DNA damage were analyzed after exposure to DMSO extracts. No toxicity was observed for extracts of virgin MPs whatever the endpoint considered. Extracts of virgin MPs coated with 250 µg.g−1 of B(a)P induced lethal effects with high embryo mortality (+81%) and low hatching rate (−28%) and sublethal effects including biometry and swimming behavior changes, increase of EROD activity (+94%) and DNA damage (+60%). Environmental MPs collected on the three selected islands exhibited different polymer, pollutant and toxicity patterns. The highest toxicity was detected for MPs extract from Hawaï with head/body length and swimming speed decreases and induction of EROD activity and DNA stand breaks. This study reports the possible sublethal toxicity of organic pollutants sorbed on MPs to fish early life stages.

Toxicity of erythromycin to Oncorhynchus mykiss at different biochemical levels: detoxification metabolism, energetic balance, and neurological impairment.

During the last decades, the presence of antibiotics in different aquatic compartments has raised increasing interest and concern, since these compounds are usually persistent and bioactive pseudo pollutants. Erythromycin (ERY) is a macrolide antibiotic, prescribed for human and veterinary medicines but also used in aquaculture and livestock production. Taking into account the recorded environmental levels of ERY, its toxicity to non-target organisms has become a still poorly studied issue, particularly in fish. In this sense, this study investigated the acute and chronic effects of realistic levels of ERY on Oncorhynchus mykiss (rainbow trout), namely, through the quantification of the activity of enzymes involved in different biochemical pathways, such as detoxification (phase I-7-ethoxyresorufin O-deethylase (EROD); phase II-glutathione S-transferases (GSTs), uridine-diphosphate-glucuronosyltransferases (UGTs)), neurotransmission (acetylcholinesterase (AChE)), and energy production (lactate dehydrogenase (LDH)). Both types of exposure caused significant increases in EROD activity in liver of O. mykiss; an increase in GST activity in gills after chronic exposure was also observed. UGT branchial activity was significantly depressed, following the long-term exposure. Thus, EROD, GST, and UGT enzymatic forms seem to be involved in the biotransformation of ERY. In terms of neurotransmission and preferential pathway of energy homeostasis, the exposed organisms appear not to have been affected, as there were no significant alterations in terms of AChE and LDH activities, respectively. The here-obtained data suggest that the observed alterations in terms of detoxification enzymes may have prevented the establishment of a set of toxic responses, namely, neurotoxic and metabolic disorders.

Toxicological and chemical investigation of untreated municipal wastewater: Fraction- and species-specific toxicity

Absence of a municipal wastewater (WW) treatment plant results in the untreated WW discharge into the recipient. The present study investigated toxic effects and chemical composition of water extracts and fractions from untreated WW and recipient Danube River (DR). Samples were prepared by solid-phase extraction and silica gel fractionation and screened for EROD activity and cytotoxicity using aquatic models, comprising of fish liver cells (PLHC-1) and a model of the early development of zebrafish embryos, while rat (H4IIE) and human (HepG2) hepatoma cells served as mammalian models. Polar fraction caused cytotoxicity and increased the EROD activity in PLHC-1 cells, and increased mortality and developmental abnormalities in developing zebrafish embryos. In H4IIE, polar fraction induced inhibition of cell growth and increased EROD activity, whereas HepG2 exerted low or no response to the exposure. Non-polar and medium-polar fractions were ineffective. Tentative identification by GC/MS showed that WW is characterized by the hydrocarbons, alkylphenols, plasticizers, and a certain number of benzene derivatives and organic acids. In DR, smaller number of organic compounds was identified and toxicity was less pronounced than in WW treatments. The present study revealed the potent toxic effect of polar fraction of untreated WW, with biological responses varying in sensitivity across organisms. Obtained results confirmed that fraction- and species-specific toxicity should be considered when assessing health risk of environmental pollution.

Biochemical and genotoxic effects of a commercial formulation of the herbicide tebuthiuron in Oreochromis niloticus of different sizes

Pesticides are serious contaminants because they are designed to eliminate pests, but they also affect non-target species. The present study aimed to evaluate the biochemical and genetic effects of the herbicide tebuthiuron in Oreochromis niloticus of different sizes. Thus, we analyzed biomarkers in small and large O. niloticus specimens exposed to 62.5, 125 and 250 mg L-1 of tebuthiuron for 72 hours. Fish exposed to 250 mg L-1 had high mortality rates; therefore, the data could not be used. The results showed an increase in EROD activity in fish exposed to 125 mg L-1, but no GST alteration. Antioxidant enzymes GPx and CAT were altered only in the liver of treated fish compared to the control group: CAT decreased in large fish, and GPx increased in small fish. The MDA analysis did not evidence lipid peroxidation. High DNA damage in exposed small fish (not in large fish) was observed using comet assay, but a micronucleus test did not show mutagenicity. Moreover, a comparison between control groups with specimens of different sizes revealed that small fish are more susceptible than large fish to the tebuthiuron effects, since increased comet scores was observed only for smaller fish.

A novel MDCKII in vitro model for assessing ABCG2-drug interactions and regulation of ABCG2 transport activity in the caprine mammary gland by environmental pollutants and pesticides

The ABC efflux transporter ABCG2 represents the main route for active secretion of xenobiotics into milk. Thus, ABCG2 regulation by aryl hydrocarbon receptor (AhR) ligands including ubiquitously environmental pollutants is of great toxicological relevance. However, no adequate in vitro model is as yet available to study AhR-dependent ABCG2 regulation in dairy animals. In this study, we therefore systematically investigated the effect of various environmental contaminants and pesticides on ABCG2 efflux activity in MDCKII cells stably expressing mammary ABCG2 from dairy goats. The AhR-agonists TCDD, Aroclor 1254, prochloraz, and iprodione caused a dose- and time-dependent increase in EROD activity. Moreover, TCDD and prochloraz significantly stimulated ABCG2 transport activity through a dose- and time-dependent induction of transporter gene expression. AhR inhibitors like CH223191 significantly reversed TCDD- and prochloraz-induced stimulation of ABCG2 efflux activity. In contrast, non-AhR activators such as PCB 101 had no significant effect on EROD activity, ABCG2 gene expression or transporter activity. As we identified various anthelmintics including monepantel as potential ABCG2 substrates this regulatory mechanism may result in increased milk residues of potentially harmful xenobiotics. Thus, MDCKII-cABCG2 cells may represent a suitable in vitro model to study mammary ABCG2 secretory activity and its potential regulation by AhR-activating contaminants.

Xenobiotic metabolism induction and bulky DNA adducts generated by particulate matter pollution in BEAS-2B cell line: geographical and seasonal influence

Airborne particulate matter (PM) toxicity is of growing interest as diesel exhaust particles have been classified as carcinogenic to humans. However, PM is a mixture of chemicals, and respective contribution of organic and inorganic fractions to PM toxicity remains unclear. Thus, we analysed the link between chemical composition of PM samples and bulky DNA adduct formation supported by CYP1A1 and 1B1 genes induction and catalytic activities. We used six native PM samples, collected in industrial, rural or urban areas, either during the summer or winter, and carried out our experiments on the human bronchial epithelial cell line BEAS-2B. Cell exposure to PM resulted in CYP1A1 and CYP1B1 genes induction. This was followed by an increase in EROD activity, leading to bulky DNA adduct formation in exposed cells. Bulky DNA adduct intensity was associated to global EROD activity, but this activity was poorly correlated with CYPs mRNA levels. However, EROD activity was correlated with both metal and polycyclic aromatic hydrocarbon (PAH) content. Finally, principal components analysis revealed three clusters for PM chemicals, and suggested synergistic effects of metals and PAHs on bulky DNA adduct levels. This study showed the ability of PM samples from various origins to generate bulky DNA adducts in BEAS-2B cells. This formation was promoted by increased expression and activity of CYPs involved in PAHs activation into reactive metabolites. However, our data highlight that bulky DNA adduct formation is only partly explained by PM content in PAHs, and suggest that inorganic compounds, such as iron, may promote bulky DNA adduct formation by supporting CYP activity.

Environmental hazard of yperite released at sea: sublethal toxic effects on fish

The aim of this study was to evaluate the potential toxicological effects on fish related to the leakage of yperite from rusted bomb shells dumped at sea. Both in vivo and field studies have been performed. As for the in vivo experiment, specimen of European eel were subcutaneously injected with 0.015, 0.15 and 1.5mg/kg of yperite and sacrificed after 24 and 48h. In the field study, specimen of Conger eel were collected from a dumping site in the Southern Adriatic Sea. The presence/absence of yperite in tissues, genotoxicity, detoxification enzymes, histological alterations and gross abnormalities were investigated. Results of the in vivo experiment showed a significant increase of EROD activity at both 24h and 48h. UGT activity increased significantly at 48h post injection. An acute inflammatory response after 24h in skin layers and muscle was observed, associated to cell degeneration and necrosis after 48h at the highest dose. On field, comet assay revealed genotoxicity in gills of fish from the dumping site. Specimen from the dumping site showed significantly higher EROD activities compared to controls, deep ulcers and papules on skin together with liver and spleen histopathological lesions.

Biomarker responses in persian sturgeon (Acipenser persicus) exposed to benzo-a-pyrene and beta-naphthoflavone

Biotransformation enzymes of xenobiotics (ethoxyresorufin-O-deethylase, cytochrome P4501A1 content and glutathione-S-transferase) were investigated in the liver of Persian Sturgeon (Acipenser persicus) after a 96-hour exposure to polycyclic aromatic hydrocarbons (PAHs), premutagenic benzo[a]pyrene (BaP) and beta-naphthoflavone (BNF). The fish were injected 10 mg/kg wet-body weight in corn oil for 96 hours every days. Ethoxyresorufin-O-deethylase activity (EROD) and glutathione s-transferase activity (GST) were measured in the fish liver. Cytochrome P4501A1 (CYP1A1) content was estimated by indirect enzyme-linked immunosorbent assay (ELISA). The response appeared as early as 12 hours post exposure. A time-dependent response was observed in the EROD activity, being significantly higher at 48 hours post exposure to 10 mg/kg of BaP. The greatest induction occurred in the fish treated with 10 mg/kg BaP, in which a 32.1- fold increase in EROD activity was observed. Results showed that EROD activity in A. persicus is significantly increased by BaP and BNF treatments. Both chemicals showed higher values of EROD activity compared to the liver CYP1A content. There was a rise in glutathione-S-transferase activity in fish exposed to BNF, but no increase was observed in fish treated with BaP. The results showed that hepatic CYP1A expression in terms of induction of EROD activity might be suited as a biomarker of organic contamination in aquatic environments and led to lower sensitivity of the second phase in the detoxification enzyme.

The Synergistic Effects of Insecticidal Essential Oils and Piperonyl Butoxide on Biotransformational Enzyme Activities inAedes aegypti(Diptera: Culicidae)

The biochemical mechanisms underlying the increased toxicity of several plant essential oils (thymol, eugenol, pulegone, terpineol, and citronellal) against fourth instar of Aedes aegypti L. when exposed simultaneously with piperonyl butoxide (PBO) were examined. Whole body biotransformational enzyme activities including cytochrome P450-mediated oxidation (ethoxyresorufin O-dethylase [EROD]), glutathione S-transferase (GST), and beta-esterase activity were measured in control, essential oil-exposed only (single chemical), and essential oil + PBO (10 mg/liter) exposed larvae. At high concentrations, thymol, eugenol, pulegone, and citronellal alone reduced EROD activity by 5-25% 16 h postexposure. Terpineol at 10 mg/liter increased EROD activity by 5 +/- 1.8% over controls. The essential oils alone reduced GST activity by 3-20% but PBO exposure alone did not significantly affect the activity of any of the measured enzymes. All essential oils in combination with PBO reduced EROD activity by 58-76% and reduced GST activity by 3-85% at 16 h postexposure. This study indicates a synergistic interaction between essential oils and PBO in inhibiting the cytochrome P450 and GST detoxification enzymes in Ae. aegypti.

Multi-biomarker approach in wild European bullhead, Cottus sp., exposed to agricultural and urban environmental pressures: Practical recommendations for experimental design

In freshwater ecosystems, a large number of chemical substances are able to disturb homeostasis of fish by modulating one or more physiological functions including the immune system. The aim of this study was to assess multi-biomarker responses including immunotoxicity induced by urban and agricultural pressure in European bullheads living in a small French river basin. For this purpose, a set of biochemical, immunological, physiological and histological parameters was measured in wild bullheads from five locations characterized by various environmental pressures. Moreover, to address effects of physiological status and contamination level variation on biomarker responses, fish were sampled during three periods (April, July and October). Results revealed a clear impact of environmental pressure on fish health and particularly on immunological status. An increase of EROD activity was recorded between upstream and downstream sites. Upstream sites were also characterized by neurotoxicological effects. Fish exhibited upstream/downstream variations of immunological status but strong differences were observed according to sampling season. Conversely, regarding biochemical and immunological effects, no significant response of physiological indexes was recorded related to environmental pressures. According to these results, the European bullhead appears as a valuable model fish species to assess adverse effects in wildlife due to urban and agricultural pressures.

Alteration of the pharmacokinetics of theophylline by rutaecarpine, an alkaloid of the medicinal herb Evodia rutaecarpa, in rats

Rutaecarpine is a main active alkaloid present in the medicinal herb, Evodia rutaecarpa. The cytochrome P450 (CYP) 1A2 substrate, theophylline, is an important therapeutic agent for the treatment of asthma, but has a narrow therapeutic index. To evaluate the pharmacokinetic interaction of theophylline with rutaecarpine, the effects of rutaecarpine on CYP1A2 activity and theophylline pharmacokinetics were investigated. Oral treatment of Sprague-Dawley rats with 50 mg kg(-1) rutaecarpine for three days through a gastrogavage caused a 4- and 3-fold increase in liver microsomal 7-ethoxyresorufin O-deethylation (EROD) and 7-methoxyresorufin O-demethylation activity, respectively. In the kidney, rutaecarpine treatment caused a 3-fold increase in EROD activity. In the lungs, EROD activity was elevated from an undetectable to a detectable level by rutaecarpine. Pharmacokinetic parameters of theophylline were determined using a microdialysis sampling method. Rutaecarpine pre-treatment increased the clearance of theophylline in a dose-dependent manner. Pre-treatment of rats with 50 mg kg(-1) rutaecarpine caused a 3-fold increase in theophylline clearance and a 70%, 68% and 68% decrease in the area under the concentration-time curve (AUC), mean residence time (MRT) and half-life, respectively. These results demonstrated that rutaecarpine treatment elevated CYP1A2 catalytic activity and theophylline excretion in rats. In patients taking theophylline, adverse effects might be noticed when a rutaecarpine-containing herbal preparation is used concomitantly.

Differential inductive profiles of hepatic cytochrome P450s by the extracts of Sophora flavescens in male and female C57BL/6JNarl mice

Aim of the study Sophora flavescens has been used as an antipyretic and analgesic agent. To assess the possible herb–drug interaction, effects of S. flavescens extracts on hepatic cytochrome P450 (P450, CYP) enzymes were studied. Materials and methods Effects of the extracts prepared by three different pharmaceutical companies on P450 enzymes were investigated in male and female C57BL/6JNarl mice. Results In male mice, extract 1 caused a dose- and time-dependent increase of 7-ethoxyresorufin O-deethylation (EROD) activity. Three-day treatment with 3 g/kg extracts 1–3 elevated EROD, 7-pentoxyresorufin O-dealkylation (PROD), coumarin hydroxylation, and nifedipine oxidation (NFO) activities. In female mice, extracts 1 and 2 increased EROD and PROD activities without affecting coumarin hydroxylation and NFO activities. However, extract 3, which lacked prenylated flavonoids, caused an induction profile in females the same as in males. Treatment with extract 3 fortified with prenylated flavonoids restored the gender difference. An alkaloid, oxymatrine was present in all extracts and increased EROD and PROD activities. At a human equivalent dose (0.18 g/(kg day)), all extracts increased EROD activity. Conclusions These results revealed that Cyp1a had a lower induction response threshold. Oxymatrine contributed at least partly to the P450 induction by S. flavescens. At a higher dose, Cyp2a, Cyp2b, and Cyp3a could be induced and the male-specific induction of Cyp2a and Cyp3a was associated with the presence of prenylated flavonoids.

Identification of mitochondrial cytochrome P450 induced in response to polycyclic aromatic hydrocarbons in the mummichog ( Fundulus heteroclitus)

Increasing evidence suggests that polycyclic aromatic hydrocarbons (PAHs) such as benzo[ a]pyrene (BaP) are localized to the mitochondria. Because the toxic effects of many PAHs are the result of metabolism by cytochrome P4501A (CYP1A), it is important to investigate whether active forms of these enzymes can be identified in the mitochondria. In this study, we identified mitochondrial P450s with a monoclonal antibody against scup ( Stenotomus chrysops) CYP1A in the isolated mitochondrial fraction of the liver from adult male mummichog ( Fundulus heteroclitus) livers. The size of the protein in the mitochondria was similar to that of microsomal CYP1A. Fish dosed with 10 mg/kg BaP had increased EROD activity in the mitochondrial fraction compared to controls. In mummichog larvae dosed with 100 µg/L BaP and 100 µg/L benzo[ k]fluoranthene, CYP1A protein levels as well as enzyme activity were elevated. However, fish from a PAH-polluted Superfund site (Elizabeth River, Portsmouth VA) showed recalcitrant mitochondrial CYP1A protein levels and enzyme activity in a similar manner to microsomal CYP1A.