Organochlorine pesticide acetofenate and its hydrolytic metabolite in rabbits: Enantioselective metabolism and cytotoxicity.

Acetofenate (AF) is a chiral organochlorine pesticide used for controlling hygiene pests. In this study, the metabolism of AF in rabbits in vivo and in vitro was investigated and the primary chiral metabolite acetofenate-alcohol (AF-A) was analyzed. The cytotoxicity of AF and AF-A was also determined. AF in rabbits in vivo was eliminated so rapidly that AF could not be detected within 10min after intravenous administration at 20mg/kg (body weight), and AF-A was quickly formed. In vitro metabolism assay, using plasma and liver microsomes, showed that AF was also quickly metabolized to AF-A and the metabolic process was significantly enantioselective with preferential degradation of (-)-AF and formation of (-)-AF-A. The cytotoxicity of AF and AF-A were investigated by assessing cell proliferation, apoptosis and generation of reactive oxygen species. The results showed that AF and AF-A induce enantioselective cytotoxicity. This study will be helpful for improving knowledge about the metabolism and toxicity of AF on an enantiomeric level and providing evidence to understand the potential environmental risk.

A fugacity-based toxicokinetic model for narcotic organic chemicals in fish.

A novel dynamic fugacity-based model is described, developed, and tested that simulates the uptake of narcotic organic chemicals in fish from water as occurs in aquatic bioconcentration and toxicity tests. The physiologically based toxicokinetic model treats the time course of chemical distribution in 4 compartments (tissue groups) in the fish, including the liver, in which biotransformation may occur. In addition to calculating bioconcentration and toxicokinetics, 5 possible toxic endpoints are defined corresponding to chemical concentration, fugacity, or activity reaching a critical value that causes 50% mortality. The mathematical description of multicompartment uptake is simplified by expressing the equations in the fugacity format. The model is parameterized and tested against reported empirical data for the bioconcentration of pentachloroethane in rainbow trout and for uptake and mortality from aquatic exposures to naphthalene and 1,2,4-trichlorobenzene in fathead minnows. Model performance is evaluated, and it is concluded that with suitable parameterization it has potential for application for assessment of both bioconcentration and toxicity expressed as median lethal concentrations, critical body residues, and chemical activity as a function of time to death.

An indirect screen for brain uptake of 1,2-diarylethane melanocortin 4 receptor antagonists in rats.

Antagonism of the melanocortin 4 receptor (MC4R) has been proposed as a therapeutic intervention for the prevention of lean body mass waste, as in cachexia. Pharmacokinetic profiles of substituted 1,2-diarylethane MC4R antagonists were determined in rats after a single intravenous (IV) administration at 1 mg/kg. Brain and plasma concentrations of these compounds were determined at 1 and 4 hours after an oral dose at 10 mg/kg, since oral administration is the intended clinical dosing route and the pharmacological target is the central nervous system. The brain to plasma concentration ratios (0.10 - 50) after oral dosing correlated well with Vd(ss) (2.21 to 81.4 L/kg; R(2)=0.810) determined after IV administration. A good correlation was also observed between the brain AUC(0-4 hr) (119 - 18400 nM*hr) and Vd(ss) (R(2)=0.981). Thus, further screening and ranking of substituted 1,2-diarylethanes for their brain uptakes could be carried out more efficiently via the simple and indirect Vd(ss) screen after intravenous administration in rats.

Branchial elimination of superhydrophobic organic compounds by rainbow trout (Oncorhynchus mykiss).

The branchial elimination of pentachloroethane and four congeneric polychlorinated biphenyls by rainbow trout was measured using a fish respirometer-metabolism chamber and an adsorption resin column. Branchial elimination was characterized by calculating a set of apparent in vivo blood:water partition coefficients (P(BW)). Linear regression was performed on the logarithms of P(BW) estimates and the log K(OW) value for each compound to give the fitted equation: log P(BW)=0.76 x log K(OW)-1.0 (r(2)=0.98). The linear nature of this relationship provides support for existing models of chemical flux at fish gills and suggests that a near equilibrium condition was established between chemical in venous blood entering the gills, including dissolved and bound forms, and dissolved chemical in expired branchial water. In vivo P(BW) estimates were combined with P(BW) values determined in vitro for a set of lower log K(OW) compounds (Bertelson et al., Environ. Toxicol. Chem. 17 (1998) 1447-1455) to give the fitted relationship: log P(BW)=0.73 x log K(OW)-0.88 (r(2)=0.98). The slope of this equation is consistent with the suggestion that chemical binding to non-lipid organic material contributes substantially to blood:water chemical partitioning. An equation based on the composition of trout blood (water content and the total amount of organic material) was then derived to predict blood:water partitioning for compounds with log K(OW) values ranging from 0 to 8: log P(BW)=log[(10(0.73 log K(ow)) x 0.16)+0.84].

Dermal absorption of three waterborne chloroethanes in rainbow trout (Oncorhynchus mykiss) and channel catfish (Ictalurus punctatus).

In vivo estimates of xenobiotic chemical flux across the dermal surface of intact fish were obtained by measuring chemical loss from venous blood to expired water. An experimental system was developed to separate the dermal route of exposure from all other routes. The system was then used to measure dermal absorption of tetrachloroethane (TCE), pentachloroethane (PCE), and hexachloroethane (HCE) in channel catfish (Ictalurus punctatus) and rainbow trout (Oncorhynchus mykiss), two fish with very different skin anatomies. The kinetics of accumulation varied among chemicals, but for each compound were similar among species. TCE accumulated rapidly, reaching steady state in blood within 48 hr. Steady state was not reached in 48 hr with PCE or HCE, although blood levels of PCE were probably close to steady-state values. Dermal flux estimates (based on branchial efflux) for TCE, PCE, and HCE were two to four times greater in catfish than in trout. Arterial blood concentrations of each compound were three to six times greater in catfish. These observations are indicative of greater flux across catfish skin, augmented by higher blood:water chemical partitioning. Trout skin is covered with scales and has no taste buds, while catfish skin does not possess scales and has numerous taste bud papillae. Both scales and taste bud papillae originate in the dermis and extend to the skin surface through the epidermis. In catfish these taste buds may offer channels through which chemicals diffuse across the epidermis to the more vascularized dermis. A comparison of dermal and branchial uptake was made by estimating zero-time dermal and branchial fluxes for all three chloroethanes. The mean dermal fluxes for TCE, PCE, and HCE ranged from 1.4 to 2.8, 1.8 to 3.6, and 1.4 to 3.2% of the total flux (branchial plus dermal) in rainbow trout and channel catfish, respectively. This research demonstrates that dermal absorption of waterborne chemicals occurs in large adult fish and results in distribution kinetics similar to those observed in inhalation exposures. Compared to branchial uptake, the dermal route of exposure appears to be relatively unimportant in large fish. It may, however, be very important in smaller fish and for juveniles of larger species.

A physiologically based toxicokinetic model for dermal absorption of organic chemicals by fish.

A physiologically based toxicokinetic model was developed to describe dermal absorption of waterborne organic chemicals by fish. The skin was modeled as a discrete compartment into which compounds diffuse as a function of chemical permeability and the concentration gradient. The model includes a countercurrent description of chemical flux at fish gills and was used to simulate dermal-only exposures, during which the gills act as a route of elimination. The model was evaluated by exposing adult rainbow trout and channel catfish to hexachloroethane (HCE), pentachloroethane (PCE), and 1,1,2,2-tetrachloroethane (TCE). Skin permeability coefficients were obtained by fitting model simulations to measured arterial blood data. Permeability coefficients increased with the number of chlorine substituent groups, but not in the manner expected from a directly proportional relationship between dermal permeability and skin:water chemical partitioning. An evaluation of rate limitations on dermal flux in both trout and catfish suggested that chemical absorption was limited more by diffusion across the skin than by blood flow to the skin. Modeling results from a hypothetical combined dermal and branchial exposure indicate that dermal uptake could contribute from 1.6% (TCE) to 3.5% (HCE) of initial uptake in trout. Dermal uptake rates in catfish are even higher than those in trout and could contribute from 7.1% (TCE) to 8.3% (PCE) of initial uptake in a combined exposure.

Three-dimensional visualization of physiologically based kinetic model outputs.

Outputs from a physiologically based toxicokinetic (PB-TK) model for fish were visualized by mapping time-series data for specific tissues onto a three-dimensional representation of a rainbow trout. The trout representation was generated in stepwise fashion: 1) cross-section images were obtained from an anesthetized fish using a magnetic resonance imaging system, 2) images were processed to classify tissue types and eliminate unnecessary detail. 3) processed images were imported to a visualization software package (Application Visualization System) to create a three-dimensional representation of the fish, encapsulating five volumes corresponding to the liver, kidney, muscle, gastrointestinal tract, and fat, Kinetic data for the disposition of pentachloroethane in trout were generated using a PB-TK model. Model outputs were mapped onto corresponding tissues volumes, representing chemical concentration as color intensity. The workstation software was then used to animate the images, illustration the accumulation of pentachloroethane in each tissue during a continuous branchial (gill) exposure.

Role of P4502E1 in the metabolism of 1,1,2,2-tetrafluoro-1-(2,2,2-trifluoroethoxy)-ethane.

1. The metabolism of 1,1,2,2-tetrafluoro-1-(2,2,2-trifluoroethoxy)-ethane (HFE), a prospective chlorofluorocarbon alternative, was studied in rat and human liver microsomes and in rat in vivo. 2. HFE was metabolized to inorganic fluoride, trifluoroacetaldehyde hydrate, trifluoroacetic acid and difluoroacetic acid, which were identified by 19F-nmr in microsomal incubation. After i.p. dosing with 200 mg/kg HFE to rat, trifluoroacetic acid and trifluoroacetaldehyde hydrate were identified as urinary metabolites. 3. The formation of inorganic fluoride from HFE was used to quantify oxidative metabolism. In liver microsomes from untreated rat, formation of inorganic fluoride could not be detected. However, microsomes from rats treated with P4502E1 (2E1) inducers ethanol and pyridine catalysed the formation of fluoride at different rates. The extent of fluoride formation in microsomes correlated with the amount of 2E1 protein as determined by immunoblots with a polyclonal antibody and with the extent of oxidation of p-nitrophenol and chlorzoxazone, two specific substrates for 2E1. 4. In different samples of human liver microsomes, the formation of inorganic fluoride correlated well with the ability of the microsomes to oxidize chlorzoxazone and p-nitrophenol and the amount of 2E1 protein as determined by immunoblots. 5. The obtained results suggest that 2E1 plays a major role in the metabolism of HFE in rat and man.

Complete biological reductive transformation of tetrachloroethene to ethane.

Reductive dechlorination of tetrachloroethene (perchloroethylene; PCE) was observed at 20 degrees C in a fixed-bed column, filled with a mixture (3:1) of anaerobic sediment from the Rhine river and anaerobic granular sludge. In the presence of lactate (1 mM) as an electron donor, 9 microM PCE was dechlorinated to ethene. Ethene was further reduced to ethane. Mass balances demonstrated an almost complete conversion (95 to 98%), with no chlorinated compounds remaining (less than 0.5 micrograms/liter). When the temperature was lowered to 10 degrees C, an adaptation of 2 weeks was necessary to obtain the same performance as at 20 degrees C. Dechlorination by column material to ethene, followed by a slow ethane production, could also be achieved in batch cultures. Ethane was not formed in the presence of bromoethanesulfonic acid, an inhibitor of methanogenesis. The high dechlorination rate (3.7 mumol.l-1.h-1), even at low temperatures and considerable PCE concentrations, together with the absence of chlorinated end products, makes reductive dechlorination an attractive method for removal of PCE in bioremediation processes.

Physiologically based toxicokinetic modeling of three waterborne chloroethanes in rainbow trout (Oncorhynchus mykiss).

A physiologically based toxicokinetic model for fish was used to simulate the uptake and disposition of three waterborne chloroethanes in rainbow trout (Oncorhynchus mykiss). Trout were exposed to 1,1,2,2-tetrachloroethane, pentachloroethane, and hexachloroethane in fish respirometer-metabolism chambers to assess the kinetics of chemical accumulation in arterial blood and chemical extraction efficiency from inspired water. Chemical residues in tissues were measured at the end of each experiment. Trout exposed to tetrachloroethane were close to steady-state in 48 hr. Fish exposed to pentachloroethane were near steady-state in 264 hr. Extraction efficiency data showed that systemic (extrabranchial) elimination of both chemicals was small. Hexachloroethane continued to accumulate in fish exposed for 600 hr. Parameterized with chemical partitioning data obtained in vitro, the model accurately simulated the uptake of all three chloroethanes in blood and tissues and their extraction from inspired water. These results provide support for the basic model structure and the accuracy of physiological input parameters.

A physiologically based toxicokinetic model for the uptake and disposition of waterborne organic chemicals in fish.

A physiologically based toxicokinetic model was developed to predict the uptake and disposition of waterborne organic chemicals in fish. The model consists of a set of mass-balance differential equations which describe the time course of chemical concentration within each of five tissue compartments: liver, kidney, fat, and richly perfused and poorly perfused tissue. Model compartmentalization and blood perfusion relationships were designed to reflect the physiology of fishes. Chemical uptake and elimination at the gills were modeled as countercurrent exchange processes, limited by the chemical capacity of blood and water flows. The model was evaluated by exposing rainbow trout (Oncorhynchus mykiss) to pentachloroethane (PCE) in water in fish respirometer-metabolism chambers. Exposure to 1500, 150, or 15 micrograms PCE/liter for 48 hr resulted in corresponding changes in the magnitude of blood concentrations without any change in uptake kinetics. The extraction efficiency for the chemical from water decreased throughout each exposure, declining from 65 to 20% in 48 hr. Extraction efficiency was close to 0% in fish exposed to PCE to near steady state (264 hr), suggesting that very little PCE was eliminated by metabolism or other extrabranchial routes. Parameterized for trout with physiological information from the literature and chemical partitioning estimates obtained in vitro, the model accurately predicted the accumulation of PCE in blood and tissues, and its extraction from inspired water. These results demonstrate the potential utility of this model for use in aquatic toxicology and environmental risk assessment.

Modeling the tissue solubilities and metabolic rate constant (Vmax) of halogenated methanes, ethanes, and ethylenes.

Experimental solvent:air and tissue:air partition coefficients for 25 halogenated methanes, ethanes, and ethylenes in saline solution; olive oil; and rat blood, muscle, liver, and fat tissues have been examined using theoretical molecular modeling techniques. The metabolic rate constant, Vmax, was also investigated by these techniques for 19 chlorinated compounds in this group. Two graph theoretical approaches (the distance method of Wiener and the connectivity index method of Randic, Kier, and Hall) and an approach utilizing ad hoc molecular descriptors were employed. Satisfactory regression models for solubility were obtained with both the Randic-Kier-Hall approach and the ad hoc descriptors approach. Fluorine substituents decrease tissue solubilities, whereas both clorine and bromine substituents increase tissue solubilities, with the relative influence being Cl less than Br. Tissue solubilities can also be represented conveniently in terms of contributions from oil and saline solubilities, a procedure reinforced by factor analysis of the data. Equations derived by these methods adequately estimated the solubilities for eight additional compounds. No approach could successfully model Vmax for all 19 compounds, but a subset of 16 compounds was modeled using the connectivity indices. The equation is limited in its use but indicated future modeling directions for Vmax.

Metabolism of ethane and pentane to carbon dioxide by the rat.

The pulmonary excretion rates of ethane and pentane have been used as indices of lipid peroxidation. This use assumes that exhalation of these hydrocarbons is directly related to their formation rate. This is true only if the elimination (metabolism plus nonpulmonary excretion) of ethane and pentane are constant in the presence of alterations in lipid peroxidation. However, the in vivo metabolic elimination profile for pentane is unknown and it has not been established with certainty that ethane is metabolized in vivo. Radiolabeled [14C]ethane and pentane were used to study the disposition of these hydrocarbons when injected into an enclosed chamber system containing a rat. The ethane and pentane concentrations in chamber air measured as 14C radioactivity were in general agreement with more selective measurements based on GC analysis. Pentane was cleared from chamber air at a much faster rate than ethane. Approximately 50 and 19.8% of the total radioactivity added to the chamber as [14C]pentane or [14c]ethane, respectively, was recovered as carbon dioxide at the end of 8 hr. The fraction of total radioactivity recovered in urine was 7.6 and 1.0% for the pentane and ethane experiments, respectively. These results indicate unequivocally that both ethane and pentane are metabolized in the intact rat.

The covalent binding of 1,1,2,2-tetrachloroethane to macromolecules of rat and mouse organs.

The in vivo interaction of the hepatocarcinogen 1,1,2,2-tetrachloroethane (1,1,2,2-TTCE) with DNA, RNA, and proteins of male Wistar rats and BALB/c mice was measured 22 hr after i.p. injection. Covalent binding index (CBI) to liver DNA was about 500 and was comparable to those of carcinogens classified as moderate initiators. It was higher than those of other chloroethanes, even than that of 1,2-dichloroethane (1,2-DCE), a symmetrically substituted haloethane whose genotoxicity has been widely demonstrated. In in vitro cell-free systems, 1,1,2,2-tetrachloroethane was bioactivated by mixed-function oxidase(s) and glutathione-S-transferase(s) (GSH-T) from microsomal and cytosolic fractions of rat and mouse liver and, to a lesser extent, of mouse lung. The in vitro activation led to formation of reactive species capable of binding to exogenous DNA and to the subcellular constituents of enzymatic fractions. These data, along with previous literature reports, provide sufficient evidence of 1,1,2,2-TTCE genotoxicity.