Lowest Observed Effect Level Research Articles

A 90-day toxicological evaluation of Compound 1080 (sodium monofluoroacetate) in Sprague-Dawley rats.

Groups of Sprague-Dawley rats received sodium monofluoroacetate (Compound 1080) at 0.025, 0.075, and 0.25 mg/kg by oral gavage once daily for 90 days and were then euthanized. The control and 0.25 mg/kg/day groups included additional rats of each sex that were treated for 90 days, then maintained without treatment for a further 56-day recovery period. Microscopic changes were restricted to the testes and the heart, and were seen only in males dosed with 1080 at 0.25 mg/kg/day and included severe hypospermia in the epididymides, severe degeneration of the seminiferous tubules of the testes, and cardiomyopathy. Sperm evaluation indicated severe decreases in all three sperm parameters evaluated (concentration, % motile, and % abnormal) at 0.25 mg/kg/day. There were no microscopic changes or 1080-related effects on sperm parameters at 0.025 and 0.075 mg/kg/day. The no observable effects level (NOEL) for rats administered 1080 via oral gavage for 90 days was 0.075 mg/kg/day. The lowest observable effects level (LOEL) dose was 0.25 mg/kg/day. After dosing with the LOEL dose of 0.25 mg/kg/day, mean concentrations of 1080 in rat plasma were 0.26 micro g/ml at 1 h and 0.076 microg/ml at 12 h. Rat urine collected from the same animals contained 0.059 microg/ml.

Photoclastogenicity-an improved protocol, its validation, and investigation of the photogenotoxicity of DMBA.

An improved protocol was developed to detect light-induced clastogenic photoproducts in Chinese hamster ovary (CHO) cells. Dishes (60 mm) containing cells and the test material or vehicle control in 3 mL of phosphate-buffered saline were exposed to light using a SUNTEST CPS solar simulation unit. Importantly, cells were exposed at about 25 cm from the light source, thereby allowing a short exposure time of 2 min. With this exposure the assay was conducted with lids removed during the UV exposure with minimal risk of contamination. After preliminary experiments an exposure of 165.6 mJ/cm(2) UVA: 17.0 mJ/cm(2) UVB was selected for treatments with the different phototoxins. Under these exposure conditions about 10-15% aberrant cells were induced in vehicle control cultures with no or minimal cytotoxicity. The well-known photoclastogens 8-methoxypsoralen (8-MOP) and chlorpromazine (CLZ) were tested. In agreement with published data, 8-MOP and CLZ were clastogenic (lowest observed effect level, LOEL, was 0.0159 microg/mL and 1.03 microg/mL, respectively). In the absence of UV, 8-MOP was clastogenic at a much higher concentration (LOEL 251 microg/mL without UV vs. 0.0159 microg/mL with UV) while CLZ was negative up to a toxic concentration of 35 microg/mL. 7,12-Dimethylbenz[a]anthracene (DMBA), which is photomutagenic in bacteria, was clastogenic at > or =0.005 microg/mL with UV light (without S9) and at > or =2.53 microg/mL with S9 (without UV light). These results demonstrate the utility of the protocol for the detection of photoclastogenicity and expand the characterization of DMBA's photogenotoxic activity.

Immunotoxicological investigations on rats treated subacutely with dimethoate, As3+ and Hg2+ in combination.

Effects of combined exposure with dimethoate (DM), HgCl2 (Hg), and NaAsO2 (As) were investigated following a 28 - day oral exposure in male Wistar rats. In preliminary experiments, the LOEL (Lowest Observed Effect Level) and NOEL (Non Observed Effect Level) doses of the substances were determined using the same experimental system [determination of body weight gain, organ weights of brain, thymus, heart, lung, kidneys, adrenals, spleen, testicles, popliteal lymph node, white blood cell (WBC) and red blood cell (RBC) count, haematocrit (Ht), mean cell volume (MCV) of RBCs, cell content of the femoral bone marrow, IgM-plaque forming cell (PFC) content of the spleen, delayed type hypersensitivity (DTH) reaction] and animal strain. In the combination studies, LOEL dose of DM (28.2 mg/kg) was combined with NOEL doses of the heavy metals ( HgCl2 = 0.40 mg/kg, NaAsO2 = 3.33 mg/kg), and vice versa (DM = 7.04 mg/kg, HgCl2 = 3.20 mg/kg, NaAsO2 = 13.3 mg/kg). In the DM-Hg combinations, significant alterations were found versus the corresponding high- dose internal control in the body weight gain, relative liver and kidney weights, and in the PFC response. When DM was combined with As, interactions were indicated by changes of relative liver weight, MCV value, and the PFC content of the spleen. These results support the theory that the interactions between pesticides and heavy metals may modify the toxic effects of the single substances, and may also change the functional detection limits of the exposure. The changes in the functional detection limits, if they occur, can lead to false-positive and false-negative results in human epidemiological studies.

An exploratory quantitative risk assessment for high molecular weight sensitizers: wheat flour

Quantitative risk assessments have been made for wheat dust and allergen exposure and wheat sensitization using classical epidemiological approaches based on simple categorizations in exposure groups. Such analyses suggest the existence of an exposure threshold level for wheat specific sensitization and were used as input in recently conducted risk assessments for wheat flour by the American Conference of Governmental Industrial Hygienists and the Dutch Expert Committee on Occupational Standards. More advanced statistical analyses were applied using generalized additive modeling and smoothed plots to evaluate the shape of the exposure response relationship in greater detail and evaluate the presence of exposure thresholds. Data were used from a recently conducted epidemiological study in bakery workers. Information was available on wheat sensitization (IgE antibodies), inhalable dust levels and wheat allergen levels. Initial analyses were based on simple exposure categorizations for inhalable dust and allergen exposure. A more detailed analysis using non-parametric generalized additive models (GAM models) and smoothing plots allowed inspection of the presence of an exposure threshold of evaluation of 'no' or 'lowest observed effect levels' (NOELs, LOELs) using exposure data on the individual level. All analyses showed an increasing sensitization risk with increasing exposure. The classical epidemiological analyses gave evidence for the existence of an exposure threshold or 'no observed effect level (NOEL)' for specific wheat sensitization between 0.5 and 1 mg/m3 of inhalable dust. The more advanced analyses did not suggest any evidence for the existence of an exposure threshold. However, estimates of a LOEL obtained by considering an arbitrary increase in sensitization risk between 1.5 and 2 as undesirable, were close to the NOEL from the classical analyses and would therefore not lead to an essentially different exposure limit. The criterion of an increase in wheat sensitization risk was based on the risk in non-wheat dust exposed populations. Exposure response modeling using different classical epidemiological approaches and advanced statistical methods resulted in health based LOEL or NOEL estimates within a relatively close range. But when sensitization accompanied by asthma or rhinitis symptoms was considered as critical endpoint, steeper exposure-response relationships were observed which would lead to lower LOEL values.

An exploratory quantitative risk assessment for high molecular weight sensitizers: wheat flour

Objective: Quantitative risk assessments have been made for wheat dust and allergen exposure and wheat sensitization using classical epidemiological approaches based on simple categorizations in exposure groups. Such analyses suggest the existence of an exposure threshold level for wheat specific sensitization and were used as input in recently conducted risk assessments for wheat flour by the American Conference of Governmental Industrial Hygienists and the Dutch Expert Committee on Occupational Standards. More advanced statistical analyses were applied using generalized additive modeling and smoothed plots to evaluate the shape of the exposure response relationship in greater detail and evaluate the presence of exposure thresholds.Methods: Data were used from a recently conducted epidemiological study in bakery workers. Information was available on wheat sensitization (IgE antibodies), inhalable dust levels and wheat allergen levels. Initial analyses were based on simple exposure categorizations for inhalable dust and allergen exposure. A more detailed analysis using non-parametric generalized additive models (GAM models) and smoothing plots allowed inspection of the presence of an exposure threshold of evaluation of ‘no’ or ‘lowest observed effect levels’ (NOELs, LOELs) using exposure data on the individual level.Results: All analyses showed an increasing sensitization risk with increasing exposure. The classical epidemiological analyses gave evidence for the existence of an exposure threshold or ‘no observed effect level (NOEL)’ for specific wheat sensitization between 0.5 and 1 mg/m3 of inhalable dust. The more advanced analyses did not suggest any evidence for the existence of an exposure threshold. However, estimates of a LOEL obtained by considering an arbitrary increase in sensitization risk between 1.5 and 2 as undesirable, were close to the NOEL from the classical analyses and would therefore not lead to an essentially different exposure limit. The criterion of an increase in wheat sensitization risk was based on the risk in non-wheat dust exposed populations.Conclusion: Exposure response modeling using different classical epidemiological approaches and advanced statistical methods resulted in health based LOEL or NOEL estimates within a relatively close range. But when sensitization accompanied by asthma or rhinitis symptoms was considered as critical endpoint, steeper exposure–response relationships were observed which would lead to lower LOEL values.

A Toxicokinetic Study of Inhaled Ethylene Glycol Monomethyl Ether (2-ME) and Validation of a Physiologically Based Pharmacokinetic Model for the Pregnant Rat and Human

Exposures to sufficiently high doses of ethylene glycol monomethyl ether (2-methoxyethanol, 2-ME) have been found to produce developmental effects in rodents and nonhuman primates. The acetic acid metabolite of 2-ME, 2-methoxyacetic acid (2-MAA), is the likely toxicant, and, as such, an understanding of the kinetics of 2-MAA is important when assessing the potential risks to humans associated with 2-ME. A previously described physiologically based pharmacokinetic (PBPK) model of 2-ME/2-MAA kinetics for rats exposed via oral or iv administration was extended and validated to inhalation exposures. Pregnant Sprague–Dawley rats were exposed for 5 days (gestation days 11–15), 6 h/day, to 2-ME vapor at 10 and 50 ppm. Validation consisted of comparing model output to maternal blood and fetal 2-ME and 2-MAA concentrations during and following 5 days of exposure (gestation days 11–15). These concentrations correspond to a known no observed effect level (NOEL) and a lowest observed effect level (LOEL) for developmental effects in rats. The rat PBPK model for 2-ME/2-MAA was scaled to humans and the model (without the pregnancy component) was used to predict data collected by other investigators on the kinetics of 2-MAA excretion in urine following exposures to 2-ME in human volunteers. The partially validated human model (with the pregnancy component) was used to predict equivalent human exposure concentrations based on 2-MAA dose measures (maximum blood concentration, Cmax, and average daily area under the 2-MAA blood concentration curve, AUC, during pregnancy) that correspond to the concentrations measured at the rat NOEL and LOEL exposure concentrations. Using traditional PBPK scale-up techniques, it was calculated that pregnant women exposed for 8 h/day, 5 days/week, for the duration of pregnancy would need to be exposed to 12 or 60 ppm 2-ME to produce maternal 2-MAA blood concentrations (Cmax or average daily AUC) equivalent to those in rats exposed to the NOEL (10 ppm) or LOEL (50 ppm), respectively.

A Toxicokinetic Study of Inhaled Ethylene Glycol Ethyl Ether Acetate and Validation of a Physiologically Based Pharmacokinetic Model for Rat and Human

The solvents ethylene glycol monoethyl ether acetate (EGEEA) and ethylene glycol monoethyl ether (EGEE), at sufficiently high doses, are known to be rodent developmental toxicants, exerting their toxic effects through the action of their metabolite 2-ethoxyacetic acid (2-EAA). Thus risks associated with exposure to these compounds are best evaluated based on a measure of the internal dose of 2-EAA. The goals of the work reported here were to develop physiologically based pharmacokinetic (PBPK) models of EGEEA and EGEE for pregnant rats and humans. These models were used to identify human exposure levels (ppm in air) equivalent to the rat no observed effect level (NOEL) and lowest observed effect level (LOEL) for developmental effects (Hanley et al., 1984). We exposed pregnant Sprague–Dawley rats to concentrations of EGEEA corresponding to the NOEL and LOEL. Maternal blood, urine, and fetal tissue concentrations of EGEE and 2-EAA measured in these experiments were used to validate the rat EGEEA and EGEE models. Data collected by other researchers were used to validate the capabilities of the rodent EGEEA and EGEE models to predict the kinetics in humans. The models for estimating circulating blood concentrations of 2-EAA were considered valid based on the ability of the model to accurately predict 2-EAA concentrations in rat blood, urine, and fetal tissue. The human inhaled concentration equivalent to the rat NOEL for EGEEA (50 ppm) was predicted to be 25 ppm using the maternal blood average daily area under the curve (AUC) and 40 ppm using the maximum concentration achieved in maternal blood (Cmax). The human inhaled concentration equivalent to the rat LOEL for EGEEA (100 ppm) was determined to be 55 ppm using the maternal blood average daily AUC and 80 ppm using the maternal blood Cmax.

An Integrated Approach to the Prediction of Systemic Toxicity using Computer-based Biokinetic Models and Biological In vitro Test Methods: Overview of a Prevalidation Study Based on the ECITTS Project

Chemical toxicity was estimated by integrating in vitro study results with physiologically-based biokinetic models for eight neurotoxic compounds (benzene, toluene, lindane, acrylamide, parathion/oxon, caffeine, diazepam and phenytoin). In vitro studies on general and specific neurotoxicity were performed and biotransformation and tissue–blood distribution studies were used in modelling the biokinetic behaviour of the compounds. Subsequently, neurotoxicity was estimated from the integrated in vitro and kinetic studies. These results were compared with in vivo data from the literature on minimal neurotoxicity for these compounds, such as lowest-observed-effect levels (LOELs). The discrepancy between estimated and experimental LOELs ranged from 2- to 10-fold. LOEL estimates for compounds with a relatively low toxicity were more accurate than for compounds with a relatively high toxicity. LOELs for the most active compounds could only be established after consideration of additional in vitro results from the literature. The present study has generated encouraging results on the risk assessment of chemicals from in vitro studies and computer simulations and has identified some key directions for future research.

Toxicity of Mercury to Hybridoma TA7 Cells.

Environmental mercury and mercury compound contamination has increased dramatically since the industrial revolution. This paper describes the toxic effects of mercury on a culture of hybridoma TA7 cells, which produce antibodies against the A-subunit of viskumin. Cells were cultivated on 96-well flat-bottomed plates with RPMI-1640 medium supplemented with 10% fetal calf serum at 37°C in 5% CO2/95% air. The cells were exposed to 0.1nM/l-10μM/l Hg2(NO3)2.2H2O (mercury nitrate) during the exponential growth phase. Toxicity was assessed by using the colorimetric MTT (tetrazolium) assay after exposure for 48 hours. Cell growth and cell survival were evaluated by using percentage indices of cellular content in exposed cells when compared to non-exposed control cells. The concentrations of the no-effect level, the lowest observed effect level and the highest toxic effect level were registered. The toxic effects of the mercury compound on the hybridoma cells occurred between 0.1μM/l and 10μM/l.

REGULATING AND ASSESSING RISKS OF CHOLINESTERASE-INHIBITING PESTICIDES: DIVERGENT APPROACHES AND INTERPRETATIONS

This document presents a revised framework for conducting worker and dietary risk assessments for less-than-lifetime exposures to organophosphate or carbamate pesticides based on red blood cell (RBC) or brain acetylcholinesterase (AChE) inhibition or the presence of clinical signs and symptoms. The proposals for appropriate uncertainty factors are based on the biological significance of the cholinesterase (ChE) inhibition noted at the lowest-observed-effect level (LOEL) and the degree of uncertainty in the extrapolation between human and animal data. An extensive evaluation of industry data, not previously summarized, and the available literature indicate that the following risk assessment principles are supportable and protective of human health: Plasma ChE inhibition is not an adverse effect, and therefore should not be utilized in risk assessments. Red blood cell AChE is not associated with the nervous system and inhibition is not per se an adverse (neurotoxic) effect. When available, cholinergic effects or brain AChE inhibition data should take precedence over RBC AChE for determining no-observed-effect levels (NOELs). When available, human RBC AChE inhibition or cholinergic effects data should take precedence over animal data for determining NOELs. Due to the lack of adversity associated with inhibition of RBC AChE, the use of a 10- fold (10x) uncertainty factor from the NOEL is adequate when RBC AChE inhibition data from either animal or human studies are used to assess human risk. Due to greater potential for adversity, NOELs for brain AChE inhibition and cholinergic effects identified in animal studies should receive a default uncertainty factor of 100x; lower uncertainty factors may be used on a case-by-case basis. NOELs based on cholinergic effects noted in human studies should only require a 10x uncertainty factor, since an interspecies extrapolation factor from animals to humans is unnecessary. For RBC and brain AChE activity the threshold for defining a NOEL should be less than or equal to 20% difference from control activity in all species. For risk assessment purposes, duration and route of the study should reflect the expected duration and route of exposure for humans (i.e., a 21-d or 28-d dermal study for subchronic occupational dermal exposure assessment). When dermal data are not available, a subchronic oral toxicity study and an appropriate dermal penetration factor should be used. A general default of 10% absorption should be used, analogous to the United Kingdom and German exposure models that are widely used in Europe. The recommendations in this document are generally consistent with current risk assessment procedures used by Canada, the European Community (EC), and the United Kingdom (UK).

Effects of tributyltin on medaka ( Oryzias latipes) embryos at different stages of development

Potential mechanisms of action for the toxicity of tributyltin (TBT) were studied in the freshwater fish embryos of medaka ( Oryzias latipes). Toxic concentrations of TBT have been found in estuaries and freshwater systems, presumably due to their use as biocides in boat, antifoulant paints and in industry for plastics production. Medaka embryos were exposed to a single concentration of TBT at developmental stages that corresponded to the formation of structures and/or organs which might be potential targets. Times of exposure included day 0, oviposition, day 3, completion of somite formation, and day 5, liver formation. Endpoints for evaluating toxicity were acute embryo lethality (96 h), rate of embryo development, hatching success, gross abnormalities, as well as hatchling eye diameter and number of somites. The clear chorion of medaka embryos allowed staging and in ova observations. Results showed that the acute toxicity of TBT was stage related. The 96 h LC 50 (LC 50: lowest concentration to cause 50% lethality in the test population) for embryos exposed on day 0 was 159 nM, which was lower than that for days 3 and 5, 360 and 340 nM, respectively. Subchronic endpoints showed that toxicity was concentration related and that embryos exposed on day 0 were more sensitive than those exposed on days 3 and 5. Lowest observable effect levels (LOELs) for hatching success were 36 nM for day 0 and 143 nM for days 3 and 5. LOELs for the combined effects of hatching success and gross abnormalities were 36 nM for day 0 and 71 nM for days 3 and 5. Developmental rate was slowed by TBT in a concentration-related manner; however, embryos treated with 36 and 71 nM were able to recover and hatch at the same time as controls. Types of gross abnormalities were similar regardless of day of exposure and consisted of tails bent at the tip, curled, and/or shortened. These abnormalities corresponded with statistically significant reductions in numbers of somites in all three age groups exposed to 71 nM ( P<0.05). Although day 0 embryos were the most sensitive, the similar abnormalities for all 3 days of exposure indicated that TBT's toxicity was not due to effects on an age-dependent target but one present throughout embryo development.

Acute effects of 3,4-dichloroaniline on blood of male wistar rats

In this work we investigated the acute effects of 3,4-dichloroaniline (DCA) on the changes of specific biochemical and cellular blood parameters using male Wistar rats, 24 hours after the intraperitoneal administration of five different doses of DCA (0, 81, 162, 324, 486, and 568 mg/Kg of body weight). We also evaluated the dose-dependent toxicity of DCA on the ratio “organ weight: total body weight” for kidney, liver, spleen and thymus, and on the changes of morphological characteristics of the spleen. The LOEL (lowest observed effect level) value for spleen and thymus injury was 324 mg DCA/Kg. The cellular blood parameters affected by DCA include: methemoglobin concentration, the number of circulating leukocytes and the number and size of platelets. The relative sensitivity of all the analysis used to assess the acute toxicity of DCA was evaluated by comparing their ED50 values, which ranged from 224 to 837 mg DCA/Kg. We concluded that the most sensitive endpoint to measure the acute toxicity of DCA was the methemoglobin formation. Platelet counts, urea and creatinine showed ED50 values slightly higher than methemoglobin (329 – 367 mg/Kg).

Heritable and cancer risks of exposures to anticancer drugs: inter-species comparisons of covalent deoxyribonucleic acid-binding agents

In the past years, several methodologies were developed for potency ranking of genotoxic carcinogens and germ cell mutagens. In this paper, we analyzed six sub-classes of covalent deoxyribonucleic acid (DNA) binding antineoplastic drugs comprising a total of 37 chemicals and, in addition, four alkyl-epoxides, using four approaches for the ranking of genotoxic agents on a potency scale: the EPA/IARC genetic activity profile (GAP) database, the ICPEMC agent score system, and the analysis of qualitative and quantitative structure-activity and activity-activity relationships (SARs, AARs) between types of DNA modifications and genotoxic endpoints. Considerations of SARs and AARs focused entirely on in vivo data for mutagenicity in male germ cells (mouse, Drosophila), carcinogenicity (TD 50s) and acute toxicity (LD 50s) in rodents, whereas the former two approaches combined the entire database on in vivo and in vitro mutagenicity tests. The analysis shows that the understanding and prediction of rank positions of individual genotoxic agents requires information on their mechanism of action. Based on SARs and AARs, the covalent DNA binding antineoplastic drugs can be divided into three categories. Category 1 comprises mono-functional alkylating agents that primarily react with N7 and N3 moieties of purines in DNA. Efficient DNA repair is the major protective mechanism for their low and often not measurable genotoxic effects in repair-competent germ cells, and the need of high exposure doses for tumor induction in rodents. Due to cell type related differences in the efficiency of DNA repair, a strong target cell specificity in various species regarding the potency of these agents for adverse effects is found. Three of the four evaluation systems rank category 1 agents lower than those of the other two categories. Category 2 type mutagens produce O-alkyl adducts in DNA in addition to N-alkyl adducts. In general, certain O-alkyl DNA adducts appear to be slowly repaired, or even not at all, which make this kind of agents potent carcinogens and germ cell mutagens. Especially the inefficient repair of O-alkyl—pyrimidines causes the high mutational response of cells to these agents. Agents of this category give high potency scores in all four expert systems. The major determinant for the high rank positions on any scale of genotoxic of category 3 agents is their ability to induce primarily structural chromosomal changes. These agents are able to cross-link DNA. Their high intrinsic genotoxic potency appears to be related to the number of DNA cross-links per target dose unit they can induce. A confounding factor among category 3 agents is that often the genotoxic endpoints occur closed to or toxic levels, and that the width of the mutagenic dose range, i.e., the dose area between the lowest observed effect level and the LD 50, is smaller (usually no more than 1 logarithmic unit) than for chemicals of the other two categories. For all three categories of genotoxic agents, strong correlations are observed between their carcinogenic potency, acute toxicity and germ cell specificity.

Comparison of Fresh and Room-Aged Cigarette Sidestream Smoke in a Subchronic Inhalation Study on Rats

Two experimental types of cigarette sidestream smoke (SS) were compared in a subchronic inhalation study on rats. Fresh SS (FSS) was generated continuously from the reference cigarette 2R1. Room-aged SS (RASS) was generated by aging FSS for 1.5 h in a room with noninert surfaces with materials typically found in residences or offices. Male Sprague-Dawley rats were head-only exposed to three dose levels of each SS type and to filtered, conditioned fresh air (sham-exposure) for 6 h/day, 7 days/week, for 90 days. Room-aging resulted in decreased concentrations of various SS components, e.g., total participate matter (TPM) and nicotine, while other components, such as carbon monoxide (CO), were not affected. The CO concentrations were 6, 13, and 28 ppm for both SS types. TPM concentrations were between 0.6 and 8.7 (μg/liter and thus up to 100-fold above the maximum of average concentrations of respiratory suspended particles reported for environmental tobacco smoke. Slight reserve cell hyperplasia in the anterior part of the nose as well as hyperplastic and metaplastic epithelial changes in the larynx were the only observed dose-dependent findings. The metabolism of benzo(a)-pyrene-as a proxy for polycyclic aromatic hydrocarbon metabolism-was induced in the nasal respiratory epithelium and in the lungs while no effect was seen in the nasal olfactory epithelium. The lowest-observed effect level was 6 ppm CO or 0.6 μg TPM/liter. Most of the effects seen were less expressed in RASS- than in FSS-exposed rats when compared on the basis of the CO concentrations. When compared on the basis of TPM, these effects were equally pronounced for both SS types, suggesting a major role of particulate matter-associated compounds. All findings reverted to sham control levels following a 42-day postinhalation period.

SYSTEMIC TOXICITY OF DERMALLY APPLIED CRUDE OILS IN RATS

Two crude oils, differing in viscosity (V) and nitrogen (N) and sulfur (S) content, were evaluated for systemic toxicity. In the Crude I (low V, low N, low S) study, the material was applied to the clipped backs of rats at dose levels of 0, 30, 125, and 500 mg/kg. In the Crude II (high V, high N, moderate S) study, the oil was applied similarly at the same dose levels. The crude oils were applied for 13 wk, 5 d/wk. Exposure sites were not occluded. Mean body weight gain (wk 1-14) was significantly reduced in male rats exposed to Crude II; body weight gain of all other animals was not adversely affected by treatment. An increase in absolute (A) and relative (R) liver weights and a decrease in A and R thymus weights were observed in male and female rats exposed to Crude II at 500 mg/kg; only liver weights (A and R) were adversely affected in male and female rats exposed to Crude I. In general, there was no consistent pattern of toxicity for serum chemistry endpoints; however, more parameters were adversely affected in Crude II-exposed female rats than in the other exposed groups. A consistent pattern of toxicity for hematology endpoints was observed among male rats exposed to Crude I and male and female rats exposed to Crude II. Parameters affected included: Crudes I and II, red blood cell count, hemoglobin, and hematocrit; Crude II, platelet count. Microscopic evaluation of tissues revealed the following treatment-related findings: Crude I, treated skin, thymus, and thyroid; Crude II, bone marrow, treated skin, thymus, and thyroid. The LOEL (lowest observable effect level) for skin irritation and systemic toxicity (based on marginal effects on the thyroid) for both crude oils was 30 mg/kg; effects were more numerous and more pronounced in animals exposed to Crude II. Systemic effects are probably related to concentrations of polycyclic aromatic compounds (PAC) found in crude oil.

Estimating low toxic effects

AbstractThe use of no‐observed‐effect levels (NOLs) and lowest‐observed‐effect levels (LOELs) assummary statistics for low toxic effects has been criticized recently. In this paper, we explore the regression‐based approach for estimating low toxic effects. Key questions with this approach include: (1) is the approach practical with typical toxicity data sets, (2) can low toxic effects be estimated with confidence, (3) what models are appropriate for typical data sets and endpoints, and (4) how do point estimates of low toxic effects (e.g., EC10) compare with their corresponding LOELs and NOELs? Analyses of 198 toxicity data sets (all but 13 unpublished) revealed that: (1) even with a range of models to choose from (three models in the logistic family, probit model, Weibull model), &gt;80% of the data sets did not produce a single adequate model fit; (2) estimates of low toxic effects were often model dependent when an extrapolation beyond the toxicity data was required; (3) confidence intervals can be quite large at 5% effect and lower; (4) of the five models, the three‐parameter logistic equation with a steep slope parameter had the best model fit in the majority of the data sets; and (5) 76.9% of the NOELs and 100% of the LOELs were higher than their corresponding 10% effect point estimates from the best‐fit model equations, suggesting that NOELs and LOELs are poor indicators of low toxic effects. We believe that the regression‐based approach is a better tool than hypothesis testing for estimating low toxic effects. The approach, however, does not produce adequate model fits unless there is an obvious concentration–response relationship and several treatments with partial effects.

Additive Interactions between Pairs of Polybrominated Dibenzo-p-dioxin, Dibenzofuran, and Biphenyl Congeners in a Rainbow Trout Early Life Stage Mortality Bioassay

Use of fish-specific toxic equivalency factors (TEFs) to estimate the risk that exposure to polybrominated dibenzo-p-dioxins (PBDDs), dibenzofurans (PBDFs), and biphenyls (PBBs) pose to fish early life stage survival depends on validation of the hypothesis that these chemicals act additively to produce mortality. A rainbow trout early life stage bioassay was used to determine how pairs of PBDD, PBDF, and PBB congeners interact to produce 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-like toxicity associated with sac fry mortality. The congener pairs tested were 2,3,7,8-tetra- bromodibenzo-p-dioxin (2,3,7,8-TBDD)/1,2,3,7,8-pentabromodi- benzop-dioxin (1,2,3,7,8-PBDD); 2,3,7,8-TBDD/1,2,3,7,8-penta- bromodibenzofuran (1,2,3,7,8-PBDF); 1,2,3,7,8-PBDD/2,3,4,7,8-pentabromodibenzofuran (2,3,4,7,8-PBDF); and 2,3,4,7,8-PBDF/3,3′,4,4′-tetrabromobiphenyl (3,3′,4,4′-TBB). Graded doses of each congener alone, or graded doses of fixed ratios of paired congeners were injected into newly fertilized rainbow trout eggs. In all cases, interactions between congener pairs were additive as tested by a probit model. Isobolographic analysis also supported the hypothesis that the PBDD, PBDF, and PBB congeners act additively. Thus, the use of fish-specific TEFs to convert fish tissue concentrations of individual PBDD, PBDF, and PBB congeners to TCDD equivalents (TEs) and then adding the TEs contributed by the various congeners to give the total TCDD equivalents concentration (TEC) in the tissue is supported by these results. By comparing the TEC in feral fish eggs to the fish egg TCDD no-observed-effect level (NOEL) and lowest-observed-effect level (LOEL) for early life stage mortality, the risk that complex mixtures of these polybrominated chemicals in eggs pose to sac fry survival can be estimated.

Biological effects of polychlorinated dibenzo‐p‐dioxins, dibenzofurans, and biphenyls in bald eagle (Haliaeetus leucocephalus) chicks

AbstractDuring the1992 breeding season, eggs of bald eagles (Haliaeetus leucocephalus) were collected within a gradient of exposure to chlorinated hydrocarbon pollutants, particularly from pulp mill point sources, on the southern coast of British Columbia, Canada. Twenty‐five eggs were placed in a laboratory incubator, of which 18 hatched; chicks were sacrificed within 24 h. Hatching success was not significantly different between eggs taken from pulp mill sites and reference sites. A hepatic cytochrome P4501A (CYP1A) cross‐reactive protein was induced nearly sixfold in chicks from near a pulp mill at Powell River compared to those from a reference site (p &lt; 0.05). Hepatic ethoxyresorufin‐O‐deethylase (EROD) and benzyloxyresorufin O‐dealkylase (BROD) activities were also significantly elevated in chicks from nests located near pulp mills compared to reference sites (p &lt; 0.0005 and p &lt; 0.02, respectively). A hepatic CYP2B cross‐reactive protein was threefold higher in chicks from pulp mill versus reference sites, but the difference was not significant. Residual yolk sacs of eggs collected near pulp mill sites contained greater concentrations of 2,3,7,8‐substituted polychlorinated dibenzo‐p‐dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) compared to reference areas. No significant differences in concentrations of polychlorinated biphenyls (PCBs), non‐ortho congeners, and organochlorine pesticides occurred among sites. Regressions showed that the hepatic CYP1A cross‐reactive protein and EROD and BROD activities were positively correlated with 2,3,7,8‐TCDD, 2,3,7,8‐TCDF, and toxic equivalents (TEQsWHO—Wrld Health Organization toxic equivalence factors) in yolk sacs. No significant concentration‐related effects were found for morphological, physiological, or histological parameters, such as chick growth, edema, or density of thymic lymphocytes. Using hepatic CYP1A induction as a biomarker, a no‐observed‐effect‐level (NOEL) of 100 ng/kg and a lowest‐observed‐effect‐level (LOEL) of 210 ng/kg TEQsWHO on a whole egg (wet weight basis) are suggested for bald eagle chicks.

Hepatocarcinogenic potential of di(2-ethylhexyl)phthalate in rodents and its implications on human risk.

The plasticizer di(2-ethylhexyl) phthalate (DEHP), to which humans are extensively exposed, was found to be hepatocarcinogenic in rats and mice. DEHP is potentially set free from objects made of synthetic materials (e.g., those used in medicine). Chronically, the greatest amounts are transferred to persons undergoing hemodialysis (up to 3.1 mg/kg b.w. per day) who would thus be considered the individuals most endangered by tumorigenesis. Although toxicokinetics seem to play a certain unclear role in the course of DEHP-related toxicity, toxicodynamic factors appear more decisive. DEHP is a representative of "peroxisome proliferators" (PP), a distinct group of substances that, in rodents, do not only induce peroxisomes but also specific enzymes in other organelles, organ growth, and DNA synthesis. The cluster of the characteristic effects of PP is generally, although perhaps not quite appropriately summarized as "peroxisome proliferation," and is strongest in the liver. The lowest observed effect level (LOEL) and the no observed effect level (NOEL) of peroxisome proliferation in the rat, as determined by the induction of specific enzymes (peroxisomal beta-oxidation, carnitine-acetyl-transferase, cytochrome P-452), DNA synthesis, and hepatomegaly, may be assumed as 50 and 25 mg/kg b.w. per day, respectively. DEHP and other carcinogenic PP are neither genotoxic nor tumor initiators, but they appear to be tumor promoters, also implicating a threshold level for the carcinogenic effect. Although a causal relationship between a particular effect of peroxisome proliferation and hepatocarcinogenesis is as yet unknown, peroxisome proliferation as a whole phenomenon appears to be associated with the potential of tumor induction, as shown by comparison of the relative strength of individual PP and by comparison of species and organ specificities. Likewise, LOEL and NOEL of rodent carcinogenesis, that is, 300 and 50 to 100 mg/kg b.w. per day, respectively, are above but not too far from the corresponding values for the investigated parameters of peroxisome proliferation. Thus, with respect to dose alone, worst-case exposure in hemodialysis patients is at least 16-fold below the LOEL of any characterized PP-specific effect of DEHP and approximately 100-fold below that of DEHP-related tumorigenesis. Also, primates are less responsive to PP than rats with respect to the investigated biochemical and morphological parameters. If this lower primate responsiveness is extrapolated to estimate carcinogenicity in humans, we might thus arrive at an even larger safety margin than when based on exposure alone. Doses of PP hypolipidemics that had clearly induced several indicators of peroxisome proliferation in rats did not cause any clear-cut enhancements in the peroxisomes of patients, even though most of these hypolipidemics were considerably stronger PP than DEHP. Thus, an actual threat to humans by DEHP seems rather unlikely. Accordingly, hepatocarcinogenesis was neither enhanced in workers exposed to DEHP nor in patients treated with hypolipidemics.

Demonstration of benzo(a)pyrene-induced DNA damage in mice by alkaline single cell gel electrophoresis: evidence for strand breaks in liver but not in lymphocytes and bone marrow.

Alkaline single cell gel electrophoresis (also known as the 'comet assay') was used to measure DNA strand breaks and alkali-labile sites in peripheral lymphocytes, bone marrow and liver cells of C57BL/6 mice orally exposed to benzo(a)pyrene. Although this polycyclic aromatic hydrocarbon is a well-known genotoxic agent, little is known about to what extent it actually induces DNA strand breaks in peripheral lymphocytes and other tissues after in vivo exposure. Significant and dose-related damage was observed in liver cells after three days of exposure (lowest observed effect level being 3 x 100 mg benzo(a)pyrene/kg b.wt. No such damage could be observed in the lymphocytes and bone marrow cells even after administration of 3 x 150 mg benzo(a)pyrene/kg b.wt. The reference substance cyclophosphamide produced pronounced DNA damage in lymphocytes and bone marrow cells already in a single dose of 100 mg/kg b.wt. The present mouse study questions the usability of DNA strand breaks in peripheral lymphocytes as an indicator of benzo(a)pyrene-induced genotoxicity.