Mechanistic relationships between hepatic genotoxicity and carcinogenicity in male B6C3F1 mice treated with polycyclic aromatic hydrocarbon mixtures.

The genotoxicity of a complex mixture [neutral fraction (NF)] from a wood preserving waste and reconstituted mixture (RM) mimicking the NF with seven major polycyclic aromatic hydrocarbons (PAHs) and benzo(a)pyrene (BaP) was investigated by determining DNA adducts and tumor incidence in male B6C3F1 mice exposed to three different doses of the chemical mixtures. The peak values of DNA adducts were observed after 24 h, and the highest levels of PAH-DNA adducts were exhibited in mice administered NF + BaP, and the highest tumor incidence and mortality were also observed in this group. DNA adduct levels after 1, 7, or 21 days were significantly correlated with animal mortality and incidence of total tumors including liver, lung, and forestomach. However, only hepatic DNA adducts after 7 days significantly correlated with liver tumor incidence. Most proteins involved in DNA repair including ATM, pATR, Chk1, pChk1, DNA PKcs, XRCC1, FANCD2, Ku80, Mre11, and Brca2 were significantly lower in liver tumor tissue compared to non-tumor tissue. Expressions of proteins involved in apoptosis and cell cycle regulation were also significantly different in tumor versus non-tumor tissues, and it is possible that PAH-induced changes in these gene products are important for tumor development and growth.

DNA damage in cichlids from an oil production facility in Guatemala.

This study focused on several wetlands in Laguna del Tigre National Park (Guatemala) as part of Conservation International's Rapid Assessment Program. Sediment and water samples were collected from a laguna near Xan field, Guatemala's largest oil facility, and three other sites for determination of levels of polycyclic aromatic hydrocarbons (PAHs). Cichlid fish (Thorichthys meeki and Vieja synspila) were collected for determination of DNA strand breakage (by gel electrophoresis), chromosomal breakage (flow cytometry), and fin erosion. For T. meeki from Xan field, chromosomal breakage and strand breakage was greater than in at least two of the three reference sites. For V. synspila, chromosomal breakage and strand breakage were greater in Xan than one of the two reference sites. Fin erosion was observed only at the Xan laguna. Genetic biomarker effects and fin erosion, along with patterns of aqueous PAH concentrations, indicate that fish are affected by anthropogenic contaminants. PAHs were elevated at some reference sites, but environmental forensic analysis suggested a pyrogenic or diagenic origin. It is possible that oil field brines injected into the ground water caused fin erosion and genotoxicity in fish at Xan field, and it is also possible that pyrogenic PAHs influence levels of DNA damage in reference sites. These analyses represent one of the first efforts to examine genotoxicity in native Mesoamerican cichlids.

Evolutionary toxicology: contaminant-induced genetic mutations in mosquitofish from Sumgayit, Azerbaijan.

This study builds on a long-term program that has shown Sumgayit, Azerbaijan to contain wetlands with high levels of a diversity of chemical contaminants. Previous contaminant and biomarker studies of turtles and frogs showed a correlation between somatic chromosomal damage and chemical contaminants at Sumgayit. The objective of this study was to determine if a recently arrived species (mosquitofish) has genetic impacts similar to native species (marsh frogs) thus confirming the pattern is not the result of historical events such as glacial cycles, but is associated with recent chemical contamination. Nucleotide sequences of the mtDNA control region of invasive mosquitofish (Gambusia holbrooki) from Sumgayit were compared to mosquitofish from pristine sites in Europe and Azerbaijan and to native North American populations. Persistent heteroplasmy for a hyper-mutable simple sequence repeat and low haplotype and nucleotide diversities were observed in all invasive populations. However, Sumgayit possessed four de novo haplotypes and heteroplasmic conditions. All of the observed variable nucleotide positions were within or adjacent to a cytosine mononucleotide repeat. This repeat was within a conserved secondary structure; the region likely undergoes expansion and contraction at a rate sufficient to prevent fixation of the common 1/3 heteroplasmy. Whereas the 1/3 heteroplasmy appeared coincident with the establishment of mosquitofish in Europe, other forms of heteroplasmy resulted from contaminant-induced de novo mutations in Sumgayit. We conclude that Sumgayit is a mutational hotspot caused by legacy contaminants from chemical factories from the era of the Soviet Union.

Role of retinoic acid in the modulation of benzo(a)pyrene-DNA adducts in human hepatoma cells: implications for cancer prevention.

Carcinogen-DNA adducts could lead to mutations in critical genes, eventually resulting in cancer. Many studies have shown that retinoic acid (RA) plays an important role in inducing cell apoptosis. Here we have tested the hypothesis that levels of carcinogen-DNA adducts can be diminished by DNA repair and/or by eliminating damaged cells through apoptosis. Our results showed that the levels of total DNA adducts in HepG2 cells treated with benzo(a)pyrene (BP, 2 µM)+RA (1 µM) were significantly reduced compared to those treated with BP only (P=0.038). In order to understand the mechanism of attenuation of DNA adducts, further experiments were performed. Cells were treated with BP (4 µM) for 24h to initiate DNA adduct formation, following which the medium containing BP was removed, and fresh medium containing 1 µM RA was added. The cells were harvested 24h after RA treatment. Interestingly, the levels of total DNA adducts were lower in the BP/RA group (390 ± 34) than those in the BP/DMSO group (544 ± 33), P=0.032. Analysis of cell apoptosis showed an increase in BP+RA group, compared to BP or RA only groups. Our results also indicated that attenuation of BP-DNA adducts by RA was not primarily due to its effects on CYP1A1 expression. In conclusion, our results suggest a mechanistic link between cellular apoptosis and DNA adduct formation, phenomena that play important roles in BP-mediated carcinogenesis. Furthermore, these results help understand the mechanisms of carcinogenesis, especially in relation to the chemopreventive properties of nutritional apoptosis inducers.

In situ biomonitoring of PAH-contaminated sediments using juvenile coho salmon (Oncorhynchus kisutch).

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous marine and freshwater sediment contaminants. Extensive data exist to confirm that PAHs are toxic to aquatic receptors. However, limited information is available regarding the bioavailability and genotoxicity of sediment PAHs to aquatic organisms. This study investigated an integrated biomonitoring approach using chemical analyses and biomarkers to characterize the bioavailability and genotoxicity of a complex PAH mixture in freshwater lake sediments associated with a former manufactured gas plant (MGP). Sediment PAH genotoxicity was assessed by flow cytometry (FCM), DNA adduct (32)P-postlabeling, and erythrocyte micronuclei in juvenile coho salmon (Oncorhynchus kisutch) caged in the water column. Significant PAH-induced genotoxicity was observed with FCM and (32)P-postlabeling, but not with erythrocyte micronuclei. Chromosome damage in peripheral blood and hepatic DNA adducts correlated with sediment, but not water column PAH concentrations. Total hepatic DNA adducts in salmon caged nearest the former MGP facility was 39+/-6.5 (RALx10(9)), while salmon caged in a reference lake had 28+/-2.3 total hepatic DNA adducts per 10(9) nucleotides. These results indicate that in situ biomonitoring using biomarkers and caged fish can be a sensitive indicator of genotoxic PAHs in sediments.

DNA adducts from alkoxyallylbenzene herb and spice constituents in cultured human (HepG2) cells.

Alkoxy derivatives of allylbenzene, including safrole, estragole, methyleugenol, myristicin, dill apiol, and parsley apiol, are important herb and spice constituents. Human exposure occurs mainly through consumption of food and drinks. Safrole, estragole, and methyleugenol are weak animal carcinogens. Experimental data reveal the genotoxicity and/or carcinogenicity of some allylbenzenes; however, except for safrole, the potential capacity of allylbenzenes for forming adducts in human cellular DNA has not been investigated. In the present study, we have exposed metabolically competent human hepatoma (HepG2) cells to three concentrations (50, 150, and 450 muM) of each of the six aforementioned allylbenzenes and shown by the monophosphate (32)P-postlabeling assay that each compound formed DNA adducts. With the exception of methyleugenol, DNA adduction was dose dependent, decreasing in the order, estragole > methyleugenol > safrole approximately myristicin > dill apiol > parsley apiol. These results demonstrate that safrole, estragole, methyleugenol, myristicin, dill apiol, and parsley apiol are capable of altering the DNA in these cells and thus may contribute to human carcinogenesis.

Prioritizing environmental issues around the world: opinions from an international Central and Eastern European environmental health conference.

BACKGROUND: As the next generation of scientists enters the field of environmental health, it is imperative that they view their contributions in the context of global environmental stewardship. In this commentary, a group of international graduate students facilitated by three experienced environmental health scientists present their views on what they consider to be the global environmental health concerns of today. This group convened initially in October 2004 at an international health conference in Prague, Czech Republic. OBJECTIVES: In this report we identify perceived environmental health concerns that exist around the world, with a focus on Central and Eastern Europe. Additionally, we address these perceived problems and offers some potential solutions. DISCUSSION: At the meeting, students were invited to participate in two panel discussions. One group of young international scientists identified several significant global environmental health concerns, including air pollution, occupational hazards, and risk factors that may exacerbate current environmental health issues. The second panel determined that communication, education, and regulation were the mechanisms for addressing current environmental challenges. CONCLUSIONS: In this commentary we expand on the views presented at the meeting and represent the concerns of young investigators from nine different countries. We provide ideas about and support the exchange of information between developed and developing countries on how to handle the environmental health challenges that face the world today.

A survey of laboratory and statistical issues related to farmworker exposure studies.

Developing internally valid, and perhaps generalizable, farmworker exposure studies is a complex process that involves many statistical and laboratory considerations. Statistics are an integral component of each study beginning with the design stage and continuing to the final data analysis and interpretation. Similarly, data quality plays a significant role in the overall value of the study. Data quality can be derived from several experimental parameters including statistical design of the study and quality of environmental and biological analytical measurements. We discuss statistical and analytic issues that should be addressed in every farmworker study. These issues include study design and sample size determination, analytical methods and quality control and assurance, treatment of missing data or data below the method's limits of detection, and post-hoc analyses of data from multiple studies. Key words: analytical methodology, biomarkers, laboratory, limit of detection, omics, quality control, sample size, statistics.

Evolutionary toxicology: population-level effects of chronic contaminant exposure on the marsh frogs (Rana ridibunda) of Azerbaijan.

We used molecular methods and population genetic analyses to study the effects of chronic contaminant exposure in marsh frogs from Sumgayit, Azerbaijan. Marsh frogs inhabiting wetlands in Sumgayit are exposed to complex mixtures of chemical contaminants, including petroleum products, pesticides, heavy metals, and many other industrial chemicals. Previous results documented elevated estimates of genetic damage in marsh frogs from the two most heavily contaminated sites. Based on mitochondrial DNA (mtDNA) control region sequence data, the Sumgayit region has reduced levels of genetic diversity, likely due to environmental degradation. The Sumgayit region also acts as an ecological sink, with levels of gene flow into the region exceeding gene flow out of the region. Additionally, localized mtDNA heteroplasmy and diversity patterns suggest that one of the most severely contaminated sites in Sumgayit is acting as a source of new mutations resulting from an increased mutation rate. This study provides an integrated method for assessing the cumulative population impacts of chronic contaminant exposure by studying both population genetic and evolutionary effects.

Degradation of organophosphorus neurotoxicity in SY5Y neuroblastoma cells by organophosphorus hydrolase (OPH).

Numerous approaches have been studied to degrade organophosphorus (OP) compounds and ameliorate their toxicity. In the current study, the potential of genetically engineered organophosphorus hydrolase (OPH) enzymes to functionally biotransform OP neurotoxicants was examined by assessing effects of OPH-hydrolyzed OPs on acute and delayed indicators of neurotoxicity. SY5Y human neuroblastoma cells were used as a model test system, as these cells respond distinctly to mipafox, which produces OP-induced delayed neuropathy, and paraoxon, which does not. Short-term effects of four OPH-treated OPs on acetylcholinesterase (AChE) and neuropathy target esterase (NTE) activities were measured in retinoic acid-differentiated or undifferentiated cells, and delayed effects of OPH-treated paraoxon or mipafox on levels of neuronal cytoskeletal proteins in nerve growth factor (NGF)-differentiated cells. The anti-AChE activity of paraoxon (maximum 3 muM) and anti-NTE activity of mipafox (250 muM) in SY5Y cells were prevented by biodegradation with OPH. Anti-AChE activities of mipafox, methyl parathion, and demeton-S were partially ameliorated, depending on OP concentration. Intracellular amounts of the 200-kD neurofilament protein NF200 were unchanged after treatment with OPH-treated or buffer-treated paraoxon, as expected, as this endpoint is insensitive to paraoxon. However, NF200 levels rose in cells treated during late differentiation with OPH-treated mipafox. This finding suggests the existence of a threshold concentration of mipafox below which SY5Y cells can maintain their viability for compensating cellular damage due to mipafox in neurite elongation. These results indicate that OPH may be used to biodegrade OPs and remediate their neurotoxic effects in vitro and that AChE and NTE are suitable detectors for OPH amelioration.

Tissue-specific attenuation of endogenous DNA I-compounds in rats by carcinogen azoxymethane: possible role of dietary fish oil in colon cancer prevention.

I-compounds are bulky covalent DNA modifications that are derived from metabolic intermediates of nutrients. Some I-compounds may play protective roles against cancer, aging, and degenerative diseases. Many carcinogens and tumor promoters significantly reduce I-compound levels gradually during carcinogenesis. Colon cancer is the second leading cause of cancer death in the United States, whereas cancer of the small intestine is relatively rare. Here we have studied levels of I-compounds in DNA of colon and duodenum of male Sprague-Dawley rats treated with azoxymethane. The effects of dietary lipids (fish oil or corn oil) on colon and duodenal DNA I-compounds were also investigated. Rats fed a diet containing fish oil or corn oil were treated with 15 mg/kg azoxymethane. Animals were terminated 0, 6, 9, 12, or 24 hours after injection. I-compound levels were analyzed by the nuclease P1-enhanced (32)P-postlabeling assay. Rats treated with azoxymethane displayed lower levels of I-compounds in colon DNA compared with control groups (0 hour). However, I-compound levels in duodenal DNA were not diminished after azoxymethane treatment. Animals fed a fish oil diet showed higher levels of I-compounds in colonic DNA compared with corn oil groups (mean adduct levels for fish and corn oil groups were 13.35 and 10.69 in 10(9) nucleotides, respectively, P = 0.034). Taken together, these results support claims that fish oil, which contains a high level of omega-3 polyunsaturated fatty acids, may have potent chemopreventive effects on carcinogen-induced colon cancer. The fact that duodenal I-compounds were not diminished by azoxymethane treatment may have been due to the existence of tissue-specific factors protecting against carcinogenesis. In conclusion, our observations show that endogenous DNA adducts may serve not only as sensitive biomarkers in carcinogenesis and cancer prevention studies, but are also helpful to further our understanding of the chemopreventive properties of omega-3 fatty acids and mechanisms of carcinogenesis.

Children's mouthing and food-handling behavior in an agricultural community on the US/Mexico border.

Children's mouthing and food-handling activities were measured during a study of nondietary ingestion of pesticides in a south Texas community. Mouthing data on 52 children, ranging in age from 7 to 53 months, were collected using questionnaires and videotaping. Data on children's play and hand-washing habits were also collected. Children were grouped into four age categories: infants (7-12 months), 1-year-olds (13-24 months), 2-year-olds (25-36 months) and preschoolers (37-53 months). The frequency and type of events prompting hand washing did not vary by age category except for hand washing after using the bathroom; this increased with increasing age category. Reported contact with grass and dirt also increased with increasing age category. The median hourly hand-to-mouth frequency for the four age groups ranged from 9.9 to 19.4, with 2-year-olds having the lowest frequency and preschoolers having the highest. The median hourly object to mouth frequency ranged from 5.5 to 18.1 across the four age categories; the frequency decreased as age increased (adjusted R(2)=0.179; P=0.003). The median hourly hand-to-food frequency for the four age groups ranged from 10.0 to 16.1, with the highest frequency being observed in the 1-year-olds. Hand-to-mouth frequency was associated with food contact frequency, particularly for children over 12 months of age (adjusted R(2)=0.291; P=0.002). The frequency and duration of hand-to-mouth, object-to-mouth and food-handling behaviors were all greater indoors than outdoors. Infants were more likely to remain indoors than children in other age groups. The time children spent playing on the floor decreased with increasing age (adjusted R(2)=0.096; P=0.031). Parental assessment was correlated with hand-to-mouth activity but not with object-to-mouth activity. The highest combined (hand and object) mouthing rates were observed among infants, suggesting that this age group has the greatest potential for exposure to environmental toxins.

Matrix-immobilized organoclay for the sorption of polycyclic aromatic hydrocarbons and pentachlorophenol from groundwater.

Sorbent materials consisting of organoclay immobilized onto the surface of a solid support were evaluated for use in pentachlorophenol (PCP) and polycyclic aromatic hydrocarbon (PAH) remediation of groundwater at a creosote-contaminated Superfund site. Cetylpyridinium-exchanged low pH montmorillonite clay (CP-LPHM) was bonded to either sand (CP-LPHM/sand) or granular activated carbon (GAC) (CP-LPHM/GAC) using the free acid form of carboxymethylcellulose as an adhesive. Effluent from an oil-water separator was eluted through equal bed volumes of composite (4 g 3:2 CP-LPHM/GAC or 13 g CP-LPHM/sand), affinity-extracted, and quantitatively analyzed by GC/MS. PCP, naphthalene, fluorene, phenanthrene, pyrene, and total PAHs were initially reduced by both CP-LPHM/GAC (> or =99%, 61%, 99%, > or =99%, 97%, and 94%, respectively) and CP-LPHM/sand (90%, 70%, 94%, 95%, 93%, and 86%, respectively). Complete breakthrough of naphthalene occurred after approximately 15 h of elution through 3:2 CP-LPHM/GAC and 22 h through CP-LPHM/sand. PCP showed complete breakthrough following 18 h of elution through 3:2 CP-LPHM/GAC and 26 h through CP-LPHM/sand. However, 50% breakthrough was not attained for higher molecular weight PAHs, as fluoranthene, pyrene, benzo[a]anthracene, and chrysene continued to be greatly reduced with both 3:2 CP-LPHM/GAC (98%, 95%, 94%, and 95%, respectively) and CP-LPHM/sand (75%, 73%, 76%, and 78%, respectively) after 48 h of continuous elution. Results confirm prior studies, indicating that these organoclay-containing composites have a high capacity for contaminants found in wood preserving waste. Further, results suggest that the inclusion of CP-LPHM may be useful as part of an effective strategy for groundwater remediation of high concentrations of PCP and PAHs, in particular high molecular weight and carcinogenic PAHs.

Biodegradation of simple chemical mixtures in soil.

Exogenous microorganisms often are used to enhance bioremediation. This study compared the capabilities of two exogenous microbial cultures and an indigenous population to detoxify a Weswood silt loam soil amended with a simple chemical mixture. The first three treatments were unamended soils inoculated with either indigenous microorganisms, Pseudomonas aeruginosa, or Phanerochaete sordida. Three additional treatments consisted of soil amended with benzo[a]pyrene, pentachlorophenol, and 2,4,6-trinitrotoluene, which were inoculated with either indigenous microorganisms, P. aeruginosa, or P. sordida. Samples were collected from the soils at several time points from 0 through 540 or 720 d, sequentially extracted with methylene chloride and methanol, and analyzed for genotoxicity (using the Salmonella/microsome assay) and chemical degradation. Although the indigenous microorganisms were effective for removal of benzo[a]pyrene, the Pseudomonas bacteria exhibited slightly greater removal rates for 2,4,6,-trinitrotoluene. The fungal cultures were significantly more effective at degrading pentachlorophenol. The day 540 extracts from all model chemical-amended treatments were genotoxic. In most cases, the day 540 extracts were more genotoxic than the day 0 extracts. The results suggest that, under appropriate conditions, enriched cultures of microorganisms may have an increased capacity to degrade individual chemicals. However, the products of degradation in some cases might be more genotoxic than the parent compounds.

The effect of pentachlorophenol on DNA adduct formation in p53 wild-type and knockout mice exposed to benzo[a]pyrene.

Previous studies have shown that pentachlorophenol (PCP) has both potentiative and antagonistic effects on the genotoxicity of benzo[a]pyrene (B[a]P). It has been suggested that these effects are due to inhibition and/or induction of enzymes involved in the biotransformation of B[a]P [Carcinogenesis 16 (1995) 2643]. However, B[a]P [J. Biol. Chem. 274 (1999) 35240] and a metabolite of PCP, tetrachlorohydroquinone (TCHQ) [Chem. Biol. Interact. 105 (1997) 1], induce p53 protein synthesis in vitro. To investigate this effect further, C57BL/6Tac trp53+/+ (wild-type, WT) and C57BL/6Tac trp53-/- (knockout, KO) mice were exposed to 55 microg B[a]P/g BW alone or in combination with 25 microg/g PCP. Hepatic and lung DNA were analyzed for the major B[a]P DNA adduct, 7R,8S,9S-trihydroxy-10R-(N2-2'-deoxyguanosyl)-7,8,9,10-tetrahydro-B[a]P (BPDE-N2G) and other minor adducts using the 32P-postlabeling assay. BPDE-N2G adducts were detected in all animals exposed to B[a]P. Similar adduct levels were observed in WT mice exposed to 55 microg/g B[a]P compared with KO mice exposed to B[a]P alone or in combination with PCP. Interestingly, hepatic and lung BPDE-N2G adducts were decreased in WT mice exposed to B[a]P with PCP (P<0.05). Total DNA adducts in the liver (P<0.05) were also decreased in WT mice exposed to B[a]P and PCP. Total DNA adducts in either hepatic or lung DNA isolated from KO mice were not different in mice treated with PCP and B[a]P. These results suggest that the decrease in BPDE-N2G adducts observed in WT mice may be a result of p53 accumulation or induction of repair pathways in response to damage induced by PCP.

Characterization of calcium oscillations in normal and benzo[a]pyrene-treated clone 9 cells.

Intracellular Ca2+ oscillations induced by oxytocin and vasopressin were analyzed in a rat liver cell line (Clone 9) in order to identify mechanisms by which benzo[a]pyrene (BaP) alters Ca2+ signaling patterns in these cells. Clone 9 cells exhibit an initial Ca2+ spike, followed by Ca2+ oscillations upon oxytocin or vasopressin treatment. The range of frequencies (maximum 110 mHz) was dependent on agonist concentration with a constant amplitude less than or equal to the amount of Ca2+ generated from the inositol trisphosphate (InsP(3))-sensitive pool. This study examined contributions of extracellular and intracellular pools to the frequency of Ca2+ oscillations and the role of membrane channels, second messengers, and different pharmacological reagents on the regulation of oscillation frequency in both control and BaP-treated cells. Results indicated that the Ca2+ oscillations are mainly due to inositol 1,4,5-triphosphate (InsP(3))-sensitive stores and that extracellular Ca2+ contributes to refilling of this intracellular Ca2+ pool. The frequency of Ca2+ oscillations is also sharply affected by protein kinase C activated by phospholipase C. In BaP-treated Clone 9 cells, basal Ca2+ levels were elevated and the frequency of Ca2+ oscillations was suppressed in a dose-dependent fashion. Suppression of Ca2+ oscillations is due, at least in part, to an effect of BaP on enhanced opening of K+ channels. This was confirmed by showing that inhibition of the K+ channel opening by tetraethylammonium chloride can reverse the effect of BaP on oxytocin-induced Ca2+ oscillations, and potentially decrease the toxicity of BaP.

Comparative in vitro and in vivo genotoxicities of 7H-benzo[c]fluorene, manufactured gas plant residue (MGP), and MGP fractions.

Manufactured gas plant residue (MGP) is a complex mixture of polycyclic aromatic hydrocarbons (PAHs) that is tumorigenic in the lungs of mice. This study compared the relative genotoxicity of 7H-benzo[c]fluorene (BC), a PAH component of MGP, with MGP and MGP fractions in order to assess the contribution of BC to the genotoxicity of MGP. An MGP sample was separated into seven fractions (F1-F7) using silica gel column chromatography with petroleum ether (PE) followed by PE:acetone (99:1 v/v, then 98:2). PAHs were quantified using gas chromatography/mass spectrometry. An aliquot of F2, the fraction with the highest BC concentration and highest weighted mutagenic activity in Salmonella typhimurium strain TA98, was further separated using silica gel thin-layer chromatography with hexane. The first F2 subfraction, sF2-a, was enriched in BC and coeluting compounds and contained 35,000 ppm BC and 216,109 ppm carcinogenic PAHs (cPAHs, the sum of seven PAHs categorized by the U.S. EPA as class B2 carcinogens). The second F2 subfraction, sF2-b, contained a ninefold lower concentration of BC, with 3,900 ppm BC and 45,216 ppm cPAHs. Female ICR mice received topical application of crude MGP, crude MGP spiked with analytical-grade BC, F2, sF2-a, sF2-b, or analytical-grade BC. DNA adduct levels were analyzed by nuclease P1-enhanced (32)P-postlabeling. In lung DNA of mice receiving 0.48 or 3.0 mg/mouse, net total RAL x 10(9) values were F2, 30.8 and 87.2; sF2-a, 24.8 and 106.7; and sF2-b, 19.6 and 151.0, respectively. Mice dosed with 0.10 mg analytical-grade BC (the mass of BC in 3.0 mg sF2-a) exhibited a net total RAL x 10(9) value of 7.03 in lung DNA. This was equal to approximately 7% of the total RAL x 10(9) value produced by 3.0 mg sF2-a. Thus, although BC appears to make an appreciable contribution to pulmonary adduct formation, the results suggest that MGP components other than BC play an important role in lung DNA adduct formation following topical MGP administration.

Seasonal pesticide use in a rural community on the US/Mexico border.

An environmental measurement and correlation study of infant and toddler exposure to pesticides was carried out in a colonia south of the city of Laredo, Texas. As part of the study, homes were visited during the late spring or summer, and during the winter of 2000-2001. At each visit, families reported on their use of pesticides in and around the home and floor wipe samples were collected and analyzed for 14 organophosphate and triazine pesticides. Selection of homes was based on the presence of infants and toddlers. A total of 27 homes participated in both seasonal visits. The interval between visits was 6 +/- 1.4 months. Univariate and multivariate nonparametric analyses were carried out using SPSS statistical software. Pesticide use within the home was more often reported than outside use and showed seasonal variation in use patterns. Indoor use was primarily associated with ants and cockroaches, and secondarily with rodents. The primary room treated was the kitchen, and the primary structures treated were the floors, lower walls, and dish cupboards. Seasonal variations were not found in the use of pesticides used outside the home and outdoor use was primarily associated with ant control. Based on parent reports, most pesticides used in the homes were pyrethroids. Several of the pesticides measured in floor wipe samples, Azinphos methyl, Fonofos, and Simazine, also showed seasonal variations. However, these pesticides are used in agriculture and were not associated with reported house and yard use patterns.

Nondietary ingestion of pesticides by children in an agricultural community on the US/Mexico border: preliminary results.

An environmental measurement and correlation study of nondietary ingestion of pesticides was carried out in a colonia in south Texas. The purpose of the study was to evaluate young children's exposure to environmental levels of organophosphate (OP) pesticides in the household. Samples were collected to measure levels of OP pesticides in housedust and on children's hands. These, in turn, were compared to levels of OP pesticide metabolites in urine. A total of 52 children, 25 boys and 27 girls, participated in the spring and summer of 2000. The children were 7-53 months of age at the time of recruitment. Univariate and multivariate regression analyses were carried out using SAS statistical software. Seventy-six percent of housedust samples and 50% of hand rinse samples contained OP pesticides. All urine samples had at least one metabolite and over 95% had at least two metabolites above the limit of detection (LOD). Total OP loadings in the housedust ranged from nondetectable (nd) to 78.03 nmol/100 cm(2) (mean=0.15 nmol/100 cm(2); median=0.07 nmol/100 cm(2)); total OP loadings on the children's hands ranged from nd to 13.40 nmol/100 cm(2) (mean=1.21 nmol/100 cm(2); median=1.41 nmol/100 cm(2)), and creatinine corrected urinary levels (nmol/mol creatinine) of total OP metabolites ranged from 3.2 to 257 nmol/mol creatinine (mean=42.6; median 27.4 nmol/mol creatinine). Urinary metabolites were inversely associated with the age of the child (in months) with the parameter estimate (pe)=-2.11, P=0.0070, and 95% confidence interval -3.60 to -0.61. The multivariate analysis observed a weak association between concentrations of OP pesticides in housedust, loadings in housedust, and concentration on hands, hand surface area, and urinary levels of OP metabolites. However, hand loadings of OP pesticides were more strongly associated (r(2)=0.28; P=0.0156) with urinary levels of OP metabolites (pe=6.39; 95% CI 0.98-11.80). This study's preliminary findings suggest that surface loadings of pesticides, on hands, are more highly correlated with urinary bioassays and, therefore, may be more useful for estimation of exposure in epidemiologic studies than levels of pesticides in housedust.

A comparison of two methods for fractionating complex mixtures in preparation for toxicity analysis.

Chemical fractionation is a widely used tool for the chemical and toxicological characterization of complex mixtures. The objective of this research was to compare two frequently employed methods for fractionating a wood preserving waste (WPW) containing polycyclic aromatic hydrocarbons (PAHs) and pentachlorophenol (PCP). The first method involved fractionation of the WPW into acid, base, and neutral fractions using a liquid-liquid acid/base/neutral (A/B/N) technique. The second method utilized alumina column chromatography to produce two fractions, A1 and A2. Gas chromatography and mass spectrometry were used to quantify the chemical components in all fractions. The alumina method recovered 473,338 mg of total PAHs (tPAHs) per kilogram crude, while the A/B/N method yielded only 193,379 mg tPAHs/kg crude. In contrast, the A/B/N method recovered 13.7 mg PCP/kg crude while the alumina method yielded only 0.5 mg PCP/kg crude. Three bioassays were used to determine the toxicity of the crude extract and fractions. The neutral and A1 fractions contained the highest levels of tPAHs and benzo[a]pyrene (BaP) but failed to induce a positive response in the Salmonella/microsome assay with concentrations containing as much as 1800 and 2500 ng BaP/plate, respectively. In the Escherichia coli prophage induction assay, the acid fraction, which contained 472 mg PCP/kg fraction, induced a positive response, as did the base fraction, which did not contain detectable PCP. Significant reduction of gap junctional intercellular communication in hepatic cells occurred with the crude extract and acid, base, and neutral fractions. Overall, the results of these bioassays suggest that PCP genotoxicity was expressed in the acid fraction, whereas the cumulative genotoxicity of genotoxic PAHs appeared to be masked in the isolates from either fractionation method. The optimal fractionation method for a mixture of chlorophenols and PAHs may involve a refined hybrid method.