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New approaches to toxicity: a seven-gas predictive model and toxicant suppressants.

Two new research approaches in combustion toxicology are: 1. the prediction of smoke toxicity from mathematical equations, which are empirically derived from, experiments on the toxicological interactions of complex fire gas mixtures and 2. the use of toxicant suppressants in materials or products to prevent the formation of toxic combustion products. The predictive approach consists of burning materials using a bench-scale method that simulates realistic fire conditions, measuring the concentrations of the primary fire gases--CO, CO2, low O2, HCN, HCl, HBr, and NO2--and predicting the toxicity of the smoke using either the 6- or 7-gas N-Gas Model. These models are based on the results of toxicological studies of these primary gases as individual gases and as complex mixtures. The predicted toxic potency is checked with a small number of animal (Fischer 344 male rats) tests to assure that an unanticipated toxic gas is not generated or an unexpected synergistic or antagonistic effect has not occurred. The results indicate if the smoke from a material or product is extremely toxic (based on mass consumed at the predicted toxic level) or unusually toxic (based on the gases deemed responsible). The predictions based on bench-scale laboratory tests have been validated with full-scale room burns of a limited number of materials of widely differing characteristics chosen to challenge the system. The advantages of this new approach are 1. the number of test animals is minimized by predicting the toxic potency from the chemical analysis of the smoke, 2. smoke may be produced under conditions that simulate the fire scenario of concern, 3. fewer tests are needed, thereby reducing the overall cost of the testing and 4, information is obtained on both the toxic potency of the smoke and the responsible gases. The N-Gas Models have been developed into the N-Gas Method (described in this paper) and these results have been used in computations of fire hazard. The 6-Gas Model is now part of the international standard ISO 13344 approved by 16 member countries of the International Standards Organization (ISO) and is also included in the U.S. national standard ASTM E1678 approved by the American Society for Testing and Materials (ASTM). In addition, the 6-Gas Model is used in the American National Standard--NFPA 269--approved by the National Fire Protection Association (Quincy, MA). The second new research approach, toxicant suppressants, examines the potential of chemical compounds, which when added to a material, to inhibit or reduce the concentration of a specific toxic gas normally generated during the material's thermal decomposition. The effectiveness of this approach was demonstrated at the National Institute of Standards and Technology (NIST) when HCN generation was reduced by 90% and the resultant toxicity of the combustion products was lowered by 50% when a flexible polyurethane foam (FPU) was treated with 0.1% (by weight) cuprous oxide (Cu2O). Copper and cupric oxide (CuO) also reduced the HCN generation but were not as efficient as Cu2O. Although melamine-treated FPU foams are being promoted as more fire safe than standard foams, a melamine-treated foam generated 10 times more HCN than a foam without melamine. The addition of Cu2O to this melamine foam also reduced the HCN generation by 90%.

Animal Testing Alternatives↗

Toxicity of Bacillus thuringiensis delta-endotoxins against bean shoot borer (Epinotia aporema Wals.) larvae, a major soybean pest in Argentina.

The toxicity of seven Bacillus thuringiensis Cry protoxins was tested against neonate larvae of Epinotia aporema, a major soybean pest in Argentina and South America. The most active protoxins were Cry1Ab and Cry1Ac, with LC50 values of 0.55 and 1.39 microg/ml, respectively. Cry1Aa, Cry1Ba, Cry1Ca, and Cry9Ca protoxins were equally toxic with LC50 values about 4 microg/ml, whereas Cry1Da was not toxic. The synergistic activity of different protoxin-mixtures was also analyzed, no synergistic effect between the Cry proteins was observed, with the exception of the poorly toxic Cry1Ba/Cry1Da mixture that was slightly synergistic. The binding capacity of individual Cry1 and Cry9Ca toxins to brush border membranes of E. aporema was also determined. The non-toxic Cry1Da toxin was the only toxin unable to bind to E. aporema membranes. In addition the heterologous competition experiments showed that Cry1Ab and Cry1Ac toxins share a common binding site. Based on these data, we propose that Cry1Ab and Cry1Ac toxins could be used in the biological control of E. aporema.

Animals↗

A mixture of dioxins, furans, and non-ortho PCBs based upon consensus toxic equivalency factors produces dioxin-like reproductive effects.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD; dioxin) and related polyhalogenated aromatic hydrocarbons (PHAHs) alter the reproductive development of laboratory animals. Therefore, we exposed animals to a mixture of dioxins, furans, and polychlorinated biphenyls (PCBs) that included TCDD, 1,2,3,7,8-pentachlorodibenzo-p-dioxin (PeCDD), 2,3,7,8-tetrachlorodibenzofuran (TCDF), 1,2,3,7,8-pentachlorodibenzofuran (1-PeCDF), 2,3,4,7,8-pentachlorodibenzofuran (4-PeCDF), octachlorodibenzofuran (OCDF), 3,3',4,4'-tetrachlorobiphenyl (PCB77), 3,3',4,4',5-pentachlorobiphenyl (PCB126), and 3,3',4,4',5,5'-hexachlorobiphenyl (PCB169). The mixture composition approximated the relative abundance of these compounds in foodstuff (L. S. Birnbaum and M. J. DeVito, 1995, Toxicology Vol. 105, pp. 391-401). Following the work of Gray et al. with TCDD (1997, Toxicology and Applied Pharmacology Vol. 146, pp. 11-20), we exposed time-pregnant dams on gestation day (GD) 15 at doses up to 1.0 microgram TCDD toxic equivalency (TEQ)/kg and the development of offspring was monitored. This mixture significantly increased the time to puberty in both male and female offspring. At postnatal day (PND) 32 seminal vesicle weights were decreased; however, only ventral prostate weight was affected at PND 49 and no effects were seen at PND 63. In female offspring, the mixture caused dose-dependent increases in the incidence of vaginal thread. Ethoxyresorufin-O-deethylase (EROD) activity was higher than with TCDD the comparable TEQ exposure. Based on the slightly lowered responsiveness to the mixture, we used 2.0 microgram TEQ/kg to examine reproductive effects. This dose elicited the responses observed with 1.0 microgram TCDD/kg. Results indicate that the mixture causes a similar spectrum of effects seen with TCDD and the slightly lowered degree of response based on administered dose appears to be due to decreased transfer of mixture components to the offspring. Thus, the use of the WHO consensus TEFs (M. Van den Berg et al., 1998, Environ. Health Perspec. 106, 775-792) reasonably predicts the developmental toxicity of this mixture of dioxin-like PHAHs.

Abnormalities, Drug-Induced↗

Effect of germination on vomitoxin level in grain.

The present investigation clarified the effect of enzymes or other substances formed during the germination process on the vomitoxin level of contaminated oats. The studies found that oats containing vomitoxin germinated very poorly; the decrease in toxins was also slight. The amount of pure vomitoxin added to toxin-free grain decreased (barley 53%, oats 40%, wheat 20%) during germination (5 d). In homogenized mixture of germinated grain (2.4 and 7 d) and toxic grain no decrease in toxin amount occurred during a 1-7 day period. In contrast, when germinating toxin free grains and toxic oats in a grain mixture the toxin level decreased at first, but later rose considerably. On the basis of these results, the utilization of germination processes for the improvement of grain containing vomitoxin is of questionable value.

Animals↗

Embryotoxic and teratogenic effects of petroleum hydrocarbons in mallards (Anas platyrhynchos).

Egg surface applications of microliter quantities of crude and refined oils of high aromatic content are embryotoxic to mallards (Anas platyrhynchos) and other avian species; applications of aliphatic hydrocarbons have virtually no effect. Mallard eggs at 72 h of development were exposed to a mixture of aromatic hydrocarbons or to aromatic compounds representative to those present in crude oil to assess their toxicity. The class composition of the mixture was similar to that of South Louisiana crude oil, an American Petroleum Institute reference oil. Application of 20 microliter of the mixture reduced embryonic survival by nearly 70%. The temporal pattern of embryonic death was similar to that after exposure to South Louisiana crude oil. Embryonic growth was stunted, as reflected by weight, crown-rump length, and bill length, and there was a significant increase in the incidence of abnormal survivors. When individual classes of aromatic hydrocarbons were tested, tetracyclics caused some embryonic death at the concentrations in the mixture. When classes were tested in all possible combinations of two, no combination appeared to be as toxic as the entire mixture. Addition of the tetracyclic compound chrysene to the aromatic mixture considerably enhanced embryotoxicity, but could not completely account for the toxicity of the crude oil. The presence of additional unidentified polycyclic aromatic hydrocarbons as well as methylated derivatives of polycyclic aromatic compounds such as chrysene may further account for the embryotoxicity of the crude oil.

Animals↗

A comparative study of Ro 03-8799: racemic mixture and enantiomers.

The maximum single dose of the 2-nitroimidazole hypoxic cell radiosensitiser Ro 03-8799 is limited to 1 g/m2 by the occurrence of a well characterised acute syndrome of sweating, nausea and mental changes. In an attempt to increase the tolerable dose, the clinical toxicity of the racemic mixture was compared with that of the R- and S-enantiomers of Ro 03-8799. Twelve patients received escalating alternate doses of racemic mixture and R- or S-enantiomer, the dose levels being 0.25 g/m2, 0.5 g/m2, 0.75 g/m2 and 1.0 g/m2. Careful monitoring of the acute syndrome failed to demonstrate any consistent differences between racemic mixture and either enantiomer. This would suggest that the toxicity is not mediated via any specific central nervous system receptor. It is concluded that separation of Ro 03-8799 into its enantiomers will not enable a clinically useful increase in dosage.

Dose-Response Relationship, Drug↗

Differential induction of podocyte heat shock proteins by prolonged single and combination toxic metal exposure.

Cadmium, mercury, and arsenite are among the most abundant toxic metals (TM) in our environment, and chronic TM exposure leads to injury to the kidney's glomerular filtration barrier. The small heat shock protein hsp25, highly expressed in glomerular podocytes, is induced during development of experimental nephrotic syndrome, and hsp25 overexpression can protect cultured podocytes from injury. Because little is known about the effect of multiple TM on podocytes, we measured the response of cultured podocytes to prolonged exposures to single and multiple TM. Podocyte viability declined by approximately 50% after 3 days of treatment with 20 microM cadmium, mercury, or arsenite, and 40 microM of any of these metals was lethal. The toxicity of equimolar concentrations of two or all three metals in combination was significantly altered compared to individual metal treatments. Single TM treatments induced only modest increases in the amounts of hsp25, alphaB-crystallin, and inducible hsp70. Toxic metal combinations induced greater stress protein accumulation, especially arsenite + cadmium or arsenite + cadmium + mercury treatments, the TM mixtures with the lowest toxicity. All TM treatments caused a rapid and sustained increase in hsp25 phosphorylation. The intracellular accumulation of cadmium was greater and that of mercury was less in cells treated with TM combinations than in cells treated with a single TM. Our results showed that multiple TM effects on podocyte viability were neither additive nor synergistic and that induction of heat shock proteins correlated with increased resistance to TM injury, suggesting that induction of small heat shock proteins may play an important role in preventing TM-induced podocyte injury.

Animals↗

Toxicological characterisation of sludge from sewage treatment plants using toxicity identification evaluation protocols based on in vitro toxicity tests.

The ecotoxicological characterisation of complex mixtures, such as sludge from sewage treatment plants, is complex. Toxicity identification evaluation (TIE) protocols, developed by the United States Environmental Protection Agency (US EPA); to identify toxic pollutants in complex effluents, are useful tools in this context; to solve the difficulties in assessing unknown organic pollutants by analytical methods, the usefulness of bioassays to detect the relevant (toxic) organic compounds present in complex samples, and the possibilities of in vitro cytotoxicity tests as screening tools, offers a profitable combination. The sludge obtained from a sewage treatment plant was extracted by acetonitrile using a microwave extractor and fractionated in an HPLC system. The toxicity of every fraction was assayed using a RTG-2 cytotoxicty test, based on the fibroblastic RTG-2 fish cell line (ATCC, CCL N. 55). At exponential growth, three endpoints, beta-galactosidase activity, culture viability assayed by the neutral red assay (NR) and inhibition of growth rate using the FRAME KB protein assay (KBP), were used. By plotting the toxicity of each fraction vs elution time, the corresponding "toxicograms" were built. The UV and fluorescence chromatograms are compared to the three toxicograms (one for each toxicity endpoint).

Animal Testing Alternatives↗

Hazard evaluation of inorganics, singly and in mixtures, to flannelmouth sucker Catostomus latipinnis in the San Juan River, New Mexico.

Larval flannelmouth sucker (Catostomus latipinnis) were exposed to arsenate, boron, copper, molybdenum, selenate, selenite, uranium, vanadium, and zinc singly, and to five mixtures of five to nine inorganics. The exposures were conducted in reconstituted water representative of the San Juan River near Shiprock, New Mexico. The mixtures simulated environmental ratios reported for sites along the San Juan River (San Juan River backwater, Fruitland marsh, Hogback East Drain, Mancos River, and McElmo Creek). The rank order of the individual inorganics, from most to least toxic, was: copper > zinc > vanadium > selenite > selenate > arsenate > uranium > boron > molybdenum. All five mixtures exhibited additive toxicity to flannelmouth sucker. In a limited number of tests, 44-day-old and 13-day-old larvae exhibited no difference in sensitivity to three mixtures. Copper was the major toxic component in four mixtures (San Juan backwater, Hogback East Drain, Mancos River, and McElmo Creek), whereas zinc was the major toxic component in the Fruitland marsh mixture, which did not contain copper. The Hogback East Drain was the most toxic mixture tested. Comparison of 96-h LC50 values with reported environmental water concentrations from the San Juan River revealed low hazard ratios for arsenic, boron, molybdenum, selenate, selenite, uranium, and vanadium, moderate hazard ratios for zinc and the Fruitland marsh mixture, and high hazard ratios for copper at three sites and four environmental mixtures representing a San Juan backwater, Hogback East Drain, Mancos River, and McElmo Creek. The high hazard ratios suggest that inorganic contaminants could adversely affect larval flannelmouth sucker in the San Juan River at four sites receiving elevated inorganics.

Animals↗

Hazardous air pollutants and asthma.

Asthma has a high prevalence in the United States, and persons with asthma may be at added risk from the adverse effects of hazardous air pollutants (HAPs). Complex mixtures (fine particulate matter and tobacco smoke) have been associated with respiratory symptoms and hospital admissions for asthma. The toxic ingredients of these mixtures are HAPs, but whether ambient HAP exposures can induce asthma remains unclear. Certain HAPs are occupational asthmagens, whereas others may act as adjuncts during sensitization. HAPs may exacerbate asthma because, once sensitized, individuals can respond to remarkably low concentrations, and irritants lower the bronchoconstrictive threshold to respiratory antigens. Adverse responses after ambient exposures to complex mixtures often occur at concentrations below those producing effects in controlled human exposures to a single compound. In addition, certain HAPs that have been associated with asthma in occupational settings may interact with criteria pollutants in ambient air to exacerbate asthma. Based on these observations and past experience with 188 HAPs, a list of 19 compounds that could have the highest impact on the induction or exacerbation of asthma was developed. Nine additional compounds were identified that might exacerbate asthma based on their irritancy, respirability, or ability to react with biological macromolecules. Although the ambient levels of these 28 compounds are largely unknown, estimated exposures from emissions inventories and limited air monitoring suggest that aldehydes (especially acrolein and formaldehyde) and metals (especially nickel and chromium compounds) may have possible health risk indices sufficient for additional attention. Recommendations for research are presented regarding exposure monitoring and evaluation of biologic mechanisms controlling how these substances induce and exacerbate asthma.

Air Pollutants↗