International Journal of Toxicology

This paper provides a general introduction to the occurrence, epidemiology, and toxicity of some of the most common contaminants of water supplies, the volatile organic chemicals (VOCs). The VOCs are formed from the reaction of chlorine during disinfection with naturally occurring carbon in the form of humic acids. The VOCs may also enter water supplies as a result of manufacturing, processing, distribution, and urban and agricultural run off. Their occurrence is summarized in this paper.

No epidemiologic studies examine the health effects where skin is the sole route of exposure. However, in several studies skin is one of the routes of exposure for VOCs. These are summarized in this paper.

Finally, the toxicity of some of the more important VOCs is summarized. Where possible, similarities in toxicity between individual members of this class of chemical contaminants are noted. There are striking similarities of toxicity of various VOCs in the liver, kidney, and hematopoietic system. These similarities should be considered as skin exposure models are being developed.

The combination of psoralens and ultraviolet light (UVA, 320–400 nm), also referred to as PUVA, is a potent modulator of epidermal cell growth and differentiation. Although it has been postulated that PUVA exerts its actions by binding to DNA, our laboratory has obtained evidence that a specific, saturable, high-affinity receptor site independent of the DNA mediates the biologic actions of these drugs. This receptor is a 22,000 molecular weight protein present in membrane and cytoplasmic fractions of responsive cell types. Treatment of cells with psoralens followed by UV light causes activation of the receptor. This leads to specific cell surface membrane alterations, in particular, phosphorylation of the receptor for epidermal growth factor (EGF). The EGF receptor is a transmembrane glycoprotein possessing intrinsic tyrosine kinase activity. Modification of the EGF receptor leads to a loss in its ability to bind EGF, as well as an inhibition of its tyrosine kinase activity. These data indicate that the psoralens act at the level of the cell membrane and that their biologic effects in the skin may be mediated by the ability of these drugs to disrupt normal growth factor functions.

Assessment of risks to humans associated with the use of chemicals requires knowledge of the hazard (toxicity) of the chemical and level of human exposure. Hazard assessment is often based on animal bioassays and quantitative exposure estimates of dermal exposure obtained from studies monitoring workers. Because human skin is an effective barrier to many chemicals, it cannot be assumed that the deposited dose is equivalent to the systemic dose. However, an estimate of systemic dose may be derived by multiplying the deposited dose by the percentage of percutaneous uptake. This correction can have major impact on the regulatory decision, because the adjusted dose used in the risk calculation may be reduced significantly, especially at high doses, when the uptake is not linearly proportional to the exposure. It is therefore important that the dermal absorption value be accurate. As outlined in this paper, numerous factors can affect percutaneous absorption. Nevertheless, many regulatory agencies will consider the use of percutaneous absorption data derived from in vivo studies to adjust the dermally deposited dose to that delivered systemically. Numerous issues must be resolved before in vitro dermal penetration studies can be used for risk assessment.

The Environmental Protection Agency has circulated a protocol for examining the dermal absorption of pesticides in rats. This protocol will be considered as a guideline for determining the dermal absorption of pesticides. Approximately 40 pesticides have been evaluated with this protocol. Male rats are dosed dermally with labeled pesticide. Doses, in mg/cm², are applied to the shaven skin of the back as the use product, diluted with water if necessary. The application site is protected with a nonocclusive device. Four rats per dose are exposed for 0.5, 1, 2, 4, 10, or 24 hrs. Samples collected are soap and water wash, skin at the application site, blood, total urine and feces, carcass, and selected tissues. Mass balance calculations include determination of pesticide that can be removed with soap and water, pesticide bound on or in the skin, total pesticide absorbed with time, blood concentrations with time, pesticide accumulation in target tissues, and pesticide excreted.

In vivo dermal penetration studies on rats have been investigated at Ciba-Geigy since 1981. Eleven different pesticides have been evaluated for dermal absorption during this time. Selected compounds with varying degrees of dermal absorption are discussed. An overview of the test protocol and nonocclusive appliance is shown. Data are presented to help evaluate whether bound skin residues after washing are permanently sequestered in skin.

Contaminants exist in ground and surface water. Human skin has the capacity to bind and then absorb these contaminants into the body during swimming and bathing. Powdered human stratum corneum will bind both lipid-soluble (alachlor, polychlorinated biphenyls [PCBs], benzene) and water-soluble (nitroaniline) chemicals. In vitro (human skin) and in vivo (Rhesus monkey) studies show that these chemicals readily distribute into skin, and then some of the chemical is absorbed into the body. Linearity in binding and absorption exists for nitroaniline over a 10-fold concentration range. Multiple exposure to benzene is at least cumulative. Binding and absorption can be significant for exposures as short as 30 min, and will increase with time. Absorption with water dilution increased for alachlor, but not for dinoseb. Soap reversed the partitioning of alachlor between human stratum corneum and water. The PCBs could be removed from skin by soap and water (70% efficiency) for up to 3 h and then decontamination potential decreased, due to continuing skin absorption. The model in vitro and in vivo systems used should permit easy estimation of this area of extensive human exposure effect on risk assessment.

Methods have been developed for the detection of exposure to carcinogens and other DNA-damaging agents in experimental animals and humans, through the detection of carcinogens or metabolic derivatives of them in body fluids or adducts bound covalently to DNA or hemoglobin. These methods are being applied in studies of exposure to environmental carcinogens, the results of which demonstrate their adequacy for detecting ambient exposures. The successful use of urinary markers of genotoxic exposures has been reported with respect to nitrosoproline as an indicator of exposure to N-nitroso compounds. The same approach has been used to detect aflatoxin B₁ (AFB₁) metabolites and AFB₁-N⁷-Gua as markers of exposure to aflatoxin B₁. Detection of adducts formed between genotoxic agents and hemoglobin has been reported in studies of populations occupationally exposed to ethylene oxide, in which 3-hydroxyhistidine and 3-hydroxyva-line have been measured, and in smokers, whose hemoglobin has been found to contain levels of 4-aminobiphenyl and 3-hydroxyvaline that were correlated with the frequency of cigarette smoking. Albumin adducts of AFB₁ have been identified in exposed people and their levels correlated with ingested amounts of the carcinogen.

DNA adducts of genotoxic agents have also been detected in the cells and tissues of exposed individuals. Several studies to date have focused on exposure to the ubiquitous polycyclic aromatic hydrocarbon benzo(a)pyrene. Immunoassays and physicochemical methods have been used to detect adducts formed through the major intermediate in the activation pathway, the benzpyrene-7,8-diol-9,10-epoxide (BPDE). BPDE adducts have been found in the DNA of peripheral leukocytes of workers in foundries, aluminum manufacturing plants, and coke oven plants, and also in roofers and cigarette smokers with the use of synchronous scanning fluorescence as well as by enzyme-linked immunosorbent assays (ELISA) or ultrasensitive enzyme radioimmunoassays (USERIA). DNA adducts of O⁶-methyl guanine have also been detected by immunoassay in the blood of populations at high risk for esophageal cancer. The method of ³²P postlabeling has been used for the detection of DNA adducts in placentas, peripheral leukocytes, and oral mucosal cells of tobacco smokers as well as coke oven and foundry workers, and increased total levels of adducts were in general indicative of elevated levels of exposure.

This paper is summarized from a report prepared by the Houston Regional Monitoring Corporation for submission to the U.S. Environmental Protection Agency explaining the methodology used in the East Harris County and West Chambers County for assessing community exposure to volatile compounds. The paper briefly discusses the program description, the rationale for selecting candidate airborne chemicals for detailed health effects assessments, and the method used to develop consensus guidance values based on accepted risk assessment methodologies. The results of a 6-month monitoring period are presented with health assessment indicators.

Both nuclease PI treatment and reverse-phase high-performance liquid chromatography (HPLC) were used to enrich hydrophobic/bulky DNA adducts in DNA digests. ³²P-postlabeling procedures and thin layer chromatography were then used to detect and quantitate aromatic/bulky DNA adducts. For both human and fish DNA from individuals exposed to environmental carcinogens, the nuclease PI and HPLC enrichment procedures generally gave similar results. The bottom sediments of the Buffalo and Detroit rivers are contaminated with polycyclic aromatic hydrocarbon carcinogens, and brown bullheads in these rivers show a high rate of liver cancer. Compared to DNA from control fish raised in aquariums, DNA of livers of brown bullheads from the polluted rivers exhibited elevated levels of DNA adducts. DNA from human oral mucosal cells and lymphocytes exhibited DNA adducts, but adducts levels did not differ significantly in smokers and nonsmokers. Adduct levels in DNA from human lung biopsy tissue, however, were elevated in smokers compared to nonsmokers. In former smokers, adducts levels were highest in those who had recently quit, and lowest in those who had not smoked for 10 years or more. Measurement of DNA adducts by ³²P-postlabeling appears to be a useful and particularly direct procedure for assessing genetic damage from environmental carcinogens.

A major concern of molecular epidemiology is the identification of individuals at increased risk of cancer by obtaining evidence of high exposure to carcinogens that may lead to pathobiological lesions in target cells. DNA is considered to be a target for modification by mutagens and carcinogens; therefore, damage to DNA can be used as an internal, molecular dosimeter of carcinogen exposure. The reactive species of these carcinogens may bind either directly to DNA to form adducts or indirectly to cause secondary DNA lesions through free radicals and aldehydes. Highly sensitive and specific methods have been developed to measure DNA lesions and DNA repair products that are found in biological specimens from humans exposed to carcinogens in the environment. For example, DNA adducts have been measured in cells and tissues from people exposed environmentally to carcinogenic polycyclic aromatic hydrocarbons or alkylating agents. Antibodies recognizing carcinogen-DNA adducts have also been detected in human sera. Carcinogen-protein adducts are also being used as molecular dosimeters of carcinogen exposure. The advantages and limitations of the various methods used to measure carcinogen-macromolecular adducts are discussed here. The use of two or more complementary assays to obtain confirmatory results is recommended.

This report presents the risk-assessment-related aspects of a multidisciplinary study of indoor coal smoke pollution and lung cancer in Xuan Wei County, Yunnan Province, China. Xuan Wei presents a unique natural experiment in environmental carcinogenesis because lung cancer mortality rates and indoor pollution exposures vary widely within the County. Current evidence links lung cancer with domestic burning of “smoky coal,” as opposed to “smokeless coal” and wood. Efforts to determine the most carcinogenic components of smoky coal pollution are in progress, as are efforts to develop a quantitative relationship of pollution dose with lung cancer response in Xuan Wei. Some available evidence suggests that the composition of indoor pollution does not vary greatly throughout Xuan Wei, and thus that lung cancer risk is a function of overall pollution exposure. Other evidence suggests that different Xuan Wei fuels exhibit different carcinogenic potencies. On-site and laboratory studies are being conducted to differentiate between these possibilities.

The Agency for Toxic Substances (ATSDR) was mandated by Superfund legislation to create two national registries: a registry of persons exposed to hazardous substances and a registry of persons with serious diseases or illness. The polices and procedures for creating the first of these (The National Exposure Registry) have been established and are discussed here. The decision logic for site selection, the criteria for population selection, the data collection and storage, and related activities are detailed.

There are some similarities and differences in the process of sexual differentiation during development in rodents and primates. The most obvious difference is in the timing of the critical periods for morphogenesis of the reproductive tract and sexual differentiation of the central nervous system (CNS). In primates these events occur late in the first trimester while in rodents they occur in the perinatal period.

The gonadal hormones involved in morphogenesis of the male reproductive tract are identical for all mammalian species studied. Testosterone is the androgen that induces differentiation of the seminal vesicles, vas deferens, and epididymis, while dihydrotestosterone (DHT) promotes differentiation of the external genitalia, urethra, and prostate.

Estrogens, derived from aromatization of testosterone, are the proximal determinants of male sexual differentiation of the CNS in rodents. In nonhuman primates, however, a nonaromatizable androgen, DHT, produces the same effect as testosterone on sexually differentiated behaviors in female offspring. Studies of male patients with 5α-reductase deficiency have suggested that maturation of male gender identity and psychosexual behavior in humans is critically dependent on testosterone but not on normal levels of DHT in prenatal and prepubertal life. Gender identity does not appear to be unalterably fixed in humans until the time of puberty and even at this and later times environmental factors have a strong impact.

Benzene is widely used by chemical industries and exposure to benzene has been shown experimentally to be immunotoxic in adult animals. This study addressed whether exposure of fetuses in utero to benzene compromises the development of fetal B lymphopoiesis and whether B-lymphocyte development recovers postnatally. Pregnant BALB/C dams were given intraperitoneal injections of benzene (100 mg/kg, twice daily) from day 12.5 of gestation through day 19.5 of gestation. Phenotypic analysis revealed that fetal liver cell suspensions from embryos exposed in utero contained fewer pre-B cells and B cells than corresponding controls. Fetal liver cell cultures established from these embryos also produced fewer B cells. In contrast, levels of pre-B cells were elevated in the livers of 8-day-old neonates that had been exposed to benzene in utero. Moreover, responsiveness to the B-cell mitogen, LPS, was significantly decreased in spleen cell cultures derived from these neonates. These results indicate that in utero exposure to high concentrations of benzene alters fetal B lymphopoiesis and may compromise immune responsiveness postnatally.

Anencephaly and spina bifida are the two most common neural tube defects (NTDs) that occur in humans. They cause considerable fetal wastage and neonatal morbidity and mortality. Numerous animal models have been discovered and have been used to study these malformations. We used the scanning electron microscope (SEM) to compare the development of embryonic stages of spina bifida and anencephaly. Pregnant 180 g Sprague-Dawley rats were given either a 1 ml subcutaneous injection of 1% trypan blue on gestational days 7-8 or an intragastric administration of 75,000 units of vitamin A on gestational days 8-10 to produce embryos with spina bifida or anencephaly. Controls were given vehicle only. The SEM examination of day 9 and 10 embryos revealed no morphologic differences between controls and subjects. Subsequent gestational days showed closure of neural tubes in controls but progressive opening of neural tubes (in the rostral and caudal regions) in experimental subjects. Growth of the dysmorphic neural tube region with subsequent sponteneous necrosis late in gestation resulted in the mature malformations of anencephaly and spina bifida. This study emphasizes the similarities in the developmental stages of spina bifida and anencephaly. We also surveyed a large series of human anencephalic autopsy specimens and noted striking similarities to the animal model.

This paper compiles sources of information dealing with environmental toxicology. Special emphasis has been put on information available from various computer databases and support networks. The information compiled will be useful for professionals entering the regulatory agencies as toxicologists or environmental health scientists.

The subchronic dermal toxicity of aqueous solutions containing the diethanolamine salt of para-tertiary butyl benzoic acid was determined in Fischer 344 rats. Para-tertiary butyl benzoic acid (pt BBA) has important applications in the manufacture of resins, polymers, and corrosion inhibitors. Male and female rats were exposed topically 5 days/week for up to 13 weeks with dosing solutions that resulted in daily exposures of 0.0, 17.5, 35.0, 70.0, or 140.0 mg/kg pt BBA. These concentrations did not produce overt clinical signs of toxicity and did not cause irritation to dermal exposure sites. However, exposure to the two highest concentrations resulted in decreased weight gain. Exposure at all concentrations resulted in dosage-related increases in relative hepatic and renal weights. Exposure of males to the two highest concentrations caused decreased testis weight. Exposure-related pathologic changes were confined to three organ systems: cytoplasmic vacuolation in the liver; tubular dilation and papillary necrosis of the kidneys; and tubular degeneration in the testes. Accompanying aberations in clinical chemistry values suggested altered hepatic and renal function. In males exposed daily to 70.0 and 140.0 mg/kg pt BBA, the testicular effects were marked, but no effects were detected in rats exposed to 17.5 or 35.0 mg/kg of pt BBA.

T-cell-dependent and t-independent humoral responses as well as cell-mediated re-sponses⁽¹⁾ were assessed with groups of young, Hormel-Hanford miniature swine. Test antigens established in various immunotoxicity studies with rodents were used. Results with the widely used t-dependent test antigen, sheep red blood cells (SRBC), are presented in this report. Groups of swine were immunized with varying dosages of SRBC to evaluate basal responses. Serum antibody titers were estimated by the standardized hemolysin (HLY) visual endpoint (EP) assay and a quantitative spectro-photometric (SP) variation using highly purified immunoglobulin (Ig) M and Ig G standards. The sensitivity of the EP HLY assay is about 0.5-1.0 μg for Ig G anti-SRBC; the SP variation is 10 times more sensitive. Peak responses were obtained 7 days after either the first or second injection for both males and females. Peak responses for females, however, were two to four times greater than those in males. Ig M antibodies had a three times higher specific HLY titer than those in the Ig G class. Use of purified antibody standards in either assay variation permitted conversion of dilution titers to specific amounts of antibody. The latter is a more meaningful parameter for various comparisons in either inter-or intraspecies evaluations in immunotoxicity studies.

To evaluate SRBC as a test antigen, low and intermediate doses of the immunotoxicant, cyclophosphamide, were given i.p. starting 2 days before SRBC injection. Antibody levels were reduced correspondingly to approximately 25 and 60% immunosuppression for the two dose levels in both primary and secondary responses. After 2 weeks of rest, animals recovered from treatment with CP. These studies demonstrate that SRBC immunization can be used in the FDA, Hormel-Hanford miniature swine to evaluate the immunotoxicity potential of chemicals without adjuvants or apparent harmful effects to the animal.