Andrew Sivak

Skin carcinogenicity of condensed asphalt roofing fumes and their fractions following dermal application to mice

Condensed roofing asphalt fumes, generated at 316 degrees C, were collected by cold trap condensation and fractionated by preparative high performance liquid chromatography. Chemical classes in each of the fractions (A-E) were identified by gas chromatography/mass spectroscopy. The fractions, various combinations of fractions, the raw and heated asphalt, the neat asphalt fume and the reconstituted asphalt were tested for carcinogenicity, and three fractions were tested for cocarcinogenicity and tumor promotion with benzo[a]pyrene (BaP). The skin application carcinogenesis bioassay was conducted by twice weekly application of test materials in 0.05 ml of acetone/cyclohexane (1:1) for 104 weeks to 40 groups of male C3H/HeJ mice (30/group). Fractions were applied at a mass in proportion to their amount in the neat asphalt fumes. In addition, the neat asphalt fume was tested on Sencar mice to determine if this strain was more susceptible to the carcinogenic effects of the fumes. Condensed neat asphalt fumes produced similar and statistically significant increased tumor yields of papillomas and carcinomas in both strains as compared to respective vehicle controls. Recombination of all fractions resulted in a tumor response similar to neat asphalt fumes. Among individual fractions, C was most potent, followed by B. The other single fractions were without significant tumorigenic activity. Combinations containing fractions B and C were most active among the mixtures that were assayed and no evidence of enhancement of tumorigenesis in the mixtures was found. No significant cocarcinogenic or tumor promoting activity was observed with fractions A, D, or E and BaP. Raw unheated asphalt produced a few tumors in C3H mice, but no tumors were seen when raw asphalt heated to 316 degrees C, with the fumes permitted to escape, was applied.

Rat Liver Foci and in Vitro Assays to Detect Initiating and Promoting Effects of Chlorinated Ethanes and Ethylenes

Nine chlorinated aliphatics (CAs) were examined in a rat liver foci assay for tumor initiating and promoting activities. In this model, young adult male Osborne Mendel rats were first subjected to a partial hepatectomy, the test chemical was then administered at the maximum tolerated dose in the initiation or promotion phase in conjunction with diethylnitrosamine (DEN; 30 mg/kg b.w.) or phenobarbital (PB; 0.05 percent, w/w, in the diet), and gamma glutamyltranspeptidase (GGT) was used as a putative preneoplastic indicator. When administered in the promotion protocol after initiation with DEN, 1,1-dichloroethane, 1,1,2-trichloroethane (1,1,2-TCE), 1,1,2,2-tetrachloroethane (1,1,2,2-TTCE), tetrachloroethylene (TTCY), and hexachloroethane induced significant increases in GGT+-foci above control levels. 1,1,2,2-TTCE, TTCY, and 1,1,2-TCE also induced significant increases in GGT+-foci when administered in the promotion protocol without DEN initiation. Two variants of GGT+-foci were observed: the classical type associated with PB promotion, and the other, which was more diffuse, less intensely stained, resembling foci undergoing redifferentiation and associated with CAs. A number of CAs were also genotoxic in short-term in vitro tests. Taken together, the studies suggest that CAs may be complete carcinogens in vivo with weak initiating activity and stronger promoting activity.

An interlaboratory evaluation of the Syrian hamster embryo cell transformation assay using eighteen coded chemicals

Eighteen coded chemicals were evaluated in the Syrian hamster embryo (SHE) cell transformation assay in three different laboratories using the same basic experimental protocol with minor modifications. In addition, individual cell and serum sources were selected. Major factors influencing intra-and interlaboratory reproducibility were the source of cells and serum, the toxicity of the chemicals, and the dose-range selected for transformation evaluation. Two or three assays from each laboratory were required to determine the transformation-inducing potential of a chemical because of the low number of transformants scored in any single assay and the difficulty of interpreting morphological variations. Rodent carcinogenicity data were available for 16 of the 18 chemicals tested and the transformation response of 14 of those chemicals was in agreement with the rodent carcinogenicity data (if the positive results are adopted for the four chemicals that produced contradictory results). Four rodent carcinogens, di-(2-ethylhexyl) phthalate, diphenylhydantoin, methapyrilene hydrochloride and o-toluidine hydrochloride, that were negative in the Salmonella/microsome assay, induced morphological transformation in the SHE assay. Although the labour, cost and lack of reproducibility might preclude application of this transformation assay for routine screening, it might, nevertheless, prove valuable for distinguishing between non-mutagenic carcinogens and non-carcinogens.

In vitro transformation of BALB/c-3T3 cells by chlorinated ethanes and ethylenes.

Eight chlorinated ethanes and 3 chlorinated ethylenes were tested in the BALB/c-3T3 cell transformation assay. Under the conditions of the assay, vinyl chloride and 1,1,1-trichloroethane induced a clear positive transformation response while 1,1,2-trichloroethane and trichloroethylene were weakly positive. Chloroethane, 1,1- and 1,2-dichloroethane, 1,1,1,2- and 1,1,2,2-tetrachloroethane, hexachloroethane and tetrachloroethylene were all negative in the assay conducted in the absence of an exogenous metabolic activation system. These results suggest that the BALB/c-3T3 cells possess capability to activate some, but not all, of the chlorinated hydrocarbons which exhibit species specificity in producing carcinogenicity in mice but not in rats.

Cell culture tumor promotion experiments with saccharin, phorbol myristate acetate and several common food materials

The BALB/c-3T3 cell neoplastic transformation system was modified to examine the tumor promoting activity of a set of substances. Following initiation of the target cells with 3-methylcholanthrene, treatment of the cultures with phorbol myristate acetate (0.01 microgram/ml; 1.5 X 10(-8) M) during the remainder of the 4-week assay interval resulted in a marked increase in both spontaneous and initiated Type III transformed foci. In contrast, a similar treatment with saccharin at 20, 100 or 500 microgram/ml (0.08, 0.4 or 2.1 X 10(-3) M) did not influence the occurrence of Type III transformed foci and did not result in a promoting response. Sodium ascorbate (2.53 X 10(-3) M) and L-tryptophan (2.45 X 10(-3) M) almost completely inhibited both spontaneous and initiated Type III transformed foci. Calcium pantothenate (2.10 X 10(-3) M) exhibited a marginal promoting effect. Under the conditions of this study in which the classical tumor promoter phorbol myristate acetate was highly active in promoting Type III transformed foci, saccharin was not active as either a direct transforming or promoting agent at doses up to 5 orders of magnitude higher.

Environmental and Human Health Aspects of Commercially Important Surfactants

Seven types of surfactants comprise the majority of those presently used in commercial detergent formulations. These are linear alkylbenzene sulfonates (LAS), alkyl sulfates (AS), alcohol ethoxylates (AE), alkyl phenol ethoxylates (APE), alcohol ethoxy sulfates (AES), alpha olefin sulfonates (AOS) and secondary alkane sulfonates (SAS). LAS surfactants, the mainstay of detergent components, have been in use the longest and their paths of biodegradation are relatively well understood. Environmental levels of methylene blue active substances, the most commonly employed but non-specific measure of anionic surfactant concentration, indicate that LAS are readily biodegradable. Nonionic (AE and APE) and the anionic AES and AS surfactants are also biodegradable with APE degrading somewhat more slowly than the others. Acute toxic effects to aquatic life forms generally occur in adult vertebrates and invertebrates at surfactant concentrations from 1 to 20 mg/L; juvenile and developmental stages show effects at somewhat lower concentrations.

Neoplastic Transformation Systems — Their Use in Studying Carcinogenesis

The cellular systems to study neoplasia essentially stem from two sources. One is the observation that one can induce alterations in cellular phenotype in a culture of cells infected with tumorigenic DNA viruses (1). The second is the finding of Berwald and Sachs in 1963 (2,3) that early passage Syrian hamster embryo cells exhibited clonal morphology not seen in untreated cultures following exposure to a chemical carcinogen. While the work of Earle and his associates (4) beginning in the nineteen thirties had demonstrated changes in cell cultures treated with carcinogens, it was the protocol and results reported in 1963 (2,3) that provided a means to obtain quantitative and reproducible findings of morphological transformation of mammalian cells induced by chemical carcinogens. Over the past two decades a considerable variety of systems have been described to study neo-plastic transformation. Of these, several have been shown to have value as bioassays for the identification of carcinogens. The properties as well as the advantages and disadvantages of these assays have been reviewed in depth recently along with a presentation of the available data base on tested chemicals (5–9). Table 1 lists these assay types along with some basic characteristics.