Eva Agurell

Results of the IPCS collaborative study on complex mixtures.

The International Programme on Chemical Safety (IPCS) sponsored a collaborative study to examine the intra- and inter-laboratory variation associated with the preparation and bioassay of complex chemical mixtures. The mixtures selected were National Institute of Standards and Technology (NIST) Standard Reference Materials (SRMs). 20 laboratories worldwide participated in the collaborative trial. The participating laboratories extracted the organic portion of two particulate samples--an air-particulate sample and a diesel-particulate sample--and bioassayed the extracts. The laboratories simultaneously bioassayed a NIST-prepared extract of coal tar and two control compounds (benzo[a]pyrene, and 1-nitropyrene). The bioassay method used was the Salmonella/mammalian microsome plate-incorporation test using strains TA98 and TA100. Study design also allowed for a comparison of sonication and Soxhlet extraction techniques. The mean extractable masses for the air particles and diesel particles were approximately 5% and 17.5%, respectively. The particulate samples were mutagenic in both strains with and without activation in all 20 laboratories. For TA100 the with and without activation slope values for the air particulate were 162 and 137 revertants per mg particles, respectively. For TA98 the respective diesel slope values were 268 and 269. The mutagenicity slope values for the diesel particles ranged from 3090 (TA98, +S9) to 6697 (TA100, +S9) revertants per mg particles. The coal tar solution was negative for both strains when exogenous activation was not used but was mutagenic in both strains with exogenous activation. The benzo[a]pyrene and 1-nitropyrene were used as positive controls and gave results consistent with the literature. This paper provides a complete summary of the data collected during the collaborative study. Companion papers provide further analysis and interpretation of the results.

Salmonella/microsome mutagenicity of monochloro derivatives of some di-, tri- and tetracyclic aromatic hydrocarbons.

A series of 8 monochloroarenes have been tested for mutagenicity in the Salmonella/microsome assay. None of the compounds was detectably active in the absence of mammalian activation whereas, depending on structure, some of the compounds were mutagenic in its presence having responses higher than those reported for the parent compounds.

Public exposure to environmental tobacco smoke.

Airborne particulate matter has been collected by personal samplers in public indoor areas and travel situations with environmental tobacco smoke pollution. Following extraction, the samples were assayed for mutagenicity in the presence of S9 with a sensitive microsuspension test using Salmonella TA98. The mutagenic responses of indoor air from public areas were much higher than those of ambient outdoor air. Depending on the circumstances, the mutagenic response varied in trains and airplanes but the results show that physical separation of non-smoking sections from smoking sections is necessary in order to achieve genuine non-smoking areas. Chemical fractionation and mutagenicity assay of the basic fraction show that Salmonella mutagenicity of airborne particulate matter might be used as a tobacco smoke-specific indicator, as the basic fraction of environmental tobacco smoke contains a large part of the mutagenic activity whereas this is not the case for outdoor ambient airborne particulate matter.

Salmonella mutagenicity of three complex mixtures assayed with the microsuspension technique. A WHO/IPCS/CSCM study

In a collaborative study on complex mixtures, three complex mixtures and two pure compounds were assayed with the Salmonella microsuspension technique. The two pure compounds were benzo[a]pyrene (BaP) and 1-nitropyrene (1-NP). The three complex mixtures were standard reference materials (SRMs) from the U.S. National Institute of Standards and Technology, SRM 1649, SRM 1650 and SRM 1597. The two samples SRM 1649, an urban dust particulate matter, and SRM 1650, a diesel particulate matter, were sonicated with dichloromethane. Sample SRM 1597 was an extract of a coal tar sample with a complex mixture of polycyclic aromatic hydrocarbons. The microsuspension assay was performed with Salmonella strains TA98 and TA100 according to Kado et al. (1983) with minor modifications (Löfroth et al., 1988). The results showed that the microsuspension technique is a more sensitive assay than the plate incorporation method. Depending on sample, strain and metabolic condition the mutagenic responses were 3-37 times higher in the microsuspension assay than in the conventional plate incorporation assay. The microsuspension method is thus useful for environmental samples which are often available in only small amounts.

The use of the Salmonella/microsome mutagenicity test for the characterization of organic extracts from ambient particulate matter

Abstract Samples of ambient particulate matter, which have been collected for a collborative research project by electrostatic precipitation and high volume sampling on glass fiber filters, have been extracted and the organic extracts have been assayed for mutagenicity in the Salmonella /microsome test. Among several solvents tested, acetone gives the highest extraction efficiency. A comparison between different sampling sites, traffic tunnel, urban street, urban rooftop, and a rural location during episodes of long-range aerosol transport, indicates that there are differences with respect to the mutagenic response per particle mass and amount of selected PAH. Tests in the absence and presence of mammalian metabolic activation, tests with enzyme deficient strains, and tests with fractionated samples all indicate that a large part of the recovered mutagenic activity is caused by relatively polar compounds containing nitro groups that can be activated by bacterial enzymes. Among several tested strains, only TA104, which responds to oxidative mutagens, gives a higher response than the conventional tester strains TA98 and TA100.

Amplification and loss of repeat units of the human minisatellite MS1 integrated in chromosome III of a haploid yeast strain

Minisatellites comprise arrays of tandemly repeated short DNA sequences which show extensive variation in repeat unit number. The mechanisms that underlie this length variation are not understood. In order to study processes influencing length changes of minisatellites, we integrated the human minisatellite MS1 into a haploid strain of the yeast Saccharomyces cerevisiae. Frequent spontaneous generation of MS1 alleles with new lengths were observed in this yeast strain. Hence it is concluded that recombination between members of a pair of homologous chromosomes is not a prerequisite for the generation of length changes in MS1 in yeast.

Salmonella/microsome mutagenicity of 1-nitropyrene-2-ol, a nitropyrene phenol formed in the photolysis of 1-nitropyrene

Abstract 1-Nitropyrene-2-ol is not detectably mutagenic in the Salmonella test in the absence of mammalian activation. It is activated by the rat liver microsom e-containing preparation S9 and gives a mutagenic response similar to that of benzo(a)pyrene.

Detection of DNA alterations in human bladder tumors by DNA fingerprint analyses.

DNA fingerprint analyses were used to examine the constitutional and tumor DNA from 22 bladder tumor patients. DNA alterations, such as loss of bands, new bands, and intensity shifts were observed in 10 of the 22 patients. The most frequent DNA alteration, occurring in 80% of the patients, was a complete loss of one or several bands. Fingerprint abnormalities were present both in low-malignant superficial tumors and in high-malignant invasive tumors, but were also lacking in the latter group. Apparently no relationship exists between fingerprint abnormalities and gross chromosomal aberrations or the proportion of S-phase cells as measured by flow cytometry or development of recurrent tumors during a limited observation period. Thus, whether fingerprint aberrations express genetic alterations directly involved in the malignancy potential of bladder carcinoma remains an open question.

Presence of Various Types of Mutagenic Impurities in Carbon Black Detected by the Salmonella Assay

Aside from their use as rubber additives, carbon blacks are used as colorants in many contexts, such as photocopying, typewriting, carbon copying, and printing. The ultimate products are commonly in close proximity to man and are often worn or turned into waste, further enhancing the dissemination of their ingredients.

Mutagenicity of extracts from typewriter ribbons and related items.

Extracts of typewriter ribbons and carbon papers were found to be mutagenic in the Salmonella/microsome assay with strain TA98. Fractionation of ribbon extracts indicates that at least 2-3 different classes of mutagenic component are present in these extracts. Nitro-containing compounds may be responsible for the high mutagenicity observed for some of the ribbon extracts in the absence of S9. The results indicate that impurities in the products may be causing part of the mutagenic effect.