Mutagenicity and chemical analysis of sequential organic extracts of airborne particulates

To obtain insight into the identity of chemicals associated with the mutagenicity of United States National Institute of Standards and Technology (NIST) Standard Reference Materials SRM 1649 (urban dust) and SRM 1650 (diesel particulate), parallel mutagenicity tests and chemical analyses were performed on dichloromethane and sequential organic extracts of these samples. SRM 1649 and 1650 were sequentially extracted with five organic solvents of increasing polarity, in order to partition mutagenic components into discrete fractions. The solvents (with associated polarity index) were as follows: (1) hexane (0.0); (2) hexane:diethyl ether 9:1 (0.29); (3) hexane:diethyl ether 1:1 (1.45); (4) diethyl ether (2.9); (5) methanol (6.6). 0.9270 g of SRM 1649, and 0.0510 g of SRM 1650 were each extracted three times with 8 ml of each of the solvents, the three aliquots were pooled, and analysed for target organics or solvent-exchanged into DMSO for mutagenicity testing in Salmonella typhimurium strains TA98 and TA100.The dichloromethane extracts of SRM 1649 and SRM 1650 contained direct-actin mutagens in Salmonella strains TA98 and TA100; SRM 1650 was significantly more potent than SRM 1649 in either strain. Addition of S9 caused a large decrease in mutagenicity of each extract, although SRM 1650 remained more potent. An interesting pattern of mutagenicity was observed for the sequential extracts of SRM 1649 and SRM 1650: the mutagenic potency of SRM 1649 extracts increased with increasing polarity of the extraction solvent while the response of the SRM 1650 extracts was the opposite. This suggests that the direct-acting mutagens in SRM 1650 are unlike those in SRM 1649. The response, though diminished, was largely unchanged when S9 was included in the test mixture.Chemical analyses on the various extracts were performed using a Hewlett-Packard model 5890 gas chromatograph equipped with a model 5970B mass selective detector (GC-MSD), and a 0.3 μm film thickness cross-linked methyl silicone capillary column (HP 1909A-101). Selected ion monitoring (SIM) methods were used to analyze for 105 target compounds including PAHs and nitro-PAHs. Chemical analysis of the dichloromethane extracts of SRM 1649 and SRM 1650 identified three main classes of compounds: polyaromatic hydrocarbons (PAH), vitro-polyaromatic hydrocarbons (NO₂-PAHs) and heterocyclics. The concentration of target compounds and the proportion of vitro-PAHs and heterocyclic compounds were considerably greater in SRM 1650 than in SRM 1649, consistent with the observed differences in their mutagenic potency. However, the different responses of the dichloromethane extracts in TA98 and TA100 suggest the presence of different (unidentified) compounds.Many of the target compounds were detected at least once in the sequential extracts from SRM 1649 and SRM 1650. There was no evident relationship between the occurrence of extracted organics, or classes of organics, and the polarity of solvents, except that, generally, the largest amount and variety of compounds were recovered in the first and second extracts (hexane; hexane:diethyl ether, 9:1). Preliminary examination of the chemical analysis results did not provide an explanation of the observed trends in mutagenic response. No single class of chemicals or individual compound was found to account for the observed pattern of mutagenicity. Compounds other than those identified must also contribute to the observed mutagenicity of any of the SRM 1649 and SRM 1650 extracts.