Stack-collected coal fly ash from western low-sulfur coal was extracted with 60:40 benzene/methanol. This extract was fractionated by preparative-scale high performance liquid chromatography (HPLC) and the mutagenic activity of 14 fractions was evaluated by microbial assay with Salmonella typhimurium TA1538. A widespread distribution of direct-acting mutagens, which probably includes both mono- and dinitroaromatics, was detected. HPLC methods were also used to isolate 1-nitropyrene from the total benzene/methanol extract. The identification of 1-nitropyrene was based on gas chromatographic and HPLC retention measurements and mass spectral data. The concentration of 1-nitropyrene in the ash extract was determined by quantitative HPLC analyses. Mutagenicity assays of the total extract and an authentic 1-nitropyrene standard with Salmonella strains TA1538, TA100, and TA98 indicated that the 1-nitropyrene accounts for approximately 0.03-0.16% of the total mutagenic activity of the extract.