Volatile metabolites that might accompany production of PR toxin by Penicillium roqueforti were investigated. Volatiles and PR toxin were evaluated for 16 strains of P. roqueforti. Solid phase microextraction was used for isolation of volatiles. Thirteen strains produced PR toxin, and all of them produced a specific set of sesquiterpene hydrocarbons including (+)-aristolochene-an intermediate in PR toxin biosynthesis, beta-bisabolene, alpha-chamigrene, diepi-alpha-cedrene, beta-elemene isomer, beta-elemene, beta-gurjunene, beta-himachalene, alpha-panasinsene, beta-patchoulene, beta-patchoulene isomer, alpha-selinene, and valencene. Aristolochene and the remainder of the sesquiterpene hydrocarbon profile were unique for P. roqueforti producing PR toxin. They were absent in nontoxigenic P. roqueforti and in 40 strains of other Penicillium species. Volatile compounds, sesquiterpene hydrocarbons, and aristolochene paralleled PR toxin synthesis. Incubation temperature (20, 24, or 27 degrees C) and water content in the medium (20, 30, or 40%) influenced the amount of produced sesquiterpenes, but not their profile, suggesting it is species specific. The sesquiterpene hydrocarbon pattern and especially aristolochene can be used as volatile markers for detecting the process of undergoing biosynthesis of PR toxin by P. roqueforti.