The production of mycotoxins and other secondary metabolites have been studied by LC-DAD-MS from six species in Aspergillus section Fumigati. This includes the three new species Aspergillus lentulus, A. novofumigatus and A. fumigatiaffinis as well as A. fumigatus, Neosartoria fisheri and N. pseudofisheri. A major finding was detection of gliotoxin from N. pseudofisheri, a species not previously reported to produce this mycotoxin. Gliotoxin was also detected from A. fumigatus together with fumagillin, fumigaclavine C, fumitremorgin C, fumiquinazolines, trypacidin, methyl-sulochrin, TR-2, verruculogen, helvolic acid and pyripyropenes. Major compounds from A. lentulus were cyclopiazonic acid, terrein, neosartorin, auranthine and pyripyropenes A, E and O. Thus in the present study A. fumigatus and A. lentulus did not produce any of the same metabolites except for pyripyropenes. The fact that A. lentulus apparently does not produce gliotoxin supports the idea that other compounds than gliotoxin might play an important role in the effective invasiveness of A. lentulus. An overall comparison of secondary metabolite production by strains of the six species was achieved by analysis of fungal extracts by direct injection mass spectrometry and cluster analysis. Separate groupings were seen for all the six species even though only one isolate was included in this study for the two species A. novofumigatus and A. fumigatiaffinis.