Twelve isolates ofFusarium graminearum were obtained from barley grains collected from different Uruguayan regions (harvest 1993–94). This was the predominant fungal species contaminating the crop due to a particular humid and warm season with cold nights conducive to toxin production The isolates were grown on moist, sterile rice, extracted with aqueous methanol, and examined for mycotoxin production. Zearalenone (ZEA) and the trichothecenes deoxynivalenol (DON), 3- and 15-acetyl-DON (AcDON), nivalenol (NIV), fusarenon-X (FX) and T-2 toxin (T-2) were analyzed by thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC) and confirmed by gas chromatography-mass spectrometry (GC-MS). Eleven of the 12 strains were DON and/or ZEA producers and 9 were AcDON positive. No NIV, or FX were detected. One strain produced T-2. The predominant acetyl-DON isomer was 15-AcDON. Mass-spectral analysis yielded detectable levels of other mycotoxins, 13-OH-apotrichothecenes, 11-epiapotrichothecenes, culmorin, sambucinol, and isotrichodermol being the most numerous. From the metabolic profiles it is suggested that UruguayanF. graminearum strains belong to the chemotype IB (DON/15-AcDON). The predominance of this chemotype is in accordance with data from Canada, United States, Mexico and Argentina which have similar climatic conditions that would favorF. graminearum growth and mycotoxin production.