In this work, the toxicological assessment towards filamentous fungi (Penicillium sp.) as model eukaryotic organisms of sixteen ionic liquids (containing an imidazolium, pyridinium, or cholinium cation) is presented. Amongst these fungi are members which show much higher tolerance towards ionic liquids than any other microorganism so far studied. Furthermore, guided by the paradigm that the choice of an ionic liquid as catalyst can alter the outcome of a given chemical reaction, the ability of ionic liquids to alter the metabolic profile in fungi was studied. The metabolic footprint, as investigated by electrospray ionisation mass spectrometry, revealed that fungal cultures respond to specific ionic liquids by changing their cell biochemistry, resulting in an altered pattern of secondary metabolites.