The production of secondary metabolites in the submerged co-cultures of Penicillium rubens Wisconsin 54-1255 and Aspergillus terreus ATCC 20542 was evaluated. The biosynthetic capabilities of the two strains were compared in a set of diverse liquid media that differed with respect to the initial levels of glucose, lactose and yeast extract, contained carrot juice or vegetable/turkey puree as additional nutrient sources or were supplemented with phenylacetic acid, the side-chain precursor of penicillin G. The main goal of the study was to investigate the interactions between A. terreus and P. rubens that might contribute to the changes of secondary metabolite titers. Briefly, the biosynthesis of octaketide metabolites (+)-geodin and asterric acid was visibly enhanced as a result of replacing the conventional monocultures with the co-culture systems, but solely in the media containing not more than 5 g L-1 of yeast extract. By contrast, no marked enhancement was observed with respect to the biosynthesis of penicillin G, lovastatin, chrysogine, 4a,5-dihydromevinolinic acid and 3a-hydroxy-3,5-dihydromonacolin L acid. It was shown that the relationships between medium composition and product titers were clearly different in monoculture variants than in the corresponding co-cultures. Finally, it was demonstrated that the utilization of penicillin precursors by P. rubens can be blocked under the conditions of co-cultivation. (C) 2020, The Society for Biotechnology, Japan. All rights reserved.