Inhibitory effects of binary combinations of microbial metabolites on the growth of tolerant Penicillium roqueforti and Mucor circinelloides

Dairy product spoilage by molds leads to substantial food waste and economic losses. Although some bioprotective lactic acid bacteria exhibit antifungal activity, some spoilage molds displayed extensive tolerance to these antifungal cultures and their metabolites. This study aimed to investigate some combinations of selected microbial compounds on two molds, Penicillium roqueforti ISI4 and Mucor circinelloides 01180023, previously found to be tolerant. Of the thirteen compounds tested in susceptibility test, octanoic acid (OA) exhibited the most potent antifungal activity, followed by diacetyl and 3-Phenylpropanoic acid (3-PPA). Six compounds were chosen for assessing the antifungal activity of binary combinations. OA combined with either diacetyl (COD) or 3PPA (COP) displayed synergistic antifungal effects in laboratory medium and yoghurt. Using oCelloScope, mold growth inhibition and morphological changes induced by COD and COP were observed. The increase of extracellular conductivity, intracellular materials leakage, and the propidium iodide-stained spores demonstrated the membrane was damaged. The accumulation of reactive oxygen species and the elevated malondialdehyde (MDA) content indicated the two combinations induced oxidative stress, leading to membrane lipid peroxidation. These findings highlight some combinations may exhibit synergistic effects and open up for opportunities to design culture and metabolite mixes with effect even towards very robust mold species.