pH-dependent effect of Congo Red on the growth of Aspergillus nidulans and Aspergillus niger

The azo dye Congo Red (CR) is frequently used as an agent to elicit cell wall integrity stress in fungi. This highly toxic aromatic, heterocyclic compound contains two azo bonds as chromophore, which are responsible for protonation under acidic conditions, leading to changes in the molecular structure of the dye and the color of the solution. The investigation of how CR affects the growth of Aspergillus nidulans and Aspergillus niger on surface cultures provided us with evidence about its pH-dependent toxicity. Reducing the starting pH of the media from 7 to 3 decreased both the toxicity of CR and the dose-dependence of its toxicity substantially. These changes can be explained by the pH-dependent structural changes of CR and its precipitation at low pH. The pH also depended on the fungi; they could induce a decrease or even an increase, which could be important in the loss of dose-dependence. Our experiments led to the conclusion that in studies to evaluate the antifungal effect of CR, properly buffered solutions with pH values adjusted to above 5 are highly recommended to achieve a well-detectable and dose-dependent antifungal effect. However, for decolorization of CR solutions, lower pH is suggested where the decreased toxicity and solubility of CR could help this process.