Bacterial cultures of lactic and propionic acid bacteria are widely used in fermented products including dairy products. Spoilage fungi may constitute a quality and safety issue in these products. The antifungal properties of some lactic and propionic acid bacteria make them potential candidates for prolonging shelf-life of food without the addition of specific preservatives. Increased interest in the use of these bacteria for biopreservation has led to identification of a range of potent strains, and in addition, isolation and identification of various antifungal metabolites produced by these cultures. The role of the different metabolites in the specific interactions and the mechanisms behind the antifungal activity of protective cultures has not been explored to a large degree because of the complexity of the interactions between microorganisms and matrices. More work is needed to clarify determining factors thereby facilitating development of new protective cultures.

The aim of the current thesis was to determine and study metabolites involved in antifungal activity of protective Lactobacillus paracasei and Propionibacterium freudenreichii subsp. shermanii. This involved development of a suitable, defined medium to simplify the initial identification of metabolites. More specifically, the study included investigation of a possible role of volatiles in antifungal activity as well as application of multivariate analysis for studying the whole exometabolome of the protective bacteria.

A chemically defined interaction medium (CDIM) was developed allowing growth of protective Lb. paracasei and P. freudenreichii subsp. shermaniii as well as the spoilage fungi, Penicillium spp., Rhodotorula mucilaginosa and Debaryomyces hansenii isolated from fermented dairy products. Lb. paracasei and P. freudenreichii subsp. shermanii grew in CDIM and showed antifungal properties similar to those observed in milk-based systems. Most of the antifungal effect of the protective bacterial ferment was lost after removal of cells. This was explained by a marked decrease in diacetyl concentration, which was the metabolite having the main activity of Lb. paracasei DGCC 2132 in CDIM. Addition of a P. freudenreichii subsp. shermanii DGCC 2053 culture stimulated the growth of Lb. paracasei and further increased the diacetyl production as well as the antifungal activity of Lb. paracasei DGCC 2132.

Metabolic footprinting was used to study the overall exometabolome of cell-free ferments of three Lb. paracasei strains. The metabolites potentially causing the main antifungal activity of these Lb. paracasei strains were identified by correlating the results of the antifungal activity test with the metabolites produced. Besides diacetyl and lactic acid, 6 antifungal hydroxy acids were identified. Of these, 3 have previously been reported from antifungal lactic acid bacteria, whereas the other 3 hydroxy acids have not previously been reported produced by antifungal lactic acid bacteria.