Antifungal activity of kefir against bread spoilage fungi

Abstract

Food spoilage is a major issue for the food industry, leading to food waste, substantial economic losses for manufacturers and consumers. In previous work, the antimicrobial capacity of kefir, an artisanal, acidic and slightly carbonated fermented drink, has been tested, showing antibacterial activity. Therefore, the objective of the present study was to evaluate the antifungal activity of kefir against bread spoilage fungi. 21 microorganisms were isolated from kefir but none presented the desired activity, indicating that microorganism need to grow in a consortium to produce it. For this reason, it was decided to continue working with kefir granules. Two kefir drinks were used, one with unsterilized mascabo sugar and the other with sterilized mascabo sugar (5 g%). A total of 3 g of kefir granules were inoculated in 30 mL of each medium and they were incubated to 30°C without and with orbital shaker at 140 rpm. Samples were taken after 72h and the cell-free supernatant (CLS) was obtained by centrifugation at 10,000 x g for 10 minutes. The SLC samples were subjected to two concentration methods: water bath at 94°C for one hour and Savant speedvac for 3 h, until a final concentration of 5x by both techniques. The antifungal activity was determined by the agar diffusion technique against six fungi isolated from bread and not yet identified. The fungus was plated onto potato dextrose agar by spreading a volume (100 L) inoculum. Sterile mascabo sugar was used as a control. All CLS showed antifungal activity, varying the degree of inhibition according to the concentration method. The CLS kefir with sterilized mascabo sugar showed greater activity than the CLS with unsterilized sugar, showing in decreasing order of fungal inhibition: SLC Savant speedvac, SLC water bath, SLC without concentrating and static culture (without shaking). The control did not show activity. In addition, thin layer chromatography was carried out with the concentrated and unconcentrated CLS, using BAW (butanol: acetic acid: water) as the mobile phase, and bioautography was carried out using as reporter the most sensitive fungus from the previous tests, obtaining one metabolite with fungal inhibition of Rf = 0.3, confirming the presence of an antifungal metabolite. It is concluded that cultivating kefir with agitation favors the production of the antifungal metabolite and that the application of the two methods to concentrate the SLC were effective in increasing fungal inhibition, however the high temperatures used would have a slight effect of decreasing this capacity. In future work it is expected to advance on the nature of the metabolite.