Fermentation Strategies to Improve Argentinian Kefir Quality: Impact of Double Fermentation on Physicochemical, Microbial, and Functional Properties

Abstract

This present study investigated the microbial dynamics, physicochemical and functional properties, and sensory characteristics of kefir produced by two different approaches: traditional kefir obtained directly from grains and kefir manufactured through a double-fermentation process in cow milk. For the first fermentation, kefir grains were inoculated in milk at different levels (1%, 3%, and 5% w/v) and incubated at 30 °C for 24 h. The lowest inoculation level promoted the greatest increase in grain biomass, whereas higher inoculation levels produced more pronounced pH decreases. All products maintained stable pH values during refrigerated storage at 4 °C for 15 days. Products derived from initial fermentations with 1% and 3% inoculum were subsequently used in a second fermentation step at two inoculation levels (1% and 10% v/v) to produce double-fermentation kefir products. These products exhibited higher counts of lactic acid bacteria and reduced yeast populations compared with traditional grain kefir. After 15 days of storage, all kefir samples maintained more than 108 CFU/mL of lactic acid bacteria, more than 107 CFU/mL of acetic acid bacteria, and around 105 CFU/mL of yeasts. Protein content was comparable among all kefir products and unfermented milk. The product obtained with 1% grains followed by 10% v/v inoculation showed enhanced biofilm formation that increased during storage and displayed the strongest antimicrobial activity, and was therefore selected for sensory evaluation, where it achieved favorable acceptance by regular kefir consumers.