Daily kefir grain increase mass was studied using fresh HTP whole fat cow’s milk (Ljubljanske mlekarne d. d.) as a culture medium. Its chemical composition is 3.2 % proteins, 4.6 % carbohydrates, 3.5 % fat and 0.13 % calcium. 3D-(+) Glucose anhydrous (Fluka) was obtained from commercial sources. Kefir grains, used as inocolum in this study, originate from Caucasian Mountain and were acquired from an internationally recognized local dairy (Kele & Kele d. o.o.). Their detailed microbial composition was not analyzed. Importantly, the microbial population (bacteria and yeasts) of kefir grains depends on many different factors (age, storage conditions and fermentation medium) and varies with the season. It is almost impossible to assure equal microbial composition during long term period, therefore for sets of experiments within one research, kefir grains with the same viability should be used.

1.2 Kefir grain biomass activation

Kefir grain biomass activation was performed in a glass lab beaker. The collected inactive kefir grains (^kG = 40 g/L) were inoculated in 1 L of fresh HTP whole fat cow’s milk. After
incubation at room temperature ($ = (22 + 2) °C) for 24 h, the grains were separated from the kefir beverages using a household sieve. After washing, they were reinoculated into the fresh milk. The same procedure was repeated over six subsequent days. After this procedure the kefir grains were considered active.