Water kefir grains are an important source of probiotics, mainly containing lactic acid bacteria and yeasts. The aim of this study is to investigate the changes in microbial and chemical properties of water kefir grains during 1-month storage at +4°C and −18°C. The initial content of lactobacilli, lactococci, and yeast in water kefir grains was 6.06, 6.33, and 5.93 log CFU/g, respectively. The number of lactobacilli, Lactobacillus acidophilus, and Bifidobacterium spp. in the water kefir grains were comparable, with slight changes at the end of refrigerated storage (p >.05). Lactococci and yeasts decreased significantly after both storage conditions compared to the initial content (p <.05). The dry matter and ash contents remained unchanged during storage (p >.05). Water kefir grains contained significant amounts of calcium, vitamin B2, vitamin B6, vitamin B7, and vitamin B12. Storage at both +4°C and −18°C did not affect the mineral and vitamin contents, except for Cu and Vitamin B2. The results indicate that the water kefir grains remained viable after storage at both temperatures. If water kefir grains need to be stored, it is recommended to store them at +4°C in sugared water as it ensures better survivability of the microbiota of the grains.