In order to prepare lyophilized cells of Bifidobacterium bifidum TMC3115 with high viability, the impact of the initial pH of the medium, different carbon and nitrogen sources, and inoculum quantity on the viable cell count was studied by the one-factor-at-a-time method. Subsequently, an orthogonal array design was applied to optimize combinations of carbon and nitrogen sources based on the pH of the medium at 16 h of culture, cell density (expressed as optical density) and viable cell count. The efficacy of 18 cryoprotectants on the probiotic strain was investigated, and the optimization of cryoprotectant blends was carried out using an orthogonal array design. The results showed that when the initial medium pH was 7.2 and the inoculum amount was 4%, the viable cell count reached the highest. The optimized medium was composed of glucose 25 g/L, tryptone 20 g/L, yeast peptone 10 g/L, yeast extract FM902 15 g/L, diammonium citrate 2 g/L, anhydrous sodium acetate 5 g/L, dipotassium phosphate 2 g/L, Tween-80 1 g/L, magnesium sulfate 0.58 g/L, manganese sulfate 0.25 g/L, and L-cysteine hydrochloride 0.5 g/L. The optimal cryoprotectant was composed of skim milk 12%, trehalose 12%, glycerol 1.2%, sodium glutamate 0.6%, and L-cysteine hydrochloride 0.15%. The viable cell count of the freeze-dried culture produced under the optimized conditions reached 4.26 × 1011 CFU/g.

A probiotic lactic acid bacterium, named LC577, was isolated from Tibetan kefir. This bacterial strain was identified as Lactococcuslactis casei based on its morphologic properties, 16S rRNA sequence analysis and physiological and biochemical characteristics. The strain had a strong ability to produce acid, and produced a cheese-like aroma after fermentation in skim milk. In addition, it showed tolerance to acid and bile. After 4 h culture in a simulated gastric fluid, the viable count increased slightly, but decreased to 6.15 × 105 CFU/mL after exposed to 3 g/L oxgall for 4 h.