Methods: Lactobacillus AF51 was grown in MRS medium to stationary phase and harvested by centrifugation. The pellet was resuspended in 0.5 M standard cryoprotectants, sucrose and trehalose, and a list of testing cryoprotecting agents. Bacteria were frozen at -30°C and dried with a Labconco lyophilizer for 24 h and stored at 4°C, 20°C and 33°C. The shelf-life of the freeze-dried lactobacilli was analyzed by colony count as a function of time.
Results: One of the testing formulae (Formula A) was more effective than the standard sucrose or trehalose formula and was further analyzed. Four weeks after freeze-drying, all three formulae had no apparent viability loss when stored at 4°C (about 10(10) cfu/gram) and lost about 1 log when stored at 20°C. When the formulae were stored at 33°C, however, no viable lactobacilli were detectable in the sucrose or trehalose formula, but Formula A showed an equally high number of viable cells as lactobacilli stored at 20°C, about 10(9) cfu/gram. Functional analysis of the dried lactobacilli in formula A showed rapid revival and equal ability to bind mannan as fresh cells.
Conclusions: We have developed a formula that can maintain viability of lactobacilli during storage in warm climate. Supported by IADR/GSK Innovation in Oral Care Award and NIH grant AI068528.