Cryoprotection of probiotic bacteria with poly-γ-glutamic acid produced by Bacillus subtilis and Bacillus licheniformis

Poly-γ-glutamic acid (γ-PGA) is a naturally occurring biopolymer made up of repeating units of glutamic acid and can be potentially used for multiple applications. This study compared the production of γ-PGA by Bacillus subtilis and Bacillus licheniformis in GS and E media. The highest γ-PGA production was achieved using initial glycerol concentration of 40 and 80 g/l, ammonium chloride as the nitrogen source, 20 g/l glutamic acid at pH 6.5 for 72 h using E medium. On characterization, it was observed that glutamic acid was the sole component of the purified material. It contained a mixture of Na-γ-PGA and H⁺-γ-PGA. The survival of probiotics during freeze drying was improved by combining them with γ-PGA polymer. For Lactobacilli, 10% γ-PGA protected the cells significantly than 10% sucrose during freeze drying. γ-PGA protection was shown to improve the viability of probiotic bacteria in orange juice for 40 days. No considerable change was observed in the concentrations of citric acid, malic acid and ascorbic acid when probiotic bacteria and γ-PGA were introduced into orange juice and hence, it could be used as a non-dairy delivery platform for these bacteria.