Bacillus subtilis natto: a non-toxic source of poly-γ-glutamic acid that could be used as a cryoprotectant for probiotic bacteria

It is common practice to freeze dry probiotic bacteria to improve their shelf life. However, the freeze drying process itself can be detrimental to their viability. The viability of probiotics could be maintained if they are administered within a microbially produced biodegradable polymer - poly-γ-glutamic acid (γ-PGA) - matrix. Although the antifreeze activity of γ-PGA is well known, it has not been used for maintaining the viability of probiotic bacteria during freeze drying. The aim of this study was to test the effect of γ-PGA (produced by B. subtilis natto ATCC 15245) on the viability of probiotic bacteria during freeze drying and to test the toxigenic potential of B. subtilis natto. 10% γ-PGA was found to protect Lactobacillus paracasei significantly better than 10% sucrose, whereas it showed comparable cryoprotectant activity to sucrose when it was used to protect Bifidobacterium breve and Bifidobacterium longum. Although γ-PGA is known to be non-toxic, it is crucial to ascertain the toxigenic potential of its source, B. subtilis natto. Presence of six genes that are known to encode for toxins were investigated: three component hemolysin (hbl D/A), three component non-haemolytic enterotoxin (nheB), B. cereus enterotoxin T (bceT), enterotoxin FM (entFM), sphingomyelinase (sph) and phosphatidylcholine-specific phospholipase (piplc). From our investigations, none of these six genes were present in B. subtilis natto. Moreover, haemolytic and lecithinase activities were found to be absent. Our work contributes a biodegradable polymer from a non-toxic source for the cryoprotection of probiotic bacteria, thus improving their survival during the manufacturing process

Poly-γ-glutamic acid: production, properties and applications

University of Wolverhampton, U

Electrospun Fibres of Polyhydroxybutyrate Synthesized by Ralstonia eutropha from Different Carbon Sources

The properties of PHB may be affected by the carbon source used in its production and this may affect nanofibres made from this polymer by electrospinning. In this study, P(3-HB) was produced from glucose, rapeseed oil, and olive oil by Ralstonia eutropha H16. Cell growth and polymer production were higher in olive or rapeseed oil supplemented media compared to glucose supplemented media. FT-IR, 1 H-, 13 C-NMR, and ESI/MS n confirmed that the synthesized polymers were P(3-HB). SEM micrograph showed the formation of nanofibres from P(3-HB) samples with the fibre diameters dependent on the source of the carbon used in polymer synthesis and the concentration of the polymer in the electrospinning solution. GPC showed that P(3-HB) from glucose (G-PHB) had a higher molecular weight ( 7.35 × 10 5  gmol −1 ) compared to P(3-HB) from rapeseed (R-PHB) and olive (O-PHB) oil. Differential scanning calorimetry (DSC) showed that the crystallinity of the electrospun polymers reduces with decreasing polymer concentration with R-PHB having lower crystallinity at all concentrations used. These observation shows that more PHB yield can be obtained using either rapeseed or olive oil compared to glucose with glucose producing polymers of higher molecular weight. It also show that electrospinning could be used to reduce the crystallinity of PHB fibres

Poly-��-Glutamic Acid: Biodegradable Polymer for Potential Protection of Beneficial Viruses

: Poly-��-glutamic acid (��-PGA) is a naturally occurring polymer, which due to its biodegradable, non-toxic and non-immunogenic properties has been used successfully in the food, medical and wastewater industries. A major hurdle in bacteriophage application is the inability of phage to persist for extended periods in the environment due to their susceptibility to environmental factors such as temperature, sunlight, desiccation and irradiation. Thus, the aim of this study was to protect useful phage from the harmful effect of these environmental factors using the ��-PGA biodegradable polymer. In addition, the association between ��-PGA and phage was investigated. Formulated phage (with 1% ��-PGA) and non-formulated phage were exposed to 50 ��C. A clear difference was noticed as viability of non-formulated phage was reduced to 21% at log10 1.3 PFU/mL, while phage formulated with ��-PGA was 84% at log10 5.2 PFU/mL after 24 h of exposure. In addition, formulated phage remained viable at log10 2.5 PFU/mL even after 24 h of exposure at pH 3 solution. In contrast, non-formulated phages were totally inactivated after the same time of exposure. In addition, non-formulated phages when exposed to UV irradiation died within 10 min. In contrast also phages formulated with 1% ��-PGA had a viability of log10 4.1 PFU/mL at the same exposure time. Microscopy showed a clear interaction between ��-PGA and phages. In conclusion, the results suggest that ��-PGA has an unique protective effect on phage particles.Iraqi cultural attach��, University of Wolverhampto