Divalent cations are antagonistic to survivability of freeze-dried probiotics encapsulated in cross-linked alginate

Alginate encapsulation is amongst the most studied probiotic encapsulation formulations in the literature, preferred for its low toxicity, low cost and ability to protect entrapped probiotics against acidic pH. Despite extensive academic studies, there has been little commercial translation of alginate-encapsulated probiotic formulations. In this study, we hypothesized that factors affecting freeze-drying survivability of alginate-encapsulated probiotics could have limited its translation potential. Factors including: 1) encapsulant material, 2) cryoprotectant, 3) cross-linker, 4) Concentration of crosslinker, and 5) other Group II divalent cations cross-linkers were investigated, using two model probiotics – Lactobacillus rhamnosus GG (LGG) and Escherichia coli Nissle 1917 (ECN). Divalent cations, which are used to cross-link alginates, were found to have antagonistic effects on the freeze-drying survivability of LGG and ECN. To mitigate the deleterious effects of divalent cations, the addition of sucrose was found to be useful, possibly due to its membrane stabilization mechanism. Agri-Food and Veterinary Authority of Singapore (AVA) Ministry of Education (MOE) Accepted version The first author is a recipient of the Nanyang President Graduate Scholarship from Nanyang Technological University, Singapore. This work was also supported by the Singapore Centre for Environmental Life Sciences Engineering (SCELSE) (MOE/RCE: M4330019.C70), Ministry of Education AcRF grant (RG19/18 & RT08/19), Agri-Food & Veterinary Authority of Singapore (APF LCK102), Biomedical Research Council (BMRC) – Therapeutics Development Review (TDR-G-004-001), NTU-HSPH grant (NTU-HSPH 17002), and the Bill and Melinda Gates Foundation (OPP1199116).