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The effectiveness of formulations using different protective agents to maintain viability of Lactobacillus salivarius I 24 during freeze drying for production of live cell in powdered form was investigated. The influence of prefreezing and cultivation conditions on viability of cells after freeze drying was also studied. Surface methodology was used to determine the most suitable combination of the protective agents. Concentrations of skim milk, sucrose, glycerol, and calcium carbonate were selected as operating variables, and survivals of cultures after freeze drying were used as results. Skim milk and sucrose were better protective agents than glycerol and calcium carbonate when used individually for preserving L. salivarius I 24 during freeze drying. Their protective abilities could be enhanced significantly when using them as a mixture (9.85% w/v skim milk and 10.65% w/v sucrose). Prefreezing of the cells at −80°C for 5 h prior to freeze drying and cultivation with regulated pH and temperature gave the highest cell viability.

The potential use of fed-batch cultivation (FBC) for improvement of the production of I 24 biomass for subsequent use as probiotics was studied using a 2-L stirredtank bioreactor. Three different constant feeding rates (0.1, 0.05, and 0.033 L/h) were applied in FBCs and their effect on carbon metabolism was evaluated. The carbon flux for cell built-up with reduction in lactic acid synthesis was observed in the fed-batch as compared to the batch cultivation mode. The viable cell number obtained in the constant FBC (CFBC) operated at a feeding rate of 0.05 L/h was 8 times higher (10.7×10 CFU/mL) than that recorded in the batch cultivation. This gave the viable cell yield based on glucose consumed for CFBC of 26 times higher (11.3×10 CFU/g) than the batch cultivation. This study demonstrated CFBC, which is simple with minimal use of process control equipment, has an industrial potential for improvement of probiotic production.

Summary This study investigated the probiotic characteristics of lactic acid bacteria (LAB) isolated from 14 Malaysian fermented food or milk products. In total, 22.3% (121 of 542) of the LAB isolated from the local fermented products exhibited antibacterial activity against Micrococcus luteus. Twelve LAB that demonstrated better antibacterial activity against M. luteus as compared to a commercial strain Lactobacillus casei strain Shirota (LABPC) were selected for further characterisation. Based on 16S rRNA gene sequence, the LAB were identified as pediococci (seven) and lactobacilli (five). All 12 LAB showed bile tolerance, but only eight were acid tolerant at pH ≥ 3.0. The highest level of adhesion to HT‐29 cells was observed among the Lactobacillus sp. LAB 1 and 10. The LAB also showed antimicrobial activity against Escherichia coli and Staphylococcus aureus through the production of organic acids. LAB isolated from Malaysian fermented food and milk products, especially fermented tapioca, contains potential probiotic candidates.

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