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Effect of Fluidized Bed Drying, Matrix Constituents and Structure on the Viability of Probiotic Lactobacillus paracasei ATCC 55544 during Storage at 4 °C, 25 °C and 37 °C.

ABSTRACT: The stabilization of probiotics for application in non-refrigerated food products is a challenging task. In the present study, probiotic Lactobacillus paracasei (Lacticaseibacillus paracasei) ATCC 55544 cells were immobilized in a dairy matrix comprising of whole milk powder, skim milk powder, or milk protein isolate using fluidized bed drying technology. The samples were taken out at different drying stages, with an apparent water activity (a_w) of a_w 0.5, a_w 0.4, and a_w 0.3, respectively, and vacuum-packed to maintain the a_w and stored at three different temperatures of 4 °C, 25 °C, and 37 °C. The study evaluated the impact of matrix constituents, milk fat, protein, and carbohydrate on the viability of encapsulated probiotic L . paracasei ATCC 55544 during storage for 1 month. The whole milk powder matrix provided superior protection to the bacteria. Confocal Laser Scanning Microscopy (CLSM) was used to investigate the structure of the immobilizing matrix and the location of the probiotic L. paracasei cells embedded within the matrix. The CLSM study revealed that the probiotic bacterial cells are mostly embedded as clusters beneath the top layer. We hypothesize that the biofilm-like structure, together with the protective whole milk powder matrix, helps to retain the superior viability of probiotic cells during storage at non-refrigerated storage conditions of 25 °C and 37 °C.

SUBMITTER: Poddar D

PROVIDER: S-EPMC8781091 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:We have developed butanol-producing consolidated bioprocessing from cellulosic substrates through coculture of cellulolytic clostridia and butanol-producing Clostridium saccharoperbutylacetonicum strain N1-4. However, the butanol fermentation by strain N1-4 (which has an optimal growth temperature of 30°C) is sensitive to the higher cultivation temperature of 37°C; the nature of this deleterious effect remains unclear. Comparison of the intracellular metabolites of strain N1-4 cultivated at 30°C and 37°C revealed decreased levels of multiple primary metabolites (notably including nucleic acids and cofactors) during growth at the higher temperature. Supplementation of the culture medium with 250 mg/liter adenine enhanced both cell growth (with the optical density at 600 nm increasing from 4.3 to 10.2) and butanol production (increasing from 3.9 g/liter to 9.6 g/liter) at 37°C, compared to those obtained without adenine supplementation, such that the supplemented 37°C culture exhibited growth and butanol production approaching those observed at 30°C in the absence of adenine supplementation. These improved properties were based on the maintenance of cell viability. We further showed that adenine supplementation enhanced cell viability during growth at 37°C by maintaining ATP levels and inhibiting spore formation. This work represents the first demonstration (to our knowledge) of the importance of adenine-related metabolism for clostridial butanol production, suggesting a new means of enhancing target pathways based on metabolite levels.IMPORTANCE Metabolomic analysis revealed decreased levels of multiple primary metabolites during growth at 37°C, compared to 30°C, in C. saccharoperbutylacetonicum strain N1-4. We found that adenine supplementation restored the cell growth and butanol production of strain N1-4 at 37°C. The effects of adenine supplementation reflected the maintenance of cell viability originating from the maintenance of ATP levels and the inhibition of spore formation. Thus, our metabolomic analysis identified the depleted metabolites that were required to maintain cell viability. Our strategy, which is expected to be applicable to a wide range of organisms, permits the identification of the limiting metabolic pathway, which can serve as a new target for molecular breeding. The other novel finding of this work is that adenine supplementation inhibits clostridial spore formation. The mechanism linking spore formation and metabolomic status in butanol-producing clostridia is expected to be the focus of further research.

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Effect of Fluidized Bed Drying, Matrix Constituents and Structure on the Viability of Probiotic Lactobacillus paracasei ATCC 55544 during Storage at 4 °C, 25 °C and 37 °C.

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