Intracellular sorbitol improves the survival rate of Lactiplantibacillus plantarum after freeze‐drying

Abstract BACKGROUND During the lyophilization (i.e., freeze‐drying) of probiotics, the addition of a protective agent is essential for the survival of the frozen bacterial cells. Although many studies have explored the effects of exogenous protective agents during freeze‐drying, whether these protective agents play an intracellular role requires study. Sorbitol is one of the most commonly used protective agents. In Lactiplantibacillus plantarum , srlD1/srlD2 is a key gene for sorbitol synthesis, and the pts‐srlD gene cluster controls sorbitol transport. The objective of this study was to analyze the effect of sorbitol as an endogenous protective agent during the lyophilization of L. plantarum . Intracellular sorbitol levels were successfully controlled by the overexpression and knockout of sorbitol synthesis ( srlD ) or transport ( pts‐srlD ) genes. Meanwhile, knockout of ldh will shift the metabolism of Fru‐6P to the synthesis of sorbitol. RESULTS In phosphate‐buffered saline alone, the survival rate of the AR113Δ ldh‐srlD2 strain, which had a large amount of intracellular sorbitol, reached 41.9% – a rate 3.2 times higher than that of AR113‐ srlD2 . The addition of sorbitol decreased the survival of transport system‐knockout strains compared with the wild‐type strain, and the transport system‐restored strain had a survival rate 3.1 times higher than that of the wild‐type strain. CONCLUSION These results show that sorbitol plays an intracellular role in L. plantarum , and exogenous sorbitol is transported into the cell, where it exerts a protective effect. This study provides a theoretical basis for the application of protective agents. © 2026 Society of Chemical Industry.