Novel combined effect of sublethal heat and ethanol for enhanced gamma-aminobutyric acid production via whole-cell catalysis of Lactiplantibacillus plantarum

Maintaining cell envelope integrity and intracellular enzyme stability while alleviating transmembrane mass-transfer limitations is important for sustained γ-aminobutyric acid (GABA) production by whole-cell biocatalysis. We investigated the effects of ethanol on the apparent whole-cell glutamate decarboxylase (GAD) activity of Lactiplantibacillus plantarum SS5 and optimized the corresponding GABA production process. Co-incubation with 6–8% (v/v) ethanol increased the apparent whole-cell GAD activity and its residual activity after prolonged heat exposure. Scanning electron microscopy and extracellular GAD assays after 30 min of treatment with 7% (v/v) ethanol showed no gross morphological disruption or detectable GAD leakage. After ethanol removal, the apparent whole-cell GAD activity of ethanol-treated cells showed no significant difference from that of untreated cells ( p > 0.05). These findings are consistent with a transient, condition-dependent effect of ethanol on the cell envelope. The optimized system consisted of a 0.20 mol·L⁻¹ L-monosodium glutamate (MSG) working solution and a 100 mg·mL⁻¹ wet cell suspension mixed at a volume ratio of 1:1 (v/v), followed by the addition of 6% (v/v) ethanol (approximately 0.09 mol·L⁻¹ MSG in the complete reaction mixture), with the reaction conditions set at pH 5.2, 42 °C, 80 rpm, and a reaction time of 56 h. In triplicate experiments, the GABA titer reached 8.87 ± 0.13 mmol·L⁻¹, corresponding to a 27.23 ± 1.83% increase over the ethanol-free control. Thus, ethanol can serve as a food-grade process additive for enhancing the efficiency of this laboratory-scale whole-cell bioconversion process.