L-lactic acid (L-LA), a naturally occurring organic acid, has extensive applications in the food, pharmaceutical, textile, leather, and chemical industries (Kim et al., 2022, Wang et al., 2016a). As the optically pure isomer of lactic acid (LA), L-LA exhibits unique properties, including participation in biochemical reactions and potential for structural optimization of biodegradable polylactic acid (PLA) (Taib et al., 2023). Microbial fermentation provides a sustainable and efficient method for L-LA production (Liu et al., 2022). Both wild-type lactic acid bacteria [e.g. Bacillus coagulans (B. coagulans) and Lactobacillus rhamnosus (L. rhamnosus)] (Chen et al., 2020b, Granget et al., 2024) and engineered microorganisms [e.g. Saccharomyces cerevisiae (S. cerevisiae) and Rhizopus oryzae (R. oryzae)] (Choi et al., 2024, Wu et al., 2025, Zhu et al., 2024) have been adopted for L-lactic acid biosynthesis. To maintain the optimal biocatalytic environment, various neutralizers have been employed in LA fermentation, including Ca(OH)2, CaCO3, NaOH, and NH3·H2O (Othman et al., 2017). However, the use of NH3·H2O, NaOH, and KOH can inhibit product accumulation and cell growth due to the toxicity of excess ammonia in addition to lactate ions to LA producers (Wang et al., 2022, Yang et al., 2015). Calcium-based neutralizers are considered mild agents that can enhance fermentation performance (Yang et al., 2015). Nevertheless, their use leads to the formation of solid waste (calcium sulfate) during the purification process (Balakrishnan et al., 2020). Moreover, conventional neutralizing agents typically result in L-LA titers ranging from 100 to 200 g/L and generate considerable wastewater due to limited product concentrations (Wang et al., 2022). In contrast, magnesium oxide (MgO) has shown potential to reduce energy consumption, substrate costs, and waste emissions during L-LA biosynthesis (Ren et al., 2022). Moreover, the presence of Mg2+ enhances microbial energy supply and accelerates metabolic activity by modulating ATP-related enzyme functions or directly forming ATP-Mg2+ complexes, thereby improving fermentation efficiency (Pawlosky et al., 2025). Additionally, Mg2+ can partially inhibit bacterial autolysis by down-regulating genes responsible for cell wall hydrolysis and up-regulating those associated with cell growth and division, ultimately enhancing fermentation stability (Rismondo and Schulz, 2021).

Although an MgO-coupled L-LA biosynthesis process has been developed, mesophilic fermentation using L. rhamnosus remains susceptible to microbial contamination during extended operations (Wang et al., 2022). In contrast, B. coagulans is capable of producing LA with high optical purity and demonstrates exceptional tolerance to elevated temperatures and nutrient limitations (Ma et al., 2016, Ouyang et al., 2020, Zhang et al., 2014). To establish MgO-based open fermentation for L-LA production, this study investigates the physiological responses of B. coagulans to high magnesium stress and explores their correlation with enhanced L-LA biosynthesis.

As one of the most critical tools for large-scale analysis of gene expression, transcriptomics focuses on gene expression at the RNA level and provides genome-wide information on gene structure and function to elucidate the molecular mechanisms underlying specific biological processes (Dong and Chen, 2013). In parallel, metabolomics offers deeper insight into the biochemical basis of stress response mechanisms (Parlindungan and Jones, 2023). Transcriptomic analysis was employed to elucidate the acid resistance mechanisms of an acid-tolerant Lactococcus lactis WH101 (L. lactis WH101) strain obtained through mutagenesis, in which intracellular pH homeostasis and energy supply were improved by enhancing carbohydrate, amino acid, and fatty acid metabolism, resulting in a 12-fold increase in survival under acid stress following overexpression of the ArcB and MalQ genes (Zhu et al., 2019). Additionally, metabolomics analysis has been used to evaluate the effects of different Cu(II) concentrations on LAB, revealing concentration-different impacts on metabolic pathways (Li et al., 2023). Additionally, untargeted metabolomics was applied to investigate the nitrite degradation mechanism in Lactobacillus fermentum RC4 (L. fermentum RC4), identifying key pathways and highlighting the role of enhanced ATP supply and exogenous glucose in promoting nitrite reduction (Shi et al., 2022). L-LA production of B. coagulans LA1507 was significantly enhanced using MgO as neutralizer in this study, with increases of 7.61 % in yield, 22.22 % in productivity, 21.81 % in titer, and 18.50 times in cell viability. And integrated transcriptomic and metabolomic analyses were conducted to explore the regulatory networks and molecular mechanisms by which B. coagulans LA1507 respond to high magnesium stress. A total of nine shared metabolic pathways and one genetic pathway were identified, involving 139 differentially expressed genes and 29 differential metabolites.