Methanol dehydrogenase inhibitors enhance methane bioconversion to methanol: Screening and comparative evaluation of highly efficient inhibitors

Methane is both a potent greenhouse gas and an abundant carbon resource, and its conversion to methanol is of environmental and industrial importance. However, methanol accumulation in biological methane conversion is limited by rapid oxidation catalyzed by methanol dehydrogenase (MDH). In this study, Methylosinus trichosporium OB3b was used to systematically compare four representative MDH inhibition strategies, including EDTA, MgCl2, cyclopropanol, and Cu2+, under a unified experimental framework. Batch experiments and enzyme activity analyses showed distinct responses among the tested inhibitors. EDTA achieved the highest short-term methanol accumulation, reaching 0.32 g/L within 24 h, approximately 220% higher than the control, but strongly impaired cell growth and operational stability. In contrast, 30 mM MgCl2 achieved relatively high methanol accumulation (0.30 g/L) while better maintaining cellular activity and MMO activity. Reactor-scale validation further showed that the MgCl2-regulated system reached 0.38 g/L methanol at 36 h under continuous CH4/air supply and exhibited comparatively better performance during repeated-batch operation. Cu2+ and cyclopropanol showed limited enhancement under the tested conditions. These results indicate that MDH inhibition strategies should be evaluated not only by peak methanol concentration, but also by cellular stability, enzyme activity, and operational robustness. MgCl2 was identified as a relatively mild and biocompatible regulator with potential for reactor-level methane-to-methanol bioconversion.