Construction of a rapid succinate concentration detection system based on the specific catalysis of succinate...

During succinic acid fermentation, substrate consumption, product accumulation, and byproduct formation significantly influence cellular metabolic state and final production yield. Rapid and accurate quantification of succinic acid is essential for process optimization, metabolic regulation, and online monitoring of fermentation. Therefore, developing a sensitive, rapid, and highly specific detection method applicable to complex fermentation samples is of considerable theoretical significance and practical application value. To achieve rapid and high-throughput quantification of succinate during fermentation, this study established an enzymatic detection system utilizing the specific catalytic properties of succinate dehydrogenase (SDH). Starting with Escherichia coli DH5α as the parental strain, a series of subunit-deletion mutants were constructed. Crude SDH preparations were extracted via ultrasonic lysis, and functional characterization demonstrated that subunit deficiencies not only compromised bacterial growth but also resulted in complete loss of SDH catalytic activity. Subsequently, using the crude enzyme solution with catalytic activity as the raw material, the extraction process of SDH was studied. After separation and purification, a protein with a specific activity of 155 U/mg of SDH was obtained. The enzymatic characteristics of the were systematically investigated. Finally, with SDH as the core component, a flexible detection system suitable for diverse application scenarios was constructed, supporting both high-throughput analysis using 96-well microplates and rapid single-sample testing. Following optimization, the detection system exhibited a linear response range of 0.5-50 g/L with R² > 0.97, satisfying the analytical demands for process monitoring in succinate fermentation and high-throughput screening of succinate-producing strains.