Carbon balancing of microbial fermentations is a valuable tool for the evaluation of the process performance and to identify the presence of undesired by-products. In this study, we demonstrate the relevance of total carbon (TC) analysis for carbon balancing in fermentations with the wild-type of Corynebacterium glutamicum by (i) quantifying significant amounts of dissolved inorganic carbonic species (TIC) in the culture medium and (ii) determining the effective (mass) carbon content of the biomass fraction (MC,X). In principle, TC based carbon balancing yielded at fully matching carbon balances. Thus, the application of our TC approach for the accurate detection of TIC and MC,X increased the total carbon recovery in standard batch fermentations with C. glutamicum on glucose from about 76\% to carbon closures of 94–100\% in contrast to conventional approaches. Besides, the origin of the missing 6\%-gap could be attributed to incomplete quantification of all carbon sources in the liquid phase. To conclude this study, the concept of TC-based balancing was transferred to an l-lysine production process, successfully quantifying relevant system carbon fractions, which resulted in matched carbon recoveries.

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