Publication: Integrated Co-Cultivation and subsequent Esterification: Harnessing Saccharomyces cerevisiae and Clostridium tyrobutyricum for Streamlined Ester Production

Together with our project partners from KIT (Karlsruhe, Germany), we have published our latest findings on co-cultivation in the journal Biotechnology for Biofuels and Bioproducts.

The rising demand for natural products is accelerating research into sustainable methods for producing bio-based flavourings like ethyl butyrate. In this study ethyl butyrate was successfully produced through the enzymatic esterification of butyric acid and ethanol, which were derived from the co-cultivation of Clostridium tyrobutyricum and Saccharomyces cerevisiae. Initial monoculture experiments with both strains were performed to investigate compromised fermentation conditions for co-cultivation. Based on these findings, anaerobic co-cultivation conditions were established at 37°C and 150 rpm, with the pH controlled at 6. The effects of varying inoculation times in co-culture were examined, considering the solvent and acid tolerance of both strains. Due to the limited acid tolerance of S. cerevisiae, with significant inhibition at butyric acid concentrations above 10 g L¯¹, a time-delayed inoculation with C. tyrobutyricum was implemented. In batch experiments, the final concentrations of butyric acid and ethanol were 13.98±3.06 g L¯¹ and 21.43±1.66 g L¯¹, respectively. Further enhancement of product concentrations was explored through a fed-batch cultivation strategy yielding up to 45.62±3.82 g L¯¹ of butyric acid and 18.61±4.11 g L¯¹ of ethanol. Ethyl butyrate was formed from the fermentation products by lipase-catalysed enzymatic esterification in a two-phase system through the addition of an organic phase. The ester concentration in the organic phase reached a maximum of 23.93±0.68 g L¯¹ (esterification yield 25%). This study presents a viable approach to the production of biobased ethyl butyrate offering a sustainable alternative to traditional chemical synthesis methods.

K. Oehlenschläger, M. Lorenz, E. Schepp, S. Di Nonno, D. Holtmann, R. Ulber; Integrated Co-Cultivation and subsequent Esterification: Harnessing Saccharomyces cerevisiae and Clostridium tyrobutyricum for Streamlined Ester Production; Biotechnology for Biofuels and Bioproducts (2025) in press