Highly efficient fermentation of 5-keto-d-fructose with Gluconobacter oxydans at different scales

Svenja Battling, Tobias Engel, Elena Ruth Herweg, Paul-Joachim Niehoff, Matthias Jan Pesch, Theresa Scholand, Marie Schöpping, Nina Sonntag, Jochen Büchs
2022 Microbial cell factories 21(1)  
The global market for sweeteners is increasing, and the food industry is constantly looking for new low-caloric sweeteners. The natural sweetener 5-keto-d-fructose is one such candidate. 5-Keto-d-fructose has a similar sweet taste quality as fructose. Developing a highly efficient 5-keto-d-fructose production process is key to being competitive with established sweeteners. Hence, the 5-keto-d-fructose production process was optimised regarding titre, yield, and productivity. Results: For
more » ... ion of 5-keto-d-fructose with G. oxydans 621H ΔhsdR pBBR1-p264-fdhSCL-ST an extended-batch fermentation was conducted. During fructose feeding, a decreasing respiratory activity occurred, despite sufficient carbon supply. Oxygen and second substrate limitation could be excluded as reasons for the decreasing respiration. It was demonstrated that a short period of oxygen limitation has no significant influence on 5-keto-d-fructose production, showing the robustness of this process. Increasing the medium concentration increased initial biomass formation. Applying a fructose feeding solution with a concentration of approx. 1200 g/L, a titre of 545 g/L 5-keto-d-fructose was reached. The yield was with 0.98 g 5-keto-d-fructose /g fructose close to the theoretical maximum. A 1200 g/L fructose solution has a viscosity of 450 mPa•s at a temperature of 55 °C. Hence, the solution itself and the whole peripheral feeding system need to be heated, to apply such a highly concentrated feeding solution. Thermal treatment of highly concentrated fructose solutions led to the formation of 5-hydroxymethylfurfural, which inhibited the 5-keto-d-fructose production. Therefore, fructose solutions were only heated to about 100 °C for approx. 10 min. An alternative feeding strategy was investigated using solid fructose cubes, reaching the highest productivities above 10 g 5-keto-d-fructose /L/h during feeding. Moreover, the scale-up of the 5-keto-d-fructose production to a 150 L pressurised fermenter was successfully demonstrated using liquid fructose solutions (745 g/L). Conclusion: We optimised the 5-keto-d-fructose production process and successfully increased titre, yield and productivity. By using solid fructose, we presented a second feeding strategy, which can be of great interest for further scale-up experiments. A first scale-up of this process was performed, showing the possibility for an industrial production of 5-keto-d-fructose.
doi:10.18154/rwth-2023-00681 fatcat:mskwzifidbbh3bcaazs62eummm