Process optimization of integrated ectoine production and secretion under osmotic stress in a cascade of continuously operated bioreactors

Process conditions for the synthesis and subsequent secretion of ectoine and hydroxyectoine were optimized in a cascade of two continuously operated bioreactors. In the first bioreactor the halophile bacterium Chromohalobacter salexigens was grown under continuous hyperosmotic and thermophilic conditions and therefore under constant stress driving the cells to accumulate large amounts of ectoines. Crossflow ultrafiltration was employed to concentrate biomass and therefore enhance high cell density cultivation. In the coupled second bioreactor the concentrated cell broth was osmotically down-shocked by addition of distilled water, forcing the cells to secrete the accumulated intracellular ectoines into the medium in order to avoid bursting. The cultivation conditions in the first bioreactor were optimized with respect to growth temperature and medium salinity to reach the highest productivity; the second bioreactor was optimized using a multi-objective approach for maximal secretion with a simultaneous minimization of cell death and product dilution. Depending on the cultivation conditions, intracellular ectoine and hydroxyectoine contents up to 540 and 400 mg per g cell dry weight were attained, respectively. With a maximum specific growth rate of 0.3 h in defined medium, productivities of approximately 2.1 g L h secreted ectoines in continuous operation were reached.