Synthesis and release of ectoines in a moderate halophile Halomonas sp. Y subjected to osmotic shocks

Abstract:

[Background] Ectoines have been applied in cosmetics, enzyme preparations and pharmaceutical industry due to their powerful stabilizing property of protecting enzyme, protein, nucleic acid and the whole cell against high temperature, freeze and dryness. Up to now, industrial-scale production of ectoines relies on bacterial milking process using moderately halophilic bacterium. Therefore, development of high-yield ectoine-producers and related fermentative technology has attracted considerable attention worldwide. [Objective] The objective of the present study was to isolate a moderately halophilic bacterium capable of synthesizing ectoines as main compatible solute, study the effects of osmotic shocks on synthesis and release of ectoines in this bacterium and explore the feasibility of producing ectoines by using bacterial milking process. [Methods] Moderately halophilic bacterium was isolated by spread-plate method and identified through analysis of its morphology, physicochemical properties and 16S rRNA gene sequence similarity. Ectoines were analyzed quantitatively and qualitatively by high performance liquid chromatography (HPLC) and mass spectrometry (MS), respectively. Ectoines were prepared by bacterial milking process. [Results] A moderately halophilic bacterium Y capable of synthesizing ectoines as main compatible solute was isolated from saltern sediment and identified as a member of the genus Halomonas according to its physiological properties and 16S rRNA gene sequence similarity. Halomonas sp. Y showed growth in lactate medium containing a wide range of NaCl concentrations of 10?250 g/L. Optimum growth was observed in the medium containing 100 g/L NaCl. HPLC-MS analysis revealed that this bacterium could synthesize both ectoine and hydroxyectoine as the main compatible solute. The yields of each compatible solute synthesized in the presence of 100 g/L NaCl were 175.5 mg/g and 47.9 mg/g for ectoine and hydroxyectoine, respectively. Maximum release rate of intracellular ectoines could be achieved within 5 min when Y cells experienced hypo-osmotic stress brought about by 0?30 g/L NaCl solutions. Hypo-osmotic saline solution with salinity of 10 g/L NaCl was the most appropriate for ectoine release during bacterial milking process. During the bacterial milking process eleven rounds of hyper/hypo osmotic shocks were exerted to the isolate and the total yields of synthesized and released ectoines were 6.0 g/L and 5.7 g/L, respectively. This corresponded to an average release rate of 64.5% and a high conversion rate of 128.9 mg/g. [Conclusion] Halomonas sp. Y is a moderately halophilic bacterium with quite high yield of ectoines. It could resist repeated osmotic shocks as characterized by ectoine release under hypo-osmotic stress and synthesis of ectoine under hyper-osmotic condition. Ectoine productivity was significantly improved by using bacterial milking process.