RESUMEN
Hydroxyectoine, an ectoine derivative, is the most common compatible solute in halophilic microorganisms for resisting harsh environments. Compatible solutes can be utilized in fields such as cosmetics, medicine, and biochemistry. Moderately halophilic microorganisms produce much less hydroxyectoine as compared with ectoine. In this study, we first evaluate the effect of medium formulation (i.e., yeast extract (YE) medium and high yeast extract (HYE) medium) on hydroxyectoine production. In addition, an investigation of hydroxyectoine production by Halomonas salina under optimal conditions for vital factors (i.e., iron and α-ketoglutarate) and hydroxylase activity was also carried out. As a result, hydroxyectoine production was obviously elevated (0.9 g/L to 1.8 g/L) when the HYE medium was utilized. Furthermore, hydroxyectoine production further increased to 2.4 g/L when both the α-ketoglutarate and iron factors were added to the HYE medium in the early stationary phase. In addition, we found that ectoine hydroxylase activity increased more when a combination of iron and α-ketoglutarate was used than when either was used alone. The results showed that the alteration of iron and α-ketoglutarate clearly stimulated the expression of ectoine hydroxylase, which in turn affected hydroxyectoine synthesis. This study also showed that hydroxyectoine production was further raised from 2.4 g/L to 2.9 g/L when 50 mM of α-ketoglutarate and 1 mM of iron were added to the HYE medium. Ultimately, the experimental results showed using the optimal conditions further elevated the hydroxyectoine production yield to 2.90 g/L, which was over 3-fold higher than the best results obtained from the original medium.
Asunto(s)
Aminoácidos Diaminos/metabolismo , Fermentación/fisiología , Halomonas/metabolismo , Reactores Biológicos/microbiología , Medios de Cultivo/química , Halomonas/enzimología , Hierro/química , Ácidos Cetoglutáricos/química , Ingeniería Metabólica/métodos , Técnicas Microbiológicas/métodos , Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Tolerancia a la SalRESUMEN
This study attempted to utilize Halomonas salina BCRC17875 to produce ectoine by optimizing the agitation speed and medium composition. In addition, the chemical structure of ectoine produced by H. salina BCRC17875 was determined. The results indicate that ectoine production reached 3.65 g/L at 38 h of cultivation when the agitation rate and NaCl concentration were fixed at 200 rpm and 2.0 M, respectively. It reached 9.20 g/L at 44 h of cultivation when the major medium components were yeast extract (56 g/L), glutamate (74.40 g/L), and ammonium sulfate (14 g/L). After the nitrogen concentration had been evaluated, evaluation of the nitrogen concentration revealed that the ectoine production reached 11.80 g/L at 44 h of cultivation when 56 g/L of yeast extract and 28 g/L of ammonium sulfate were used. Ectoine production reached 13.96 g/L at 44 h of cultivation when the carbon/nitrogen ratio was fixed at 3/1 using 84 g/L of yeast extract and 28 g/L of ammonium sulfate. Furthermore, the identification of ectoine were identified and characterized by fast atom bombardment mass spectrometry (FAB-MS) and 1H NMR. The results demonstrated a fermentation strategy was successful in increasing ectoine production, and that the fermentation medium of ectoine had commercialization potential.