Wax ester and lipophilic compound profiling of Euglena gracilis by gas chromatography-mass spectrometry: toward understanding of wax ester fermentation under hypoxia

Lipids are being increasingly used as biodiesel feedstock, and several saturated wax esters from Euglena gracilis are candidates for outdoor bulk production. Wax ester fermentation in Euglena is strongly increased by hypoxia, but key events underlying the metabolic shift toward wax ester biosynthesis are poorly understood. Profiling of wax esters and other lipophilic compounds is the first step for research toward the clarification of wax ester fermentation molecular mechanisms, and thus, a simple and comprehensive platform for their profiling is required. In this study, we established a profiling method for wax esters and lipophilic compounds in Euglena using gas chromatography-mass spectrometry (GC-MS). Using this method, we compared accumulation profiles of wax esters and lipophilic compounds between a wild-type Euglena Z strain and a bleached SM-ZK strain. Both the wild-type and the bleached strains contained C14:0 fatty acid-C14:0 fatty alcohol as a dominant wax ester. Wax ester fermentation initiated 4 h after the cessation of oxygen supply by halting the culture agitation resulting in linear increase and proportional changes of wax ester amounts during 24 h. However, complete anoxia by nitrogen gas aeration inhibited wax ester production and the addition of bicarbonates reversed the inhibition, suggesting that there is a need for an additional carbon source for wax ester fermentation under anoxia. Our simple method enables the investigation of metabolic responses leading to wax ester fermentation in Euglena.