Study On Impact Factors And Related Physiology Of Lipid And EPA Accumulation In Nannochloropsis Oculata

Nannochloropsis sp. with several advantages such as good adaptive capability to theenvironment, short growth cycle, high photosynthetic efficiency, could accumulate highcontent of lipid and EPA. In this study, the strain of Nannochropsis oculata C170was selected,the impact factors and related physiology of lipid and EPA in algae cells were investigatedpurposefully. And then, the initiatives that could promote the accumulation of lipid and EPAthrough adding intermediate metabolites were studied in detail. The main results wereillustrated as follows:The method with Nile Red as fluorochrome and dimethylsulfoxide as permeation toassay algal lipid content in Nannochropsis oculata C170was established. The optimumconditions were: excitation and emission wavelength of480and565nm respectively,dimethylsulfoxide volumn fraction of20%, Nile Red concentration of1.5μg·mL-1, stainingtime of20min, cell density of0.9-3.1×107cells per milliliter. The relative fluorescence ofstained cells had a linear relationship with the lipid content (R2=0.9923). The lipid body withobvious yellow fluorescence was also observed by the laser scanning confocal microscopy inoptimum staining conditions.Several impact factors that would affect the accumulation of lipid and EPAwere studied.It was found that the algae had the highest proportion of EPA(18.76%of total fatty acid) withthe nitrogen concentration of42.5mg·L-1. The lipid content was the highest which was29.17%of dry weight supplemented with12.5mg·L-1nitrogen. When the concentration ofFe3+was6.51mg·L-1, the algae biomass and lipid content were the highest of0.54g·L-1and22.03%, respectively. With the sanlinity of24‰, the algae biomass and productivity were thehighest of0.52mg·L-1and42.99mg·L-1·d-1, respectively. The highest lipid content (33.68%)was obtained under the salinity of54‰. Furthermore, the algae biomass and lipid cotent werethe highest when the pH value was8.5and the EPAcontent peaked at pH5.5.Under nitrogen-limiting conditions, the related physiological basis of algae cells and therelationship between lipid and EPA were analyzed. When the algae cells were cultured for6days, the lipid content increased to23.07%gradually, the protein content and the percentageof EPA decreased to31.31%and8.13%, respectively. At the same time, the proportion ofC16:1and C18:1increased to23.04%and12.15%, respectively. By the method of TEM, thedecline in the proportion of chloroplast and the serious degradation of thylakoid wereobserved. A significant accumulation of lipid body also occurred in algae cell.Intermediate metabolites were added to promote the accumulation of lipid and EPA. Itwas found that the acetic acid and propanediol could contribute to the syntheses of lipid.When supplemented with the acetic acid and propanediol, the lipid content increased by17.2%and31.5%respectively compared with the control groups. It was also found that thesalinity stress with adding of acetic acid and propanediol was more advantageous to the lipidaccumulation. The lipid content increased to43.38%which was1.6times than that of thecontrol group. When cultured with the ethanol, the algae growth was inhibited while theaccumulation of EPA was promoted. When the ethanol concentration was4%, the proportion of EPA was highest which was improved by64.5%compared to the control group.