(751f) Study of a Two- Stage Culture of Docosahexaenoic Acid Producing Marine Algae Schizochytrium Limacinum SR21 with Shifting Dissolved Oxygen Level

Docosahexaenoic acid (DHA, C22:6), an important omega-3 fatty acid, is known as an essential nutrient for the development of brain and eye for the fetal and infant. Production of DHA from marine microalgae is taking more and more important role for providing the increasing demand of DHA, since fish oil will be not sustainable in the future, and there is potential pollutant from the environment. To decrease the production cost, cull potato and crude glycerol from biodiesel industry has been used to culture S. limacinum SR21 to produce DHA. However, difficulty has been encountered when pursuing a high density biomass production. Reviewing the life cycle of S. limacinum SR21 found that this alga exists as two statuses: zoospores and vegetative cells. A phenomena was observed that the zoospores, swimming rapidly in the culture broth, like to gathering at the surface of air-liquid surface. Based on this, a hypothesis was come up that the limitation of biomass production could be caused by the low oxygen concentration in the culture and produced too less cells in this condition. This was proved in this work, since the culture of DO controlled at 50% produced 181 million cells/ml , but the culture with 10% DO produced a cell density only 98.4 million cells/ml. Even lower density of 22.5 million cells/ml was produced in the culture with a fixed agitation rate of 150 rpm. It was also found that the size of new generated vegetative cells and lipid accumulated vegetative cells in the later phase of the culture is significantly different, due to the lipid accumulation inside the cell. The experiments also showed that high oxygen as 50% led to pH decreasing and be harmful to the lipid accumulation, and low oxygen concentration was obligatory for lipid accumulating. Thus, the culture of Schizochytrium limacnum SR21 can be divided into two stages: (1) cell number increasing stage: cell reproduction and cell number increasing with little increase in the size and weight of each cell and (2) cell size increasing stage: cells stopped reproduction but cell size enlarged due to lipids accumulation. A shifting oxygen control strategy was used to produce more cells with dissolved oxygen controlled at 50%, and then providing a limiting oxygen concentration for fatty acid accumulation. With this protocol, the production of algae biomass and DHA was improved to 37.9 g/L and 6.56 g/L, respectively. This work revealed an important feature of oleaginous microorganisms that high oxygen concentration is demanded for the reproduction while low or limiting oxygen concentration is obligatory for the lipid accumulation. Shifting the oxygen level is necessary to obtain high production of biomass and lipids, since the optimal condition for lipid accumulation conflicts with cell proliferation if one stage culture was used. This two-stage culture process could be used not only in the omega-3 fatty acids production process, but also more widely, in all single cell oil (SCO) producing processes, including biodiesel production from algae, which is the largest potential of SCO application.