Study On The Cell Growth And Lipid Accumulation Of Chlorella Sp. Intensified By Magnetic Fe₃O₄ Nanoparticles

Microalgae are very important bioresources,which play a key role in the matter cycle and energy flow of ecosystems and have high ecological values.In addition,microalgae possess a lot of advantages,including wide distribution,numerous species,large quantity,rapid reproduction,and rich nutritional value.Therefore,they have been widely used in different areas such as food,feed,medicine,energy,and environmental protection.The technology of microalgal cultivation is one of the key factors affecting the industrialization of microalgae production.Increasing biomass and target product content is one of the effective ways to achieve the large-scale production of microalgae.In this study,the autotrophic liquid culture of Chlorella sp.UJ-3 was used as the research system,and magnetic Fe₃O₄ nanoparticles（nano-Fe₃O₄）and alternating magnetic field were tested as the methods of process intensification.The effect of nano-Fe₃O₄on Chlorella sp.UJ-3 growth and lipid metabolism was studied,and the regulation mechanism of nano-Fe₃O₄ on the metabolism of Chlorella sp.UJ-3 was investigated by transcriptome analysis.The synergistic effect of alternating magnetic field and nano-Fe₃O₄ on cell growth and lipid accumulation was also explored the culture of Chlorella sp.UJ-3.The main conclusions are as follows:（1）The effects of nano-Fe₃O₄ at different concentrations on biomass,lipid production and fatty acid composition were studied the cultivation of Chlorella sp.UJ-3.As a process intensification method,nanoparticles increased biomass and lipid production during the microalgae culture under tested conditions.After the optimization of nano-Fe₃O₄ concentration,the algal biomass was increased by 52.9%at 20 mg/L and the total lipid content was improved by 42.9%at 100 mg/L.The enhancement in biomass could be attributed to the increase of trace element and gas-liquid mass transfer rate.The enhanced accumulation of lipids may be due to oxidative stress induced by nano-Fe₃O₄.Based on the above results,a new strategy was established.In this case,algae were cultured at low nano-Fe₃O₄ concentration of 20 mg/L for better cell growth,and then the concentration of nano-Fe₃O₄ was increased to 100 mg/L in logarithmic growth phase for higher lipid accumulation.Compared to the untreated algal cells,algal biomass and total lipid production under this strategy increased by 50%and 108.7%,respectively.The addition of nano-Fe₃O₄ caused an increase in the content of ROS and the activity of related antioxidant enzymes in algal cells.（2）The differential expression of transcriptome genes and the expression level of key enzymes genes in important metabolic pathways were studied based on transcriptome analysis under different nano-Fe₃O₄ treatments.The results showed that the addition of nano-Fe₃O₄promoted the synthesis and desaturation of fatty acids,and weakened theβ-oxidation process of fatty acid in microalgal cells.The expression levels of most enzyme genes in the glycolytic pathway were down-regulated,while the expression levels of multiple enzyme genes in the pentose phosphate pathway were up-regulated.It may be due to the response of algal cells to exotic stress resulted from nano-Fe₃O₄ addition,which regulated the glucose decomposition from glycolysis to the pentose phosphate pathway.In addition,the gene expression of enzymes related to Calvin cycle and C₄ pathway varied under the nano-Fe₃O₄ treatment.The carbon fixation of algal cells was mainly based on Calvin cycle,and C₄ pathway could also exist,which increased the collection and concentration of CO₂,and provided sufficient inorganic carbon to sustain energy for other metabolism in algal cells.The gene expression of superoxide dismutase（SOD）and catalase（CAT）,which are the key enzymes of hydrogen peroxide metabolism in the antioxidant system,was up-regulated under the nano-Fe₃O₄ treatment.It proved the protection effect of antioxidant enzyme system from the oxidative stress in algal cells.（3）The synergetic effect of alternating magnetic field and nano-Fe₃O₄ on the cell growth and lipid metabolism of Chlorella sp.UJ-3 were studied.The results showed that the growth of microalgal cells was promoted with the treatment of magnetic field.The biomass of microalgal cells treated by magnetic field alone increased by 38.0%compared to the untreated control.Among the tested magnetic field at different frequencies,the alternating magnetic field at 200Hz exhibited the best synergetic effect with nano-Fe₃O₄,where the maximal biomass increased by 64.5%and the highest total lipid production（0.65 g/L）was enhanced by 170.8%compared to the untreated control.The highest concentrations of chlorophyll a and chlorophyll b were obtained in the algal cells treated with magnetic field alone.The photosynthesis of algal cells treated with nano-Fe₃O₄ was attenuated due to the shading effect of nano-Fe₃O₄.As an antioxidant,carotenoids possessed an increased concentration in the algal cells of the treated groups.However,alternating magnetic field with high frequency of 400 Hz brought an inhibition in carotenoids synthesis.Under magnetic field and nano-Fe₃O₄ treatment,algal cells can regulate their antioxidant defense system,and the activities of related antioxidant enzymes increased to resist the increased ROS induced by the magnetic field and nano-Fe₃O₄ in algal cells to protect cells effectively.