Research On The Circadian Rhythm Response Of Chlorella And Its Lipid Production Technology

Microalgae were widely applied in energy,environment treatment,as well as food and medicine industry.However,large scale cultivation,increase lipid productivity and collection of microalgae were the major problems restricting the development of algae application.Chlorella cells usually grow photosynthetically during the day,DNA replication and fission occur during the night,and this circadian cycles is the mechanism by which algae divide and multiply.After optimization of culture conditions and culture stages to ensure algae growth,division behavior and circadian rhythms,adaptation is the basis of culture optimization design.After studying the correlations between chlorella biomass,biochemical components accumulation and circadian rhythm,optimized the lipid production technology of chlorella by the two-stage culture method through the optimized combination of culture conditions and culture stages,combined with the advantages of self-flocculation of microalgae.(1)To study the effect of autotrophic and mixotrophic cultivation on the growth,division and biochemical components accumulation of Chlorella pyrenoidosa,and then to explore the response mechanism of Chlorella pyrenoidosa on circadian rhythm under artificial cultivation and optimize the corresponding technology.Glucose had a significant growth promoting effect on Chlorella pyrenoidosa.The growth and proliferation as well as the biochemical components accumulation of Chlorella pyrenoidosa were regulated by circadian rhythm,the algae could grow autotrophically during the day when exposed to light and undergo cell proliferation in dark.The machanism for increasing the biomass of Chlorella pyrenoidosa by mixotrophic cultivation were shortening its silent period,promoting the growth of algae cells in circadian rhythm and increasing the proportion of larger cells,which ensures that cells have passed the commitment point of cell-cycle,with the effect during light period was especially obvious.The protein and starch contents of Chlorella pyrenoidosa showed regular circadian rhythm,their best harvest time were 8 and 12 hours after in light,respectively.(2)The growth and lipid content of Chlorella pyrenoidosa at different concentrations of nitrogen stress were studied.The low nitrogen group with 6 mmol/L nitrogen concentration showed rapid growth effect on the first 3 days after the conversion.When the algae cells were harvested on the 4th day,the 6 mmol/L low-nitrogen group had the highest lipid content.Considering the concentration of algal cells and lipid production,the nitrogen stress threshold of Chlorella pyrenoidosa was 6 mmol/L.Under low nitrogen stress,the intracellular extracellular protein and starch of algal cells decreased significantly.Compared with the control,the synthesis of protein and starch at low nitrogen level was inhibited.In addition,both autotrophic and mixotrophic algae cells enhanced their flocculation ability under low nitrogen and flocculated better with the incubation time.(3)Based on the self-flocculation characteristics of algae cells,aiming at improving the lipid production of Parachlorella kessleri,the optimization of the first-stage glucose-culturing and the second-stage elemental stress treatment of two-stage culture method were carried out.After 10 days,the concentration of algae cells and the dry weight reached 4.74E+07 cell/mL and 3.54 g/L.In the two-stage culture,the effect of three kinds of elemental constraints and low-glucose and low-nitrogen co-treatment on the lipid production of Parachlorella kessleri was studied.The results showed that the low-glucose treatment was the best,lipid production reached 242.64 mg/L/d,suitable for harvesting algae cells.The flocculation rates of first-stage and second-stage low-glucose treatments reached 96.11%and 93.21%,respectively,which saved the cost of harvesting and facilitated the two-stage cultivation and efficiency enhancement.(4)Parachlorella kessleri was used as the research object to investigate the circadian rhythms of algae cells under carbon and nitrogen constraints.The results showed that the circadian rhythm response under carbon signal stimulus conditions was significantly changed.Algae cell division and proliferation to self-support dark period-based conversion to light-based trends.After the transfer of seed cells under mixotrophic cultivation,the circadian rhythm of algae began to reverse again and gradually returned to the dominant trend of proliferation in dark phase.The response rate of low-glucose and low-nitrogen and circadian rhythms moved rapidly,which indicated that Parachlorella kessleri responded more rapidly to the changes of low glucose and low nitrogen environment.Compared with the control,the concentration of algae in 2 d was the highest,the concentration of low glucose and low nitrogen was the highest,which indicated that the growth of algae cells accelerated the synergism of circadian rhythms and environmental factors and accelerated the proliferation of algae cells.