Study On Haematococcus Pluvialis Transformational Conditions And Astaxanthin Extraction

Astaxanthin has powerful antioxidantive activities and the function ofscavenging Free Radicals. It has wide applications in the aquaculture, food, medicineetc. At present, Haematococcus pluvialis is regarded as the best biological source ofastaxanthin. As is well-known Haematococcus pluvialis has the obvious life cycle.Through the environmental stress, algal cells switch from green swimming to thickwall spores and astaxanthin is produced. In this thesis, firstly, with astaxanthin contentfor index, by the CCD experiment, the highest production of astaxanthin was11.05mg/L, when the concentration of sodium acetate was1.46g·L1, but the productionless than dextrose culture medium. Results showed that the optimal culture andtransformational medium was decided as: glucose3g/L, KNO₃0.3g/L, KH₂PO₄0.03g/L, MgSO₄·7H₂O0.2g/L, CaCl₂0.08g/L, the production of astaxanthin reached to16.98±0.68mg/L, enhanced about53.67%. Then, by the CCD experiment, thehighest production of astaxanthin was33.80±1.56mg·L1,enhanced about99%, whenthe transformation light intensity7203.95lx, transformation temperature29.92℃,transformation time10.6days and pH10. Finally, the effects of liquid mediumvolume and transformation time on the production of astaxanthin were investigated.The experimental results demonstrated that increasing the relative surface area caneffectively improve the efficiency of transformation and the production of astaxanthin.By the CCD experiment, the highest producion of astaxanthin was36.91mg/L,enhanced about10%, when the liquid medium volume was49.49mL and thetransformation time was10.36days.The effects of red light on the growth of Haematococcus pluvialis and blue lighton the cells transformation were further investigated. It was concluded that thecombination of continuous spectra （1600-1700lx） and red light （800-850lx） can obtain a good balance of speed and efficiency, the cell dry weight reached0.72±0.03g/L, enhanced about6.3%. The combination of continuous spectra（6000-7000lx） andblue light（2000lx） was better for transformation than white light, the production ofastaxanthin reached38.44±1.64mg/L, enhanced about14.9%. Blue light caneffectively improve the efficiency of transformation, applying to industrial productioncan not only improve the productivity but also save cost.In this experiment, succeeding preservation （liquid-culture-medium method andsolid-liquid-culture-medium method）and immobilization storage were used topreserve Haematococcus pluvialis in low temperature and dim lights. According tothe survival rate, the specific growth rate and the content of chlorophyll-a, the resultsshowed the descending sequence of preservation methods is: liquid-culture-mediummethod, solid-liquid-culture-medium method and immobilization storage. After fourmonths, the survival rate reached93.84±3.88%, the specific growth rate of the algaereached0.4648±0.0021d^-1, and the content of chlorophyll-a decreased by11.91%inliquid-culture-medium method. So succeeding preservation is suitable to be used inresearch for its simplicity and high survival rates, while immobilization storage issuitable to be used for relative long term preservation. After finished the experiment,DMSO and glycerol were used as protective agent to preserve Haematococcuspluvialis in-20℃a nd-40℃. When10%DMSO was used as protective agent,microalgae was preserved in-20℃, the survival rate and the specific growth ratewere42.27±1.88%and0.1550±0.0063d^-1. When20%DMSO was used as protectiveagent, microalgae was preserved in-40℃, the survival rate and the specific growthrate were49.05±2.03%and0.1750±0.0065d^-1. This method was not easy to dye thebacteria, algae was preserved in this way not easy to degenerate.By the preliminary studies on astaxanthin extraction from the chlamydospore,the optimal breaking conditions were: high speed disperse homogenizer （50Hz,23000rpm） acted6minutes. The wall-broken rate was about96±3.93%, and it was a goodmethod to break the wall. As to the extracting astaxanthin from Haematococcuspluvialis directly, the results suggested that the mixture of DMSO: acetone（4:1） wasproper to exact Astaxanthin from Haematococcus pluvialis. The production of astaxanthin was34.04±1.53mg/L, in comparision with DMSO, the production wasincreased by4.4%. The crude astaxanthin need further purified, and more research isneeded.