Research On The Stress Induced Condition And Pilot Test Of Astaxanthin-rich Haematococcus Pluvialis

Astaxanthin is a kind of valuable pigment, due to its high antioxidant and stain activity, it has be used in aquaculture, cosmetic, nutraceutical and pharmaceutical industries widely. Haematococcus pluvialis is the best natural source for astaxanthin. It has two distinct phases in its life cycle, viz. a green-motile stage in favorable conditions and a red non-motile resting stage called a cyst in unfavorable conditions, astaxanthin accumulates in cytoplasmic lipid vesicles in its red stage. Haematococcus pluvialis can grow rely on photosynthesis as plant, in addition, it is able to grow heterotrophically and mixotrophically on organic carbon source.Here, triangular flask and flat-or tubular-photobioreactor in outdoor were employed to study Haematococcus pluvialis from the following aspects:(1) Influence of experimental conditions on the accumulation of astaxanthin in Haematococcus pluvialis in lab. Results showed that Haematococcus pluvialis accumulated more astaxanthin in nitrogen-free condition than in other nitrogen starvation conditions, the concentration of astaxanthin could reach13.92±0.24mg/L. phosphate-free condition was the best phosphate starvation level for Haematococcus pluvialis to accumulate astaxanthin, the concentration of astaxanthin could reach17.52±1.43mg/L. The content of astaxanthin in Haematococcus pluvialis would be enhanced with the increase of NaCl concentration, but when the NaCl was excessive, it inhibited the growth of Haematococcus pluvialis, further more, it would kill cells, the optimum NaCl concentration to obtain a high astaxanthin concentration was0.6%, the concentration of astaxanthin could reach13.86±0.83mg/L. High light increased the content of astaxanthin in Haematococcus pluvialis, however when the light intensity exceeded8000Lux, it inhibited the growth of Haematococcus pluvialis as well as damaged its ability to accumulate astaxanthin, the most suitable light intensity for Haematococcus pluvialis to accumulate astaxanthin was between6000Lux-8000Lux, the concentration of astaxanthin could reach22.98±0.11mg/L in nitrogen-free condition. The change of culture volume affected the accumulation of astaxanthin in Haematococcus pluvialis as it changed the light path and gas exchange, the concentration of astaxanthin could reach20.29±1.01mg/L in100mL culture volume (500mL triangle flask). When Haematococcus pluvialis grown in the condition composited the favorable aspects stated above, the concentration of astaxanthin could reach44.58±2.68mg/L.(2) Haematococcus pluvialis cultivated in composited stress condition was selected for carotenogenic genes expression studies. Results showed that the expression of psy, pds, lcy bkt, chy were enhanced during the12days except the expression of lcy deceased slightly on day10compared to those without stress-induced. The highest expression of psy, pds, lcy, chy appeared on second day, were3.5-,2.5-,5.9-,2.4-fold higher than those without stress-induced respectively. The highest expression of bkt was found to be3-fold higher than that without stress-induced and appeared on8th day.(3) Flat-and tubular-photobioreactor were used to culture Haematococcus pluvialis in medium scale. In green stage, Haematococcus pluvialis grown in indoor or outdoor condition, the biomass reached0.88g/L and1.04g/L respectively, biomass productivity before the algae entered stable phase were0.0485gL-1d-1and0.0595gL-1d-1respectively. In red stage, Haematococcus pluvialis was induced in flat-photobioreactor in semi-outdoor condition, the biomass of1.17g/L and the content of astaxanthin of1.49%were obtained, the concentration and productivity of astaxanthin were17.49mg/L and1.05mgL-1d-1respectively.(4) Influence of initial sodium acetate concentration on the growth of Haematococcus pluvialis in green stage and the accumulation of astaxanthin in red stage. Results showed that the optimum range of initial sodium acetate concentration to obtain a high biomass in light was between1g/L and1.5g/L, cultured in the initial sodium acetate concentration of1.5g/L obtained highest biomass in dark. When the algae was induced in stress condition after grew in mixotrophic conditions, the biomass was almost the same as that grew in autotrophic condition, but the content of astaxanthin was lower. In stress-induced process,the sodium acetate did not affect the content of astaxanthin in Haematococcus pluvialis and increased the biomass when the initial concentration was below2g/L. When initial sodium acetate concentration exceeded2g/L, the content of astaxanthin increased slightly, but the biomass of Haematococcus pluvialis decreased.