Optimization And Regulation Of Growth And Astaxanthin Accumulation In Haematococcus Pluvialis

Astaxanthin(3,3'-dihydroxy-?,?-carotene-4,4'-dione),a liposoluble red ketocarotenoid,has potential clinical applications in the poultry industry,aquaculture,cosmetics,nutraceutical and medicine because of its powerful antioxidant capacity.Haematococcus pluvialis,a unicellular Chlorophyceae,is widely known as an important natural source for natural astaxanthin and it has been cultivated at mass-scale in industry prodution.However,the method to increase cell density and astaxanthin content need to be further studied.Understanding the essence of interactions between photosynthetic performance and changing actinic irradiances in different phases would provide some new insights into large-scale industrial production.Far less attention has been paid to the interrelationship between other metabolic pathways and astaxanthin accumulation in H.pluvialis.The alga Haematococcus pluvialis(strain H6)was used in this study.And different carbon sources were added in different stages by combining two-stage cultivation and adding carbon source.The effects of additional carbon source on the growth of Haematococcus pluvialis were studied by adding 1 g/L sodium acetate,glucose,sodium bicarbonate at initial stage of culturing.Moreover,mixotrophic conditions were used in the astaxanthin accumulation stage by adding 1 g/L sodium acetate and glucose.The basic parameters of cell density,specific growth rate,dry weight,biomass productivity and pigment content were studied,and physiological and biochemical parameters were preliminary analysed.The effects of additional carbon source in different stages involves many synthesis and metabolis,which need to be further studies,and then extended to large-scale industrial production.Besides,this study clarifies the short-term variability in the photosynthetic activity of different phases in H.pluvialis in closed photobioreactors outdoors via rapid light curves(RLCs).The cell morphological changes and pigment contents with increased time were analysed in this process.Furthermore,this thesis examines the relationship between photorespiration and astaxanthin accumulation.The biomass,pigment contents,physiological and biochemical parameters were studied by adding specific inhibitor of photorespiration,carboxymethoxylamine(CM).Understanding the basic interactions between photorespiration and astaxanthin accumulation would provide some new insights into mechanisms impacting astaxanthin production in H.pluvialis.The main results were as follows:1.At the early stages of culturing,additional carbon source increased biomass productivity significantly,in which sodium acetate had the best effect,with 2.11 times than control cells on 0-2 d.On 1 d of incubation,additional carbon source increased total photosynthetic O2 evolution capacity.Sodium acetate incereased specific growth rate(0.89 day-1)only in the initial stage of culture(0-2 d)with the maximum respiratory O2 consumption capacity,while the photosynthetic parameters of(37)PSII,Fv'/Fm',qP and NPQ were much lower than.Glucose incereased specific growth rate(0.17 day-1)in the late culture(after 4 d),with 1.7 times than control cells.Sodium bicarbonate was not conducive to the increase of specific growth rate,while the photosynthetic parameters of(37)PSII,Fv'/Fm',qP and NPQ were much higher than adding sodium acetate or glucose.These results suggest that sodium acetate increased cell growth on 1-2d when heterotrophy played an important role,and improved photosynthesis on 3-4 d to maintain cell growth and biomass accumulation.2.On the astaxanthin accumulation stage,the effects of additional carbon source on cell density were similar to the results of initial stage of culturing.And additional carbon source increased dry weight(2.27 times than control cells)and astaxanthin contents(2.95 times than control cells)significantly,in which sodium acetate had the best effect,with inhibition of photosynthesis accompanied by the decrease of(37)PSII,Fv'/Fm',qP and NPQ.The results indicate that heterotrophy played an important role in astaxanthin accumulation in H.pluvialis.The effects of additional carbon source in different stages involves many synthesis and metabolis,which need to be further studies,and then extended to large-scale industrial production.3.Astaxanthin accumulation was significant increased with the increased time of incubation in outdoor mass-scale cultivation of H.pluvialis,from 2.72 mg/L(0 d)to 39.73 mg/L(12 d).And With the increased astaxanthin contents,(37)PSII decreased 78.9%,80.9%,79.9%,86.4%,90.1% and 89.6% from 0 to 1600 ?mol m-2 s-1,respectively.The effective quantum yield of photosystem II((37)PSII)decreased with the increased photosynthetically active radiation(PAR)because of increased Fs and decreased Fm'.The photosynthetic efficiency(?)of rapid light curves was significant higher when astaxanthin started to accumulate(4 d)and astaxanthin covered the whole cell(12 d).While,there was no significant difference of photoinhibition parameter(?),the maximum of relative electron transport rate(rETRm)and the minimum light saturation irradiance(IK)with increased astaxanthin in H.pluvialis.4.In H.pluvialis,the accumulation of astaxanthin was reduced(decreased 45.45% than control cells)significantly when the photorespiratory pathway was inhibited by CM.Inhibition of the photorespiratory pathway did not change DW accumulation,chlorophyll content or OJIP transients during the incubation.In contrast,the inhibition of the photorespiratory pathway decreased the photochemical activity of PSII(decreased 34.56% than control cells)and total photosynthetic O2 evolution capacity(decreased 42.69% than control cells)significantly.The suppression of photorespiration correlated with the decrease of astaxanthin content.Taken together,these results suggest that the photorespiratory pathway plays an important role in astaxanthin accumulation in H.pluvialis.