A Study On The Systhesis Of Astaxanthin And The Analysis Of Its Geometric Isomers In Haematococcus Pluvialis

Astaxanthin's powerful antioxidant properties endow it many important physiologic functions,such as,improving animal immunity,being a foe of free radicals,precventing and treating of cancer,preventing Alzheimer's disease,coloring agent and antioxidants.Therefore,the natural astaxanthin has a widespread application and prospect on auaculture and poultry,health-food,osmetics and so on.Furthermore,Haematococcus pluvialis is regard as the best biological source of astaxanthin.Natural astaxanthin mostly was trans-astaxanthin,but exists a little cis-astaxanthin,tran-astaxanthin and cis-astaxanthin have differences on physiologic.It will be very significant that the separation and analysis of the astaxanthin isomers,and explore the reason of astaxanthin isomerization.The work in this dissertation is as follows:1,to explore the ability of astaxanthin acculumation and the differential characteristic of cis/trans geometrical isomers during different algal strains,different Haematococcus pluvialis stains H₀,H₂,H₃,H₆ and T₂ are taken as research objects and the research is focus on that the cell growth,the variations of photosynthetic rate and respiratory rate,and the content of chlorophyll and astaxanthin,and the ratio of astaxanthin isomers.Results show that strainT₂ was significantly better than the other four strains?H₀,H₂,H₃,H₆?on the cell growth,photosynthesis,respiration rate,content of chlorophyll,carotenoid and algal cells resistance to adversity and astaxanthin content and so on.But five algal strains have no significant difference?P > 0.05?on the ratio of astaxanthin isomer.2,to explore the change of cells morphology and internal composition during different stages of Haematococcus pluvialis?cell transformation phase and astaxanthin accumulation phase?,stain H₆ are taken as research objects and the research is focus on that the cell growth,the variations of photosynthetic rate and respiratory rate,and the content of chlorophyll and astaxanthin,and the ratio of astaxanthin isomers and the content of protein.Results show that cells have a significantly change on the content of pigment and protein during cell transformation phase?0-3days?,mainly was the sharply decrease of chlorophyll content and quickly increase of carotenoids and astaxanthin,and fluctuation changes of isomers ratio.All above changes was slowly on the astaxanthin accumulation phase?3-15days?,mainly was the increase of astaxanthin content.3,to explore the content of astaxanthin geometric isomers,this article use the acetone grinding method under low temperature to prevent the isomerization of astaxanthin,by cholesterol esterase enzyme to get free astaxanthin,and use high-performance liquid chromatography to analyze the astaxanthin isomers and other carotenoids from the pigment's extract of Haematococcus pluvialis.Results show that different strains?H₀,H₂,H₃,H₆ and T₂?have no significant difference on the ratio of astaxanthin isomer under same culture conditions.The ratios of astaxanthin isomer have a significant difference on different cell phases in Haematococcus pluvialis,and the ratio have no difference on the late astaxanthin accumulation phase.4,to explore the reason of astaxanthin accumulation and geometric isomers,western blot technique is used to analyse the protein expression about catalytic the cis/trans carbon atoms.Results show that FFPS and GGPS have a higher expression on cell transformation phase and lower on astaxanthin accumulation phase.Over the whole culture time,the content of FFPS and GGPS decrease gradually.But the above proteins are trans-prenyl enzyme,there is still need further research on whether Haematococcus pluvialis exists cis-prenyl enzyme,and how the cis-prenyl and trans-prenyl enzymes interact with each other in Haematococcus pluvialis.5,in this dissertation,the pigment content and cell morphological and protein content are combined,expounds the relationship between light and pigment,ROS and astaxanthin,analyzes the mechanism of astaxanthin accumulation under stress conditions.Results show that: high light destroys chlorophyll,which reduce the ability of photosynthesis causes the green color decline.In addition,the breaken of chlorophyll causes the decline of related protein D1,PsaA and RBCL content,which is mediated the electron transport chain is damaged,further causing the generation of ROS and residual electron.Algal cell enhance the ability of chloroplast and mitochondrial respiratory to eliminate ROS.But the cell's respiration ability is not enough to scavenging ROS produced by light inhibition.Speculated that the algal cells through synthesize a large number of astaxanthin and other carotenoids to cope with stress conditions.6,the indexes such as carotenoid/chlorophyll,astaxanthin/carotenoid and the ratio of the isomers of astaxanthin are used as indicators of astaxanthin quality.Results show that: with the culture time increase,the ratios of carotenoid/chlorophyll and astaxanthin/carotenoid gradually are increased,and the proportion of astaxanthin isomers tends to be stable.The addition of the above indicators can help to regulate the quality of astaxanthin and astaxanthin market.