Protective Effects Of Astaxanthin From Haematococcus Pluvialis On The Zebrafish Embryos Induced By The Stress Of Microcystins

Microcystins(MCs)was one of the most widely distributed and toxic substances in aquatic ecosystems occurring cyanobacterial blooms.MCs could inhibit the growth of aquatic organisms and would bioaccumuate in food chain,thus indirectly threaten human health.Haematococcus pluvialis is single-celled green alga that is widely present in the natural environment.Due to its rich astaxanthin with strong antioxidant capacity,it has attracted more and more attention.Under unfavorable conditions,red dormancy cysts were formed and a large amount of astaxanthin was accumulated in the cell.Astaxanthin has been widely used in food,medicine,cosmetics,aquaculture and other industries due to its many outstanding physiological characteristics,such as improving resistance,preventing cell damage,inhibiting tumors,and eliminating free radicals.In this study,we explored the effects of different environmental factors on the growth and accumulation of astaxanthin in H.pluvialis and compared the algal cell disruption efficiency.We successfully obtained natural astaxanthin,and then exposed the zebrafish embryos to acute toxicity through microcystins.Astaxanthin was evaluated for its antioxidant effects.The specific results are as follows:(1)The present study discussedthe effects of different light intensities(0,2000,4000,6000,8000 lux),different temperatures(18,22,26,30?),different nitrogen concentration(gradient control of nitrogen element content in BG₁₁ medium,that was 0 N,1/4 N,2/4 N,3/4 N and all N)on the growth and accumulation of astaxanthin in H.pluvialis.It was suggested that the best growth condition for H.pluvialiswas 4000 lux,18?,and 2/4 N content,where as the highest astaxanthin accumulation occurred at 8000 lux,30?,and 0 N content.The study compared the break efficiency of different methods on algal cell,that was ultrasonic crusher,homogenizer,tissue cell crusher,and liquid nitrogen repeatedly freeze-thaw to break algal cell.It was concluded that the ultrasonic crusher's crushing was significantly more effective than other methods,and the optimal conditions of ultrasonic crushing were 350 W,5 min,frequency of 1 time.(2)The research also illustrated the effects of microcystins with different concentration(0,10,20,30,40,50?g/L)on the development of zebrafish embryos.The results showed that MC-LR concentration and embryo developmental toxicity were in dose-effect relationship.MC-LR could increase the mortality and deformity rate and reduce the hatching rate during zebrafish embryo development.The result was statistically significant at a MC-LR concentration of 50?g/L(p<0.05).The results also showed that MC-LR had no significant effects on body lengthof zebrafish juveniles.The study also discussed the combined effects of two concentrations of astaxanthin(10,100?g/L)and microcystin on the development of zebrafish embryo.The results showed that compared with the MC-LR alone exposure group,the mortality and deformity rates decreased significantly in the combined exposure group.The results of the combined exposure of high concentration astaxanthin(100?g/L)were significantly different,indicating that astaxanthin could alleviate embryonic developmental toxicity caused by MC-LR,which was positively correlated with astaxanthin concentration.However,no significant effects of astaxanthin on hatchability and body length were found.(3)The results of microcystin in oxidative stress showed that MC-LR could also induce oxidative stress and cause oxidative damage in zebrafish bodies at different concentrations(0,10,20,30,40,50?g/L).After MC-LR exposed zebrafish embryos at 120 hpf,the ROS and MDA content in juveniles increased,and GSH,SOD,and T-AOC decreased significantly.The higher the MC-LR concentration was,the stronger the oxidative stress response was detected.The results of each group were significantly different at a concentration of 50?g/L(p<0.05).Astaxanthin could significantly improve the oxidative stress response and oxidative damage in zebrafish caused by microcystins.In the experimental group of astaxanthin combined with MC-LR at different concentrations(10,100?g/L),compared with the MC-LR exposure group,after adding astaxanthin,the ROS and MDA contents of juvenile fish decreased,mean while,GSH,SOD,T-AOC increased significantly,and the higher the concentration of astaxanthin was,the more significant effects on the improvement of oxidative stress was.The results of each group were significantly different in the group exposed to the combination of high concentration astaxanthin(100?g/L)(p<0.05).