Effects Of 1-aminocyclopropane-1-carboxylic Acid And Fulvic Acid On The Accumulation Of Carotenoids Components In Dunaliella Bardawil

Microalgae carotenoids having a variety of significant biological activities play an important positive role in human health.Due to the potential carcinogenicity,hepatotoxicity and nephrotoxicity of synthetic pigments,the world's demand for natural carotenoids is increasing.Microalgae are natural resources for carotenoids production.Dunaliella bardawil was the research subject to investigate the effects of seven chemicals [fulvic acid(FA),betaine,thiamine,choline chloride,tricine,1-aminocyclopropane-1-carboxylic acid(ACC)and spermidine] on the biomass,pigment content and caroteniods accumulation of D.bardawil.And the concentrations of ACC and FA were optimized to enhance carotenoid yield.To further increase the yield of caroteniod,the ACC-FA two-stage joint cultivation strategy was adopted.The PCR technology was used to explore the effects of ACC,FA and ACC-FA treatment on the gene expression of key enzymes in caroteniod anabolic metabolism.From the preliminary screening of seven chemical substances,the compounds that can significantly increase the yield of caroteniods in D.bardawil are ACC and FA.The optimization results of ACC concentration are consistent with the preliminary screening results.ACC can significantly promote the biomass and at the same time significantly promote carotenoids(especially in ?-carotene)such as lutein,?-carotene and ?-carotene in D.bardawil in a time-dependent manner and the promotion effect is more significant in the stable phase.Different concentrations of ACC treatments had inconsistent responses to the production of caroteniod components in D.bardawil cells.11 m M ACC is conducive to the accumulation of ?-carotene and 15 m M ACC is conducive to the accumulation of ?-carotene.Moreover,ACC treatment can change the content of caroteniod components in D.bardawil cells(?-carotene>?-carotene),so that the content of ?-carotene is higher than that of?-carotene.The results of laser confocal observation of lipid distribution indicate that ACC may realize the co-production of carotenoids and lipid.The results of FA concentration optimization show that FA inhibited the biomass of D.bardawil and has no obvious effect on the accumulation of lutein in the logarithmic phase.In the stationary phase,it is more conducive to 600 mg/L FA to promote the accumulation of caroteniods in algal cells,especially in lutein;but the promoting effect on ?-carotene and?-carotene can be seen in the logarithmic phase.The laser confocal results further prove that FA enhances lipid accumulation in a concentration-dependent manner,which is consistent with the inhibition of biomass and various caroteniods accumulation in concentration-dependent.The results of the ACC-FA two-stage joint cultivation strategy showed that the first stage ACC treatment alone can increase the biomass and significantly promote the accumulation of caroteniods.In the second stage,compared with the treatment with ACC alone,the addition of FA could reduce the accumulation of lutein in logarithmic phase and promote the accumulation of lutein in stable phase,and further significantly increase the accumulation of?-carotene and ?-carotene.The effect of ACC alone in improving each caroteniod is stronger than that of FA alone.The 11 m M ACC-500 mg/L FA combined treatment group had the best promotion effect on lutein,?-carotene and ?-carotene in algae cells.RT-PCR resulted showed that ACC or FA can promote the accumulation of lutein and ?,?-carotene in D.bardawil cells,which was related to the up-regulation of Db LYCB,Db LYCE,Db BCH,Db CYPA and Db CYPC gene expression.The reduction of zeaxanthin may be related to the dissipation of excessive excitation energy;ACC or FA regulates other metabolic pathways related to the synthesis of zeaxanthin,and the high activity and bias of the Db BCH and Db CYPA enzyme genes in the ?-carotene synthesis lutein pathway.ACC-FA combined treatment further promoted the accumulation of ?,?-carotene than ACC treatment alone,which is closely related to its further up-regulation of the expression of Db LYCB and Db LYCE,and down-regulation of the expression of Db BCH,Db CYPA and Db CYPC.