| Dunaliella salina is a single-cell prokaryotic plant withoutcell wall.It is also one of the few species of algae that has been industrialized so far.This microalga is rich in?-carotene,oil and pigment.Beta-carotene is a natural pigment with the functions of anti-lipid peroxidation,thereby improving human immunity and curing certain chronic diseases.Meanwhile,it is also an important compound essential to the human body.Beta-carotene is used in health care,medicine,cosmetics,foods and other fields with broad application prospects.This study usedtwo industrial Dunaliella salina strains of CCAP 19/18 and B01 as research materials.Firstly,the cultivation condition and medium composition were optimized for Dunaliella salina;secondly,the influence of light intensity and carbon source on the growth of Dunaliella salina was further studied;The growth of Dunaliella salina and their accumulation of?-carotene in the photosynthetic reactor were explored through environmental inhibition and metabolic inhibitors.The main results are as follows:(1)Dunaliella salina CCAP 19/18 were cultivated with different culturing conditions and media.The research showed that the ATCC media is the optimal medium for the growth of CCAP 19/18 byachieving maximal cell concentration of 4.35×10~6/m L.Using ATCC medium as the substrate,stirring speed of 200 rpm/min and air flow of 300 m L/min showed the fastest growth ratein Dunaliella salina CCAP 19/18.Under this condition,the chroophyll in microalga can reach up to 18.36?g/10~6cells,which is 1.2 timesof that in control.Under the same condition,the F/2 medium is most suitable for?-carotene accumulation with yield of1.503?g/10~6cells,which is 1.8 times of that in control.Adding additional carbon sources to ATCC medium showed that the addition of 5 g/L glucose can promote the growth of Dunaliella salina with a maximal cell concentration of 2.928×10~6/m L.The yield of?-carotene reached up to 2.69?g/10~6cells,but resulting in flocculation of the algae during lag growth period.In contrast,adding 3 g/L of acetic acid to the ATCC medium showed no effect on the growth of Dunaliella salina.(2)Dunaliella salina grown under different wavelengths of light sources showed that red light,green light,purple light,and blue light can promote the growth of algae cells to various degrees.The red light and??light promotes the growth of CCAP 19/18 and B01 the most with the maximal cell concentration reached to 10.24×10~6/m L and 17.2×10~6/m L,respectively,1.53 and 1.40 times that of the control group.The?-carotene yield of the two microalgal strains cultured under a purple light source were 0.340?g/10~6cells and 0.325?g/10~6cells,which were 1.25 and 1.36 times that of the control group.Under the white light sources,different light intensities were used.The results showed that the light intensity of 7000 Lux can best promote the growth of Dunaliella salina,with the highest cell concentration of6.5×10~6/m L.When the light intensity exceeds 13000 Lux,the growth of this mciroalga was inhibited.However,when the light intensity exceeds 16000 Lux,it can promote the accumulation of?-carotene in the cells with the yield of 0.423?g/10~6cells,which is about 2.5times of that in the control(3)The three induction conditions of high light,high salt,and nitrogen deficiency were used to investigate Dunaliella salina growth and carotene accumulation.The results showed that high salt(210 g/L)had the strongest inhibitory effect on cell growth,with the maximal cell concentration of 4.32×10~6/m L.Under high light induction conditions(1.6×10~5Lux),the accumulationof?-carotene significantly increasedwith a yield of 0.175?g/10~6cells.Under the high salt induction conditions,the?-carotene yield reached up to 0.123?g/10~6cells.Different concentrations of Na Cl have significant effects on the growth and carotene accumulation of Dunaliella salina.The maximum cell density of Dunaliella salina grown at90 g/L Na CL reached 4.8×10~6/m L cells,which is 1.4 times of that in control The maximal?-carotene yield was achieved at Na CL concentraiton of210 g/L,with yield of 5.71?g/10~6cells,which is 2.7 times that of the control group.(4)Adding different concentrations of triethylamine and nicotine to the ATCC medium showed that 80 mg/L of triethylamine can stimulate the accumulaiton of?-carotene in short term,reaching 0.0847?g/10~6cells on the third day,which was twice that of the control group.Adding nicotine to the substrate has no effect on the growth of CCAP 19/18,and the higher the concentration of nicotine added,the stronger the inhibitory effect on the growth of the algae were seen.When nicotine was added in 20 mg/m L,the chlorophyll yield is 2.690?g/10~6cells,when nicotine of 60 mg/m L was added,the yield of carotene reached to 3.586?g/10~6cells,which is 1.2 times to that of control.(5)Dunaliella salina CCAP 19/18 and B01 were cultivated in a continuous photosynthetic reactor.The maximal cell concentration of two microalgae strains reached 8.95×10~6/m L and5.26×10~6/m L,respectively,in CCAP19/18 and B01.During the induction phase of the continuous photosynthetic reactor,different environmental stresses and inhibitor stresses affected the growth of two Dunaliella salina strains and promoted the production of?-carotene.Among the inhibitors,triethylamine showed negative effect on the growth of two Dunaliella salina.Nitrogen deficiency had the least impact on the growth of the two Dunaliella salina.Under the high light condition of 16000 Lux in the continuous photosynthetic reactor,the?-carotene yield of the two Dunaliella salina reached the highest amount,with B01 accumulated 0.351 mg/m L and CCAP 19/18 accumulated 0.318 mg/m L,respectively,3.6 times and 2.3 times of the control.Based on above experimental results,the optimal light intensity for growth of Dunaliella salina CCAP19/18 and B01 is 7000 Lux,and the optimal light intensity for inducing?-carotene accumulation is 16000 Lux.The addition of diphenylamine to the medium can promote the formation of?-carotene to a certain extent,especially for Dunaliella salina B01,indicating that?-carotene hydroxylase is a key enzyme involved in the formation of?-carotene.In summary,this study has improved the understanding of the two industrial microalgae strains,proposed the most suitable conditions in industrial production of?-carotene using Dunaliella salina These data further help to develop a dedicated photosynthetic reactor model suitable for Dunaliella salina cultivation and?-carotene production. |
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