Optimization Of High-density Heterotrophic Conditions For Lutein Production In Chlorella Sp.

Lutein is an oxygen-containing carotenoid,and microalgae is considered as an emerging source of lutein.Due to its fast growth rate,strong environmental adaptability,short growth cycle and easy to carry out large-scale industrial culture,microalgae-based lutein has great application value and development potential.In this study,the nutrient and culture conditions for heterotrophic production of lutein by Chlorella sorokiniana FZU60 were optimized,and an efficient fed-batch strategy was established to achieve high-density heterotrophic cultivation,and the biochemical composition of algal cells was characterized.The aim of this study was to establish an easy and scalable heterotrophic model for microalgal lutein production.The main research results are as follows:The effects of different nutritional conditions on heterotrophic production of lutein were investigated by batch culture in shaking flask.The results showed that the optimal medium composition was glucose 10.0 g/L,sodium nitrate 1.75 g/L,potassium dihydrogen phosphate trihydrate 0.10 g/L,sodium chloride 5.0 g/L,magnesium sulfate heptanehydrate 0.4 g/L,calcium chloride dihydrate 0.15 g/L,potassium chloride 0.6 g/L,Tris 0.5 g/L and trace element 25 mL/L.Under these nutritional conditions,strain FZU60 could obtain a lutein content of 3.39 mg/g and a lutein productivity of 5.40 mg/L/d,which increased by 175.6% and 191.9% compared with the initial nutritional conditions,respectively.The effects of different culture conditions on heterotrophic production of lutein were investigated by shaking flask and 5 L fermenter.The results of shaking flask experiment showed that biomass productivity and lutein content should be considered comprehensively in the batch culture process,and the best time to finish the culture was when the lutein productivity reached the maximum.In addition,initial cell concentration and culture temperature should be controlled at 1.40 g/L and 30℃,respectively.On this basis,batch culture was further carried out in a 5 L fermenter.It was found that under the conditions of pH 7.5 and initial stirring speed of 200 r/min,strain FZU60 could obtain higher lutein content and productivity,which could reach3.23 mg/g and 12.15 mg/L/d,respectively.Compared with the result in Chapter 1,the lutein productivity increased by 125.0%.To further improve the production efficiency of lutein,the effects of three fed-batch strategies on heterotrophic lutein production of strain FZU60 were investigated in a 5 L fermenter.The results showed that strain FZU60 could achieve the highest biomass productivity,lutein production and productivity,which were17.96 g/L/d,289.59 mg/L and 57.01 mg/L/d,respectively,under the Fed-batch Ⅲ strategy(i.e.,feeding medium with urea as the nitrogen source).The obtained results were superior to those reported in most literatures.The results of cell composition analysis showed that Fed-batch Ⅰ strategy(i.e.,only feeding carbon and nitrogen sources)was more conducive to the accumulation of lipid and carbohydrate,while Fed-batch Ⅱ strategy(i.e.,feeding medium with sodium nitrate as the nitrogen source)and Fed-batch Ⅲ strategy were more conducive to the accumulation of protein and pigment.