Study On The Optimization Of Culture Conditions For The Production Of Lutein From Chlorella Sp.and The Stability Of Lutein

Chlorella is an unicellular microalga.It contains chloroplasts and other photosynthetic apparatus,which makes it good for autotrophic growth.Several species of microalgae have shown the ability to utilize sugars or other organic carbon sources for heterotrophic cultivation in the dark.Chlorella is rich in protein,unsaturated fatty acids,lutein and vitamins;therefore it has high nutritional value and physiological effects such as comprehensive and balanced nutrient.Among several value-added pigments,lutein has a wide range of biological activity:it can effectively eliminate free radicals,inhibite growth of tumor cells,protect eyesight,prevent senile macular degeneration,and have abilities of anti-oxidation,anti-atherosclerosis,anti-cancer,anti-aging,prevention of ulcers and have other medicinal health benefits.Optimization of heterotrophic culture conditions of Chlorella sp.for lutein production and lutein stability have been investigated in this study.The main findings are as follows:(1)In order to locate key factors that affect Chlorella sp.'s growth and lutein production,we performed single factor experiment.The results of single factor test reflected the concentration of glucose,potassium nitrate concentration,and inoculum concentration had great effects on the microalga Chlorella sp.'s growth and lutein yield,based on which we do response surface experiment to optimize.The results revealed that glucose was the best carbon source and potassium nitrate was the best nitrogen source to support sustained and rapid growth of this strain of Chlorella sp.as well as lutein production.The results of optimization were as follows:the glucose concentration was 49.21 g·L-1;potassium nitrate concentration was 5.40 g·L-1;inoculum concentration was 0.23 g·L-1;initial pH was 6.0 and temperature was 30?.When cultured in dark for 7 days with shaking of 200 rpm,Chlorella final dry weight was 18.90 g·L-1,and the maximal lutein concentration was 30.06 mg·L-1.(2)In order to determine the best way to extract lutein from algae,we tested several pretreatment method and ultimately find thaw broken was the most effective.Compared to dry extraction,wet extraction was better.Best extraction solvent was methanol/dichloromethane in the ratio of 2:1(v/v).Through single factor experiment and response surface methodology optimization,we determined the optimum extraction conditions for Chlorella lutein were as follows:extraction temperature of 42.8?,algal dry weight to solvent ratio of 1 |:94(w/v),treating time of 41min.The efficiency of lutein relative extraction was 96.83%achieved under the optimized conditions.(3)Response surface methodology experimental results from light induced lutein production of Chlorella showed that inoculation concentration and potassium nitrate concentration were significantly affected dry weight and yield lutein of Chlorella.The best parameters were as follows:the inoculation concentration was 3.26 g·L-1,potassium nitrate concentration was 7.30 g·L-1.In this condition,the final dry weight and lutein concentration were 1.97 and 9.20 times of those recorded before induction.Lutein yield was improved greatly through this method.(4)The stability and antioxidant activity of Chlorella lutein were investigated.The influences of illumination,sterilization,oxidants,antioxidants,edible acids and preservatives on the stability of Chlorella lutein were studied.The samples were placed in a certain period of time under different conditions,then the lutein production was determined by high performance liquid chromatography.The preservation rate was calculated and the influence of respective factors was analyzed.The results showed that Chlorella lutein was sensitive to light and it faded quickly under the sunlight.The influence of temperature showed that it could keep the preservation rate in lower temperature under darkness,so it is suitable to store it in low temperature and darkness.It was relatively stable at high-temperature and short-time sterilization.Ascorbic acid could protect the stability of lutein,while high concentrations of antioxidants,edible acid and preservatives had damage to lutein.Chlorella lutein had strong antioxidant capacity.When the DPPH radical scavenging rate was 50%,the Chlorella lutein concentration was 211.41 mg·L-1.