Research On The Regulation Of DHA Production By Schizochytrium Limacinum Mutant Strain

In 2010,DHA algae oil was approved as a new resource food in China,including marine microalgae originated from Schizochytrium sp.,Crypthecodinium cohnii,and Ulkenia amenoboida..In this paper,the Schizochytrium limacinum mutant strain was used as the original strain.The cell growth and lipid accumulation process of Schizochytrium sp was first observed in the culture process,and its morphological characteristics were described.Then,the effect of fatty acids(SFAs,PUFAs and DHA)on the synthesis of fatty acids(SFAs,PUFAs and DHA)was studied by adding chemical modulators.The composition and content of fatty acids were analyzed by gas chromatography-mass spectrometry(GC-MS).Finally,the response surface optimization of the enzymatic wall breaking process of Schizochytrium sp.mutants strain was carried out to improve the oil yield and DHA yield,and 50 L fermenter was used to amplify the Schizochytrium limacinum mutant strain under the optimum conditions of enzymolysis.Docosahexaenoic acid(Docosahexaenoic acid)is a polyunsaturated fatty acid,abbreviated as DHA.This polyunsaturated fatty acid has a positive effect on the development of the human brain,and plays a key role in the healthy growth of infants and young children.However,it is difficult for us to synthesize the body itself and we need to get it through the outside world.DHA traditionally originated from deep-sea fish oil,this DHA is accumulated when fish eat DHA-containing algae and plankton.Due to marine pollution and many other factors,the market is in short supply and can not meet people's needs.Therefore,the production of DHA by microbial fermentation has become the main research direction.So the main research results are as follows:1.Observe and analyze morphological characteristics and lipid accumulation of Schizochytrium limacinum mutant strain.Through the morphological studies on the growth and development of Schizochytrium limacinum mutant strain,its morphology,morphology and electron microscopic morphology were described.It can be found that,with the extension of culture time,the algae cells proliferate through division and continue to fission and grow,the number of cells gradually increases,the volume becomes larger,the metabolism of life is exuberant and the intracellular fat accumulation.The cells of Schizochytrium limacinum mutant strain began to divide and reproduce into log phase growth,a substantial increase in the number of cells,the nutrients in the culture medium is consumption gradually,at the same time culture due to the growth ofSchizochytrium limacinum mutant strain become thickened,culture color darker.After the cultivation of about 120 h,the intracellular oil became more and more and gradually connected into sheets.At this time,the growth of Schizochytrium limacinum mutant strain cells entered the stable period and produced a large amount of oil.2.Effect of chemical modulators on DHA production by fermentation of Schizochytrium limacinum mutant strain.The effects of exogenous chemical additives on SFAs,PUFAs and DHA were investigated.First found crack pot of Schizochytrium limacinum mutant strain through the experiment in fermentation logarithmic phase of adding moderate concentration of ethanol amine(ETA),naphthalene acid(BNOA)and salicylic acid(SA)can improve crack pot of DHA content of Schizochytrium limacinum mutant strain.It was found that the amount of saturated fatty acids decreased and the amount of unsaturated fatty acids increased relative to the control group by chromatography-mass spectrometry(GC-MS).Through the single factor experiment found that adding just the right amount of three kinds of chemical modulators can improve the ability of crack pot synthesis DHA of Schizochytrium limacinum mutant strain.Adding 150 mg/L of ethanolamine(ETA),10 mg/L of naphthalene acid(BNOA)and 0.5 mg/L of salicylic acid(SA),compared with the control group,the results show that the mass fraction of DHA respectively increased by 16.81%,15.52%,17.94%.Then through the orthogonal experiment and found that at the same time add 150 mg/L of ethanolamine(ETA),10 mg/L naphthalene acid(BNOA)1mg/L and salicylic acid(SA)can improve the ability of crack pot synthesis DHA of Schizochytrium limacinum mutant strain,DHA production reached 6.18 g/L,up12.77% than the control group.3.Optimization of enzymatic cell disruption from Schizochytrium limacinum mutant strain by Response Surface Methodology.In order to explore the optimum conditions of algae oil extraction technology in the process of cell wall,using alkaline to disrupt cell of Schizochytrium limacinum mutant strain to extract oil.The effects of enzymatic lysis temperature,enzymatic lysis time,enzyme dosage and pH value on oil yield and DHA yield were studied.Then based on the single-factor experiment,design response surface experiment was used to screen out the best conditions for cell disruption.The fatty acid composition and content of algal oil under the optimum conditions were analyzed by gas chromatography-mass spectrometry(GC-MS).The results showed that the oil andDHA yield reached the highest level at the enzymatic lysis temperature 55 ?,enzymatic lysis time 9h,enzyme dosage 3% and the pH value 8,which were 14.52 g /L and 7.12 g / L.At the same time,At the same time,a 50 L fermentor scale-up experiment was conducted.The oil yield and DHA yield reached 26.27 g/L and 12.89g/L,respectively.