平板式光生物反应器中紫球藻培养条件的优化
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摘要
紫球藻(Porphyridium cruentum)是属于红藻门的一种海洋微藻,在生长过程中能合成硫酸酯多糖、多不饱和脂肪酸和藻胆蛋白等重要生物活性物质。本论文主要运用自行设计的平板式光生物反应器培养紫球藻,在培养基优化的基础上,对紫球藻在平板式光生物反应器中的生长参数进行了测定,对紫球藻的培养工艺进行优化,获得以下主要结果:
摇瓶培养实验表明,初始pH为原始pH(7.5)时,有利于藻体生物量、胞外多糖的积累;当接种量为10%的,藻细胞生物量收获最大。当接种量为10%和20%时,胞外多糖的收获量最大,分别可以达到460mg/L和475mg/L。硝基氮可促进紫球藻生物量和胞外多糖的积累;肌醇对紫球藻生物量、胞外多糖的积累没有明显的影响。
采用自行研制的平板式光生物反应器培养紫球藻,测定了紫球藻在不同光径平板式光生物反应器中的生长参数(溶氧、pH、光衰减、氮源的消耗、磷源的消耗),确定了紫球藻的光衰减模型以及平板式光生物反应器中平均光强计算方程;发现30 mm小光径的反应器有利于藻对光的利用效率,提高了藻的培养密度以及多糖的产量。
运用均匀设计的方法对紫球藻的平板反应器培养条件进行了优化,考察了更新率等与半连续培养过程中细胞总采收量和多糖总采收量之间的关系;探讨了流加时间等对流加培养过程中最大细胞密度和多糖产量的影响。结果显示:在NaNO3 1.504 g/L,更新率32.93%,更新周期1 d的培养条件下,细胞总采收量最大,达到69.5 g;在NaNO3 3.5 g/L,更新率26.83%,更新周期2.9 d的条件下,多糖总采收量最大,达到17.6 g。
Porphyridium cruentum,a kind of the marine microalgae, attracts increasing interest for it can synthsis some valuable compounds such as phycoerythrin, sulfated exopolysaccharides and polyunsaturated fatty acids. To investigate the effect of culture conditions on the growth and some metabolites production of P. cruentum, the light attenuation model, the some key growth parameters and the optimal culture process were researched in the flat plate photobioreactor. The results were showed as follow.
In the flask cultures, the productivity of biomass and extracellular polysaccharides of P. cruentum arrived to the biggest value when the original pH was 7.5. When the vaccination was 10%, the largest biomass was harvested; however, the largest extracellular polysaccharides of 460mg/ and 475mg/L were obtained when vaccination was 10% and 20%, respectively. And the nitro nitrogen could increase the productivity of the biomass and accumulation of extracellular polysaccharides of P. cruentum.
The optical attenuation equation and the average light intensity calculation equation of P. cruentum were obtained in the flat plate photobioreactors. And some important growth parameters were determinated such as DO, pH, optical attenuator, the consumption of phosphorus and nitrogen in the different light-path flat plate photobioreactors. The results showed that the photobioreactor of small light-path (30 mm) could improved the light utilization efficiency, and a high of cells density and polysaccharide production could be obtained.
To improve the cultural density and polysaccharide production, the cultural conditions of P.cruentum were optimized by the method of uniform design in flat plate photobioreactors. And the relationship between the renewal rate and the total harvest of biomass and polysaccharides was studied under the cultural modle of semi-continuous culture. The results showed that the total productivity of biomass reached to 69.5 g under the conditions of NaNO3 1.504 g/L, 32.93% renewal rate, and 1d renewal cycle; and the total harvest of polysaccharides amounted to 17.6 g with the conditions of NaNO3 3.5 g/L, 26.83% renewal rate and 2.9d renewal cycle.
摇瓶培养实验表明,初始pH为原始pH(7.5)时,有利于藻体生物量、胞外多糖的积累;当接种量为10%的,藻细胞生物量收获最大。当接种量为10%和20%时,胞外多糖的收获量最大,分别可以达到460mg/L和475mg/L。硝基氮可促进紫球藻生物量和胞外多糖的积累;肌醇对紫球藻生物量、胞外多糖的积累没有明显的影响。
采用自行研制的平板式光生物反应器培养紫球藻,测定了紫球藻在不同光径平板式光生物反应器中的生长参数(溶氧、pH、光衰减、氮源的消耗、磷源的消耗),确定了紫球藻的光衰减模型以及平板式光生物反应器中平均光强计算方程;发现30 mm小光径的反应器有利于藻对光的利用效率,提高了藻的培养密度以及多糖的产量。
运用均匀设计的方法对紫球藻的平板反应器培养条件进行了优化,考察了更新率等与半连续培养过程中细胞总采收量和多糖总采收量之间的关系;探讨了流加时间等对流加培养过程中最大细胞密度和多糖产量的影响。结果显示:在NaNO3 1.504 g/L,更新率32.93%,更新周期1 d的培养条件下,细胞总采收量最大,达到69.5 g;在NaNO3 3.5 g/L,更新率26.83%,更新周期2.9 d的条件下,多糖总采收量最大,达到17.6 g。
Porphyridium cruentum,a kind of the marine microalgae, attracts increasing interest for it can synthsis some valuable compounds such as phycoerythrin, sulfated exopolysaccharides and polyunsaturated fatty acids. To investigate the effect of culture conditions on the growth and some metabolites production of P. cruentum, the light attenuation model, the some key growth parameters and the optimal culture process were researched in the flat plate photobioreactor. The results were showed as follow.
In the flask cultures, the productivity of biomass and extracellular polysaccharides of P. cruentum arrived to the biggest value when the original pH was 7.5. When the vaccination was 10%, the largest biomass was harvested; however, the largest extracellular polysaccharides of 460mg/ and 475mg/L were obtained when vaccination was 10% and 20%, respectively. And the nitro nitrogen could increase the productivity of the biomass and accumulation of extracellular polysaccharides of P. cruentum.
The optical attenuation equation and the average light intensity calculation equation of P. cruentum were obtained in the flat plate photobioreactors. And some important growth parameters were determinated such as DO, pH, optical attenuator, the consumption of phosphorus and nitrogen in the different light-path flat plate photobioreactors. The results showed that the photobioreactor of small light-path (30 mm) could improved the light utilization efficiency, and a high of cells density and polysaccharide production could be obtained.
To improve the cultural density and polysaccharide production, the cultural conditions of P.cruentum were optimized by the method of uniform design in flat plate photobioreactors. And the relationship between the renewal rate and the total harvest of biomass and polysaccharides was studied under the cultural modle of semi-continuous culture. The results showed that the total productivity of biomass reached to 69.5 g under the conditions of NaNO3 1.504 g/L, 32.93% renewal rate, and 1d renewal cycle; and the total harvest of polysaccharides amounted to 17.6 g with the conditions of NaNO3 3.5 g/L, 26.83% renewal rate and 2.9d renewal cycle.
引文
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2. Kathleen L. et al. Ultrastructure of Cell Division in the Unicellular Red Alga Porphyridium cruentum, Can. J. Bot., 1982, 60: 85-97.
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2. Kathleen L. et al. Ultrastructure of Cell Division in the Unicellular Red Alga Porphyridium cruentum, Can. J. Bot., 1982, 60: 85-97.
3. Lin H. et al. Light and Electron Microscope Observations on Motile Cells of Porphyridium purpureum (Rhodophyta), J. of Phycol., 1975, 11:452-457.
4.王明兹,施巧琴,郑梅清.紫球藻的培养与利用.亚热带植物科学,2001,30(2):66-69.
5. Schmidt P et al. Chromophore protein interaction controls the complexity of the phytochrome photocycle, J. Photochem. Photobiol. Biology, 1996, 340):73-77
6. Glazer A N. Phycobiliprotein-a family of valuable, widely used fluorophores, J. Appl. Phycology, 1994, 6:105-112
7.许实波.海洋生物制药.北京:化学工业出版社,2002.
8.王明兹,施巧琴,郑梅清等.紫球藻的培养与利用.亚热带植物科学.2001,30 (2):66-69
9.王长海,李叙风,鞠宝等.紫球藻及其应用研究.海洋通报.1998,17 (3):79-84.
10. Rebolloso Fuentes M M, Acien Fernandez G G, Sanchez Perez J A et al. Biomass nutrient profiles of the microalga Porphyridium cruentum. Food Chemistry.2000,70:345-353.
11. Ginzberg A, Cohen M, Sod-Moriah UA et al. Chickens fed with biomass of the red microalga Porphyridium sp. have reduced blood cholesterol level and modified fatty acid composition in egg yolk. J Apll Phyco1.2000, 12(3/5):325-330.
12. Dvir I, Chayoth R, Sod-Moriah U et al. Soluble polysaccharide and biomass redmicroalga Porphyridium sp. alter intestinal morphology and reduce serum cholesterolin rats. Br J Nutr.2000,84(4):469-476.
13.肖华山,林和,范子南等.红外线和植物生长物质对紫球藻生长及代谢的影响.热带海洋.2000, 19 (2) : 64-68
14.王明兹,陈必链,庄惠如等.倍频Nd:YAG激光对紫球藻生长与胞外多糖的影响.激光生物学报.2002, 11(1) :6-9.
15.黄鹭强,王明兹,欧琳等.半导体激光对紫球藻生物学效应的影响.生物技术.2002, 12 (1):14-16.
16.王长海.微藻的光生物反应器培养中国科学院博士论文,1998.
17.王长海,郭尽力,鞠宝等.(1998)微藻与微藻生物技术.全国第二届海洋生命活性物质与天然生化药物学术会议论文集,上海,p194-198.
18.温少红,王长海,鞠宝.紫球藻细胞中AA, EPA及色素提取的研究.中国海洋药物.1999, 2: 28-31.
19.温少红,王长海.光照和培养时间对紫球藻细胞脂肪酸含量的影响.中国海洋药物.2000, 1:47-50.
20. Wang C H, et a1. Studies on high concentration culture of Porphyridium cruentum[J]. Proceedings of Young Asian Biochemical Engineers Community 1997, Tianjin University,1997, 37-39.
21. Guil-GuerreroJL, Belarbi EH, Rebolloso-Fuentes MM. Eicosapentaenoic andarachidonic acids purification from the red microalga Porphyridium cruentum. Bioseparation. 2000,9(5):299-306.
22. Shiran D,Khozin I, Heimer YM et a1.Biosythesis of eicosapentaenoic acid in the microalga Porphyridium cruentum. I: The use of externally supplied fatty acids. Lipids. 1996,31(12):1277-1282
23. Khozin-Goldberg I, Hu ZhY, Adlerstein D et al. Triacylglycerols participate in the eukaryotic pathway of PUFAs biosynthesis in the red microalga Porphyridium cruentum. In Williams,JP, Mobashsher,UK and Nora WL(eds) Physiology,Biochemistry and molecular biology of plant lipids. Kluwer AcademicPublishers,Dodrecht,Netherlands,90-92.
24. Khozin I. And C. Zvi, Differential response of microalgae to the substitutedpyridazinone. Phytochem. 1996,42(4):1025-1029.
25. Khozin-Goldberg I, Bigogno C, Cohen Z. Salicylhydroxamic acid inhibits delta desaturation in the microalga Porphyridium cruentum. Biochim Biophys Acta.1999,1439(3):384-394.
26. Khozin-Goldberg I, Hu ZhY, Adlerstein D et al. Triacylglycerols of the redmicroalgaPorphyridium cruentum can contribute to the biosynthesis of eukaryoticgalactolipids. Lipids. 2000, 35(8):881-889.
27.王长云,管华诗.多糖抗病毒作用研究进展1.多糖抗病毒作用.生物工程进展.2000, 20 (1):17-20.
28.王长云,管华诗.多糖抗病毒作用研究进展2.硫酸多糖抗病毒作用.生物工程进展.2000, 20 (2) : 3-8.
29. Rebolloso Fuentes M M, Acien Fernandez G G, Sanchez Perez J A et al. Biomass nutrient profiles of the microalga Porphyridium cruentum. Food Chemistry.2000,70:345-353.
30. Geresh S, Mamontov A, Weinstein J. Sulfation of extracellular polysaccharides of red microalgae:preparation, characterization and properties. J Biochem Biophys Methods. 2002, 50 :179-187.
31. Gloaguen V, Ruiz G, Morvan H. The extracellular polysaccharide of Porphyridium sp. an NMR study of lithium-resistant oligosaccharidic fragments. Carbohydr Res. 2004, 339(1):97-103.
32.王长海,温少红,鞠宝.紫球藻多糖的提取和测定.中国海洋药物.1999, 18(1):22-25.
33. Heaney- Kieras J, Chapman DJ. Structural studies on the extracellular polysaccharide of the red alga,Porphyridium.Carbohydr Res.1976,52:169-77.
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