光径及氮浓度对缺刻缘绿藻生长、油脂和花生四烯酸积累的影响
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摘要
以缺刻缘绿藻(Parietochloris incisa)为实验材料,采用BG-11培养基,分别在2种氮浓度和3种不同光径(LP)的柱状和平板光生物反应器中进行培养,并探究其生长、油脂和花生四烯酸(AA)的积累规律。结果显示:在两种光生物反应器中,光径越小,越有利于缺刻缘绿藻的生长。其中,最大生物量均在17.6 mmol/L氮浓度时获得,分别为5.09 g/L(2.5 cm-柱状)和2.98 g/L(3.0 cm-平板);而最高油脂和AA绝对含量则均在1.0 mmol/L氮浓度和最大光径处获得,分别为39.23%、13.21%(6.0 cm-柱状)和40.74%、11.33%(5.0 cm-平板);另外,两种光生物反应器中的最大油脂单位体积产率分别可以达到216.39 mg/(L·d)(17.6 mmol/L; 2.5 cm-柱状)和135.93 mg/(L·d)(1.0 mmol/L; 1.5 cm-平板);而最高的AA单位体积产率均在1.0 mmol/L低氮条件,最大光径处达到最大,分别为21.65 mg/(L·d)(6.0 cm-柱状)和19.42 mg/(L·d)(5.0 cm-平板)。因此,根据实际生产需要,在1.0 mmol/L低氮条件下,选择6.0 cm光径的柱状光生物反应器或5.0 cm光径的平板光生物反应器,培养缺刻缘绿藻生产AA,能有效降低生产成本。
This study explored effects of 3 light-paths(LP) of column and flat plat photobioreactors under two kinds of nitrogen concentrations on the growth, total lipid and AA content of Parietochloris incisa using BG-11 medium. The results showed that cell growth rate increased by the smaller light-path of two kinds of photobioreactors. The maximal biomass were 5.09 g/L in column photobioreactor(2.5 cm LP) and 2.98 g/L(3.0 cm LP) in flat plate photobioreactor with nitrogen concentration at 17.6 mmol/L. The highest total lipid and AA content at the biggest length of light-path under 1.0 mmol/L nitrogen concentration were 39.23% and 13.21% in column photobioreactor(6.0 cm LP) and there were 40.74% and 11.33% in flat plate photobioreactor(5.0 cm LP). In addition, the maximal volumetric productivity of total lipid was 216.39 mg/(L·d) in column photobioreactor(2.5 cm LP) under 17.6 mmol/L nitrogen concentration and it was 135.93 mg/(L·d) in flat plate photobioreactor(1.5 cm LP) under 1.0 mmol/L nitrogen concentration. The AA volumetric productivity reached the maximum level in the biggest light-path of column [21.65 mg/(L·d)] and flat plate photobiorector [19.42 mg/(L·d)] under 1.0 mmol/L nitrogen concentration. These results indicate that the 6.0 cm lightpath column photobioreactor or 5.0 cm light-path of flat plate photobioreactor under 1.0 mmol/L nitrogen concentration are the best condition to culture Parietochloris incisa for AA production.
This study explored effects of 3 light-paths(LP) of column and flat plat photobioreactors under two kinds of nitrogen concentrations on the growth, total lipid and AA content of Parietochloris incisa using BG-11 medium. The results showed that cell growth rate increased by the smaller light-path of two kinds of photobioreactors. The maximal biomass were 5.09 g/L in column photobioreactor(2.5 cm LP) and 2.98 g/L(3.0 cm LP) in flat plate photobioreactor with nitrogen concentration at 17.6 mmol/L. The highest total lipid and AA content at the biggest length of light-path under 1.0 mmol/L nitrogen concentration were 39.23% and 13.21% in column photobioreactor(6.0 cm LP) and there were 40.74% and 11.33% in flat plate photobioreactor(5.0 cm LP). In addition, the maximal volumetric productivity of total lipid was 216.39 mg/(L·d) in column photobioreactor(2.5 cm LP) under 17.6 mmol/L nitrogen concentration and it was 135.93 mg/(L·d) in flat plate photobioreactor(1.5 cm LP) under 1.0 mmol/L nitrogen concentration. The AA volumetric productivity reached the maximum level in the biggest light-path of column [21.65 mg/(L·d)] and flat plate photobiorector [19.42 mg/(L·d)] under 1.0 mmol/L nitrogen concentration. These results indicate that the 6.0 cm lightpath column photobioreactor or 5.0 cm light-path of flat plate photobioreactor under 1.0 mmol/L nitrogen concentration are the best condition to culture Parietochloris incisa for AA production.
引文
[1]Watanabe S,Hirabayashi S,Boussiba S,et al.Parietochloris incisa comb.nov.(Trebouxiophyceae,Chlorophyta)[J].Phycological Research,1996,44(2):107-108
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[3]Funk C D.Prostaglandins and leukotrienes:advances in eicosanoid biology[J].Science,2001,294(5548):1871-1875
[4]Hansen J,Schade D,Harris C,et al.Docosahexaenoic acid plus arachidonic acid enhance preterm infant growth[J].Prostaglandins Leukotrienes and Essential Fatty Acids,1997,57(2):196
[5]Singh A,Ward O P.Production of high yields of arachidonic acid in a fed-batch system by Mortierella alpina ATCC 32222[J].Applied Microbiology and Biotechnology,1997,48(1):1-5
[6]Yu C Q,Li L N.Research advancement on arachidonic acid[J].Academic Periodical of Farm Products Processing,2007,(4):10-12[于长青,李丽娜.花生四烯酸研究进展.农产品加工(学刊),2007,(4):10-12]
[7]Bigogno C,Khozin-Goldberg I,Boussiba S,et al.Lipid and fatty acid composition of the green oleaginous alga Parietochloris incisa,the richest plant source of arachidonic acid[J].Phytochemistry,2002,60(5):497-503
[8]Bigogno C,Khozin-Goldberg I,Cohen Z.Accumulation of arachidonic acid-rich triacylglycerols in the microalga Parietochloris incisa(Trebuxiophyceae,Chlorophyta)[J].Phytochemistry,2002,60(2):135-143
[9]Gim G H,Ryu J,Kim M J,et al.Effects of carbon source and light intensity on the growth and total lipid production of three microalgae under different culture conditions[J].Journal of Industrial Microbiology and Biotechnology,2016,43(5):605-616
[10]Solovchenko A E,Khozin-Goldberg I,Didi-Cohen S,et al.Effects of light intensity and nitrogen starvation on growth,total fatty acids and arachidonic acid in the green microalga Parietochloris incisa[J].Journal of Applied Phycology,2008,20(3):245-251
[11]Li X,Hu H Y,Zhang Y P.Growth and lipid accumulation properties of a freshwater microalga Scenedesmus sp.under different cultivation temperature[J].Bioresource Technology,2011,102(3):3098-3102
[12]Rai M P,Gautom T,Sharma N.Effect of salinity,pH,light intensity on growth and lipid production of microalgae for bioenergy application[J].Online Journal of Biological Sciences,2015,15(4):260-267
[13]Kaewkannetra P,Enmak P,Chiu T Y.The effect of CO2and salinity on the cultivation of Scenedesmus obliquus for biodiesel production[J].Biotechnology and Bioprocess Engineering,2012,17(3):591-597
[14]Ning Z,Richmond A.Effect of light-path length in outdoor fiat plate reactors on output rate of cell mass and of EPA in Nannochloropsis sp[J].Journal of Biotechnology,1999,35(1-3):351-356
[15]Singh S,Arad S,Richmond A.Extracellular polysaccharide production in outdoor mass cultures of Porphyridium sp.in flat plate glass reactors[J].Journal of Applied Phycology,2000,12(3-5):269-275
[16]Richmond A,Zhang C W.Optimization of a flat plate glass reactor for mass production of Nannochloropsis sp.Outdoors[J].Journal of Biotechnology,2001,85(3):259-269
[17]Zhao P,Zou N,Tan Q,et al.Effect of light-path on the growth and oil yield of Phaeodactylum tricornutum[J].China Oils and Fats,2013,38(3):39-42[赵萍,邹宁,谭青,等.光径对三角褐指藻生长及油脂产量的影响.中国油脂,2013,38(3):39-42]
[18]Zhang C W,Cohen Z,Khozin-Goldberg I,et al.Characterization of growth and arachidonic acid production of Parietochloris incisa comb.nov.(Trebouxiophyceae,Chlorophyta)[J].Journal of Applied Phycology,2002,14(6):453-460
[19]Hu Q A,Richmond A.Environmental Effects on Cell Composition[M].Handbook of Microalgal Culture:Applied Phycology and Biotechnology,Second Edition.John Wiley and Sons,Ltd.2015,83-94
[20]Tong M,Yu S Y,Ouyang L L,et al.Comparison of increased arachidonic acid content in Myrmecia incisa cultured during the course of nitrogen or phosphorus starvation[J].Journal of Fisheries of China,2011,35(5):763-773[童牧,于水燕,欧阳珑玲,等.氮饥饿与磷饥饿促使缺刻缘绿藻花生四烯酸含量增加的比较.水产学报,2011,35(5):763-773]
[21]He S S,Gao B Y,Lei X Q,et al.Effects of initial nitrogen supply on the growth,morphology and lipid accumulation of oleaginous microalga Eustigmatos vischeri(Eustigmatophyceae)[J].Acta Hydrobiologica Sinica,2015,39(3):574-582[何思思,高保燕,雷学青,等.初始硝酸钠浓度对魏氏真眼点藻的生长、形态和油脂积累的影响.水生生物学报,2015,39(3):574-582]
[22]Solovchenko A E,Merzlyak M N,Chivkunova O B,et al.Effects of illumination and nitrogen starvation on accumulation of arachidonic acid by the microalga Parietochloris incisa[J].Moscow University Biological Sciences Bulletin,2008,63(1):44-48
[23]Khozin-Goldberg I,Shrestha P,Cohen Z.Mobilization of arachidonyl moieties from triacylglycerols into chloroplastic lipids following recovery from nitrogen starvation of the microalga Parietochloris incisa[J].Biochimica Biophysica Acta,2005,1738(1-3):63-71
[24]Yin Y Q,Huang L D,Hu Q,et al.Effects of light intensity,nitrogen sources,and nitrogen levels on the growth,total lipids,and arachidonic acid accumulation of Parietochloris incisa[J].Plant Science Journal,2017,35(4):592-602[印尤强,黄罗冬,胡强,等.光强和氮源及其浓度对缺刻缘绿藻生长、油脂和花生四烯酸积累的影响.植物科学学报,2017,35(4):592-602]
[25]Laurens L M L,Quinn M,Wychen S V,et al.Accurate and reliable quantification of total microalgal fuel potential as fatty acid methyl esters by in situ transesterification[J].Analytical and Bioanalytical Chemistry,2012,403(1):167-178
[26]Christie W W,Han X L.Lipid Analysis[M].4th ed.Cambridge:Woodhead Publishing,2012,145-158
[27]Borowitzka M A.Commercial production of microalgae:ponds,tanks,tubes and fermenters[J].Journal of Biotechnology,1999,70(3):313-321
[28]Pulz O,Scheibenbogen K.Photobioreactors:Design and performance with respect to light energy input[J].Bioprocess and Algae Reactor Technology Apoptosis,1998,38:123-152
[29]Liu J N,Hu P,Yao L,et al.Advance of photobioreactor on microalgal cultivation[J].Food Science,2006,27(12):772-777[刘娟妮,胡萍,姚领,等.微藻培养中光生物反应器的研究进展.食品科学,2006,27(12):772-777]
[30]Brennan L,Owende P.Biofuels from microalgae-A review of technologies for production,processing,and extractions of biofuels and co-products[J].Renewable and Sustainable Energy Reviews,2010,14(2):557-577
[31]Ugwu C U,Aoyagi H,Uchiyama H.Photobioreactors for mass cultivation of algae[J].Bioresource Technology,2008,99(10):4021-4028
[32]Zhu J Y,Rong J F,Zong B N.Factors in mass cultivation of micrroalgae for biodiesel[J].Chinese Journal of Catalysis,2013,34(1):80-100[朱俊英,荣峻峰,宗保宁.影响微藻规模化培养的因素.催化学报,2013,34(1):80-100]
[33]Zhang W,Sang M,Li A F,et al.Effects of light paths on the growth and lipid accumulation of two strains of soil green algae[J].Renewable Energy Resources,2010,28(3):67-71[展望,桑敏,李爱芬,等.不同光径对两株土壤绿藻生长及脂类积累的影响.可再生能源,2010,28(3):67-71]
[34]Liu J G,Liu W,Zvi C,et al.The high productivity of biomass andarachidonic acid in continuous high density culture of Parietochloris incise[J].Oceanologia et Limnologia Sinica,2002,33(5):499-508[刘建国,刘伟,Zvi C,等.雪藻高密度连续培养中生物量和花生四烯酸的高产率.海洋与湖沼,2002,33(5):499-508]
[35]Huang X,Huang Z,Wen W,et al.Effects of nitrogen supplementation of the culture medium on the growth,total lipid content and fatty acid profiles of three microalgae(Tetraselmis subcordiformis,Nannochloropsis oculata and Pavlova viridis)[J].Journal of Applied Phycology,2013,25(1):129-137
[36]Solovchenko A E,Khozin-Goldberg I,Didi-Cohen S,et al.Effects of light and nitrogen starvation on the content and composition of carotenoids of the green microalga Parietochloris incisa[J].Russian Journal of Plant Physiology,2008,55(4):455-462
[37]Zhao D X.Advances on microalgal arachidonic acid[J].Fisheries Science,2004,23(10):42-44[赵大显.微藻花生四烯酸的研究进展.水产科学,2004,23(10):42-44]
[38]Liu J G,Zvi C,Amos R.Fatty acids profile in high cell density culture of arachidoncic acid-rich Parietochloris incise(Trebouxiophyceae,Chlorophyta)exposed to high PFD[J].Chinese Journal of Oceanology and Limnology,2002,20(2):149-156
[2]Khozin-Goldberg I,Bigogno C,Shrestha P,et al.Nitrogen starvation induces the accumulation of arachidonic acid in the freshwater green alga Parietochloris incisa(Trebuxiophyceae)[J].Journal of Phycology,2002,38(5):991-994
[3]Funk C D.Prostaglandins and leukotrienes:advances in eicosanoid biology[J].Science,2001,294(5548):1871-1875
[4]Hansen J,Schade D,Harris C,et al.Docosahexaenoic acid plus arachidonic acid enhance preterm infant growth[J].Prostaglandins Leukotrienes and Essential Fatty Acids,1997,57(2):196
[5]Singh A,Ward O P.Production of high yields of arachidonic acid in a fed-batch system by Mortierella alpina ATCC 32222[J].Applied Microbiology and Biotechnology,1997,48(1):1-5
[6]Yu C Q,Li L N.Research advancement on arachidonic acid[J].Academic Periodical of Farm Products Processing,2007,(4):10-12[于长青,李丽娜.花生四烯酸研究进展.农产品加工(学刊),2007,(4):10-12]
[7]Bigogno C,Khozin-Goldberg I,Boussiba S,et al.Lipid and fatty acid composition of the green oleaginous alga Parietochloris incisa,the richest plant source of arachidonic acid[J].Phytochemistry,2002,60(5):497-503
[8]Bigogno C,Khozin-Goldberg I,Cohen Z.Accumulation of arachidonic acid-rich triacylglycerols in the microalga Parietochloris incisa(Trebuxiophyceae,Chlorophyta)[J].Phytochemistry,2002,60(2):135-143
[9]Gim G H,Ryu J,Kim M J,et al.Effects of carbon source and light intensity on the growth and total lipid production of three microalgae under different culture conditions[J].Journal of Industrial Microbiology and Biotechnology,2016,43(5):605-616
[10]Solovchenko A E,Khozin-Goldberg I,Didi-Cohen S,et al.Effects of light intensity and nitrogen starvation on growth,total fatty acids and arachidonic acid in the green microalga Parietochloris incisa[J].Journal of Applied Phycology,2008,20(3):245-251
[11]Li X,Hu H Y,Zhang Y P.Growth and lipid accumulation properties of a freshwater microalga Scenedesmus sp.under different cultivation temperature[J].Bioresource Technology,2011,102(3):3098-3102
[12]Rai M P,Gautom T,Sharma N.Effect of salinity,pH,light intensity on growth and lipid production of microalgae for bioenergy application[J].Online Journal of Biological Sciences,2015,15(4):260-267
[13]Kaewkannetra P,Enmak P,Chiu T Y.The effect of CO2and salinity on the cultivation of Scenedesmus obliquus for biodiesel production[J].Biotechnology and Bioprocess Engineering,2012,17(3):591-597
[14]Ning Z,Richmond A.Effect of light-path length in outdoor fiat plate reactors on output rate of cell mass and of EPA in Nannochloropsis sp[J].Journal of Biotechnology,1999,35(1-3):351-356
[15]Singh S,Arad S,Richmond A.Extracellular polysaccharide production in outdoor mass cultures of Porphyridium sp.in flat plate glass reactors[J].Journal of Applied Phycology,2000,12(3-5):269-275
[16]Richmond A,Zhang C W.Optimization of a flat plate glass reactor for mass production of Nannochloropsis sp.Outdoors[J].Journal of Biotechnology,2001,85(3):259-269
[17]Zhao P,Zou N,Tan Q,et al.Effect of light-path on the growth and oil yield of Phaeodactylum tricornutum[J].China Oils and Fats,2013,38(3):39-42[赵萍,邹宁,谭青,等.光径对三角褐指藻生长及油脂产量的影响.中国油脂,2013,38(3):39-42]
[18]Zhang C W,Cohen Z,Khozin-Goldberg I,et al.Characterization of growth and arachidonic acid production of Parietochloris incisa comb.nov.(Trebouxiophyceae,Chlorophyta)[J].Journal of Applied Phycology,2002,14(6):453-460
[19]Hu Q A,Richmond A.Environmental Effects on Cell Composition[M].Handbook of Microalgal Culture:Applied Phycology and Biotechnology,Second Edition.John Wiley and Sons,Ltd.2015,83-94
[20]Tong M,Yu S Y,Ouyang L L,et al.Comparison of increased arachidonic acid content in Myrmecia incisa cultured during the course of nitrogen or phosphorus starvation[J].Journal of Fisheries of China,2011,35(5):763-773[童牧,于水燕,欧阳珑玲,等.氮饥饿与磷饥饿促使缺刻缘绿藻花生四烯酸含量增加的比较.水产学报,2011,35(5):763-773]
[21]He S S,Gao B Y,Lei X Q,et al.Effects of initial nitrogen supply on the growth,morphology and lipid accumulation of oleaginous microalga Eustigmatos vischeri(Eustigmatophyceae)[J].Acta Hydrobiologica Sinica,2015,39(3):574-582[何思思,高保燕,雷学青,等.初始硝酸钠浓度对魏氏真眼点藻的生长、形态和油脂积累的影响.水生生物学报,2015,39(3):574-582]
[22]Solovchenko A E,Merzlyak M N,Chivkunova O B,et al.Effects of illumination and nitrogen starvation on accumulation of arachidonic acid by the microalga Parietochloris incisa[J].Moscow University Biological Sciences Bulletin,2008,63(1):44-48
[23]Khozin-Goldberg I,Shrestha P,Cohen Z.Mobilization of arachidonyl moieties from triacylglycerols into chloroplastic lipids following recovery from nitrogen starvation of the microalga Parietochloris incisa[J].Biochimica Biophysica Acta,2005,1738(1-3):63-71
[24]Yin Y Q,Huang L D,Hu Q,et al.Effects of light intensity,nitrogen sources,and nitrogen levels on the growth,total lipids,and arachidonic acid accumulation of Parietochloris incisa[J].Plant Science Journal,2017,35(4):592-602[印尤强,黄罗冬,胡强,等.光强和氮源及其浓度对缺刻缘绿藻生长、油脂和花生四烯酸积累的影响.植物科学学报,2017,35(4):592-602]
[25]Laurens L M L,Quinn M,Wychen S V,et al.Accurate and reliable quantification of total microalgal fuel potential as fatty acid methyl esters by in situ transesterification[J].Analytical and Bioanalytical Chemistry,2012,403(1):167-178
[26]Christie W W,Han X L.Lipid Analysis[M].4th ed.Cambridge:Woodhead Publishing,2012,145-158
[27]Borowitzka M A.Commercial production of microalgae:ponds,tanks,tubes and fermenters[J].Journal of Biotechnology,1999,70(3):313-321
[28]Pulz O,Scheibenbogen K.Photobioreactors:Design and performance with respect to light energy input[J].Bioprocess and Algae Reactor Technology Apoptosis,1998,38:123-152
[29]Liu J N,Hu P,Yao L,et al.Advance of photobioreactor on microalgal cultivation[J].Food Science,2006,27(12):772-777[刘娟妮,胡萍,姚领,等.微藻培养中光生物反应器的研究进展.食品科学,2006,27(12):772-777]
[30]Brennan L,Owende P.Biofuels from microalgae-A review of technologies for production,processing,and extractions of biofuels and co-products[J].Renewable and Sustainable Energy Reviews,2010,14(2):557-577
[31]Ugwu C U,Aoyagi H,Uchiyama H.Photobioreactors for mass cultivation of algae[J].Bioresource Technology,2008,99(10):4021-4028
[32]Zhu J Y,Rong J F,Zong B N.Factors in mass cultivation of micrroalgae for biodiesel[J].Chinese Journal of Catalysis,2013,34(1):80-100[朱俊英,荣峻峰,宗保宁.影响微藻规模化培养的因素.催化学报,2013,34(1):80-100]
[33]Zhang W,Sang M,Li A F,et al.Effects of light paths on the growth and lipid accumulation of two strains of soil green algae[J].Renewable Energy Resources,2010,28(3):67-71[展望,桑敏,李爱芬,等.不同光径对两株土壤绿藻生长及脂类积累的影响.可再生能源,2010,28(3):67-71]
[34]Liu J G,Liu W,Zvi C,et al.The high productivity of biomass andarachidonic acid in continuous high density culture of Parietochloris incise[J].Oceanologia et Limnologia Sinica,2002,33(5):499-508[刘建国,刘伟,Zvi C,等.雪藻高密度连续培养中生物量和花生四烯酸的高产率.海洋与湖沼,2002,33(5):499-508]
[35]Huang X,Huang Z,Wen W,et al.Effects of nitrogen supplementation of the culture medium on the growth,total lipid content and fatty acid profiles of three microalgae(Tetraselmis subcordiformis,Nannochloropsis oculata and Pavlova viridis)[J].Journal of Applied Phycology,2013,25(1):129-137
[36]Solovchenko A E,Khozin-Goldberg I,Didi-Cohen S,et al.Effects of light and nitrogen starvation on the content and composition of carotenoids of the green microalga Parietochloris incisa[J].Russian Journal of Plant Physiology,2008,55(4):455-462
[37]Zhao D X.Advances on microalgal arachidonic acid[J].Fisheries Science,2004,23(10):42-44[赵大显.微藻花生四烯酸的研究进展.水产科学,2004,23(10):42-44]
[38]Liu J G,Zvi C,Amos R.Fatty acids profile in high cell density culture of arachidoncic acid-rich Parietochloris incise(Trebouxiophyceae,Chlorophyta)exposed to high PFD[J].Chinese Journal of Oceanology and Limnology,2002,20(2):149-156