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微藻脂肪酸测定步骤一般是先将微藻样品进行前处理(脂类抽提、皂化、甲酯化以及脂肪酸甲酯的萃取),然后将得到的脂肪酸甲酯在气相色谱-质谱仪上分析并计算,得出脂肪酸的含量和组成。文中概述了测定微藻脂肪酸时,样品前处理过程中不同的皂化、甲酯化和萃取方法,以期为微藻科学研究和开发利用提供理论依据。样品前处理过程中皂化的方法主要有氢氧化钠甲醇法、氢氧化钾甲醇法等。甲酯化的方法主要有酸催化法(盐酸甲醇法、硫酸甲醇法、三氟化硼甲醇法和三氯化硼甲醇法等)、碱催化法(氢氧化钾甲醇法)以及其他一些甲酯化的方法(乙酰氯甲醇法、硅烷化试剂法以及重氮甲烷法等);其中乙酰氯甲醇法可直接对干燥的微藻样品进行甲酯化处理而得到脂肪酸甲酯,具有操作简便、用时短、效果好的优点。脂肪酸甲酯的萃取方法主要有正己烷萃取法、石油醚萃取法、戊烷萃取法以及其他一些萃取方法(乙醚、环己烷、乙酸乙酯等)。上述各种处理方法各有优缺点及适用范围,在实际应用时,根据实际需要选择合适的测定方法。
Abstract：Determining microalgal fatty acid composition includes pretreating(lipid extraction,saponification,methyl esterification and fatty acid methyl esters extraction) and quantifying fatty acid methyl esters with gas chromatography and deducing fatty acids and their content.In this review,diverse methods of saponification,methyl esterification and fatty acid methyl esters extraction at sample pretreatment step are compared in order to facilitate theoretical and practical researches of microalgae.Sodium hydroxide methanol and potassium hydroxide methanol methods and among others are available for saponification.Acid catalytic actions with hydrochloric acid methanol,sulfuric acid methanol,boron trifluoride methanol and boron trichloride methanol,base catalytic action with potassium hydroxide and other catalytic actions with acetyl chloride methanol,silane reagent and diazomethane are available for methyl esterification.Of these methods,acetyl chloride methanol is highly performing;it is convenient for use,time saving and highly effective.With this method,fatty acids in dry microalgal powder can be methyl esterified.Hexane,petroleum ether and pentane extractions and among other extractions with ether,cyclohexane,ethyl acetate and so on are available for fatty acid methyl esters extraction.The methods mentioned above each holds advantages and limitations at the same time,which should be evaluated ahead of use.
[1]邹树平,吴玉龙,杨明德,等.微藻的综合开发利用[J].水产科学,2007,26(3):179-181.
[2]梁英,麦康森.微藻EPA和DHA的研究现状及前景[J].水产学报,2000,24(3):289-296.
[3]Yoo C,Jun S Y,Lee J Y,et al.Selection of microalgae for lipid production under high levels carbon dioxide[J].Bioresource tech-nology,2010,101(1):71-74.
[4]Damiani M C,Popovich C A,Constenla D,et al.Lipid analysis in Haematococcus pluvialis to assess its potential use as a biodiesel feedstock[J].Bioresource technology,2010,101(11):3801-3807.
[5]Knothe G.Dependence of biodiesel fuel properties on the structure of fatty acid alkyl esters[J].Fuel Processing Technology,2005,86(10):1059-1070.
[6]余珠花.气相色谱法中油脂脂肪酸衍生化方法及其选择[J].粮食加工,2004,29(6):64-66.
[7]彭兴跃,王大志,徐立,等.毫克级微藻样品中脂肪酸的分离及测定[J].台湾海峡,1998,17(3):289-293.
[8]陆开宏,林霞,钱云霞.三角褐指藻不同藻株脂肪酸组成的研究[J].中国水产科学,2000,7(1):20-24.
[9]Desvilettes C,Bourdier G,Breton J C.On the occurrence of a possible bioconversion of linolenic acid into docosahexaenoic acid by the copepod Eucyclops serrulatus fed on microalgae[J].Jour-nal of plankton research,1997,19(2):273-278.
[10]Ishida Y,Hiragushi N,Kitaguchi H,et al.A highly CO2‐tol-erant diatom,Thalassiosira weissflogii H1,enriched from coastal sea,and its fatty acid composition[J].Fisheries Science,2000,66(4):655-659.
[11]Lavaniegos B E,Lopez-Cortes D.Fatty acid composition and community structure of plankton from the San Lorenzo Channel,Gulf of California[J].Estuarine,Coastal and Shelf Science,1997,45(6):845-854.
[12]Cho E S,Kim G Y,Choi B D,et al.A comparative study of the harmful dinoflagellates Cochlodinium polykrikoides and Gyro-dinium impudicum using transmission electron microscopy,fatty acid composition,carotenoid content,DNA quantification andgene sequences[J].Botanica Marina,2001,44(1):57-66.
[13]黄冠华,陈峰.环境因子对异养小球藻脂肪酸组分含量和脂肪总酸产量的影响[J].可再生能源,2009,27(3):65-69.
[14]Yang H L,Lu C K,Chen S F,et al.Isolation and characteriza-tion of Taiwanese heterotrophic microalgae:screening of strains for docosahexaenoic acid(DHA)production[J].Marine Biotech-nology,2010,12(2):173-185.
[15]梁英,麦康森,孙世春,等.对筒柱藻属33株总脂含量及脂肪酸组成的比较研究[J].青岛海洋大学学报:自然科学版,1999,29(3):457-462.
[16]Liang Y,Mai K S,Sun S C.Effects of harvest stage on the total lipid and fatty acid composition of four Cylindrotheca strains[J].Chinese Journal of Oceanology and Limnology,2002,20(2):157-161.
[17]Liang Y,Mai K S,Sun S C.Effect of NaNO3Concentrations on the Growth and Fatty Acid Compositions of Nitzschia closterium and Chaetoceros gracilis[J].Marine Science Bulletin,2003,5(1):69-74.
[18]Liang Y,Mai K S.Effect of growth phase on the fatty acid com-positions of four species of marine diatoms[J].Journal of Ocean University of China(English Edition),2005,4(2):157-162.
[19]Liang Y,Mai K S,Sun S C.Differences in growth,total lipid content and fatty acid composition among60clones of Cylin-drotheca fusiformis[J].Journal of Applied Phycology,2005,17(1):61-65.
[20]Kumari P,Kumar M,Gupta V,et al.Tropical marine macroal-gae as potential sources of nutritionally important PUFAs[J].Food Chemistry,2010,120(3):749-757.
[21]李宪璀,范晓,韩丽君,等.中国黄、渤海常见大型海藻的脂肪酸组成[J].海洋与湖沼,2002,33(2):215-224.
[22]Rowland S J,Allard W G,Belt S T,et al.Factors influencing the distributions of polyunsaturated terpenoids in the diatom,Rhizosolenia setigera[J].Phytochemistry,2001,58(5):717-728.
[23]吴瑞珊,魏东.Fe对眼点拟微球藻的生长和脂肪酸组成的影响[J].海洋科学,2008,32(7):93-96.
[24]刘平怀,李兰芝.三种螺旋藻的脂肪酸组成的比较研究[J].热带作物科技,1999(3):48-49.
[25]Gapasin R,Duray M N.Effects of DHA-enriched live food on growth,survival and incidence of opercular deformities in milkfish(Chanos chanos)[J].Aquaculture,2001,193(1-2):49-63.
[26]梁惠,冷凯良,贺娟,等.两种海藻物提取物的制备及脂肪酸GC-MS分析[J].食品科学,2005,26(12):184-186.
[27]马夏军,洪爱华,梁智渊,等.中国南海总状蕨藻中脂肪酸和甾醇GC-MS分析[J].广东化工,2005,32(4):19-21.
[28]刘梅芳,王海英.氮源及其浓度对三角褐指藻生长及其脂肪酸组成的影响[J].中南民族大学学报:自然科学版,2008,27(2):32-35.
[29]吕冬伟,宋茜,王旭晨.小球藻和三角褐指藻中脂肪酸和长链烷烃组成的比较研究[J].海洋科学,2009,33(5):28-32.
[30]Xu X Q,Beardall J.Effect of salinity on fatty acid composition of agreen microalga from an antarctic hypersaline lake[J].Phyto-chemistry,1997,45(4):655-658.
[31]蒋汉明,张凤珍,孙凌云,等.Fe3+对三角褐指藻生长和脂肪酸组成的影响[J].食品科学,2005,26(6):43-46.
[32]徐继林,严小军,王晓申,等.巴夫藻脂肪酸和甾醇的同步测定研究[J].海洋科学,2004,28(4):19-24.
[33]Riebesell U,Revill A T,Holdsworth D G,et al.The effects of varying CO2concentration on lipid composition and carbon iso-tope fractionation in Emiliania huxleyi[J].Geochimica et Cos-mochimica Acta,2000,64(24):4179-4192.
[34]徐继林,严小军,周成旭,等.一种海洋席藻淡化过程中脂肪酸组成的应急变化[J].水生生物学报,2004,28(4):374-379.
[35]Fabregas J,Aran J,Morales E D,et al.Modification of sterol concentration in marine microalgae[J].Phytochemistry,1997,46(7):1189-1191.
[36]Ghazala B,Naila B,Shameel M.Fatty acids and biological activi-ties of crude extracts of freshwater algae from Sindh[J].Paki-stanJournal of Botany,2010,42(2):1201-1212.
[37]李润,周春玉.金藻Isochrysis Galbana中DHA的分离与分析[J].应用化学,2001,18(6):454-456.
[38]Blanchemain A,Grizeau D.Eicosapentaenoic acid content of Skeletonema costatum as a function of growth and irradiance;re-lation with chlorophyll a content and photosynthetic capacity[J].Journal of experimental marine biology and ecology,1996,196(1-2):177-188.
[39]周秋香,张德强,孙汉文.三角褐指藻的培养条件研究[J].河北大学学报:自然科学版,2001,21(1):68-71.
[40]林学政,李光友.11种微藻脂类和EPA/DHA组成的研究[J].黄渤海海洋,2000,18(2):36-40.
[41]李文权,王宪,陈清花,等.超声波对湛江等鞭金藻生长和脂肪酸组成的影响[J].海洋学报,2002,24(3):94-100.
[42]Schneider J C,Roessler P.Radiolabeling studies of lipid and fatty acid in Nannochloropsis(Eustigmatophyceae),an oleaginous ma-rine alga[J].Journal of phycology,1994,30(4):594-598.
[43]Sheng J,Vannela R,Rittmann B E.Evaluation of methods to extract and quantify lipids from Synechocystis PCC6803[J].Bioresource Technology,2011,102(2):1697-1703.
[44]Dembitsky V M,Rezankov H,Rezanka T,et al.Variability of the fatty acids of the marine green algae belonging to the genus Codium[J].Biochemical Systematics and Ecology,2003,31(10):1125-1145.
[45]Tatsuzawa H,Takizawa E.Changes in lipid and fatty acid com-position of Pavlova lutheri[J].Phytochemistry,1995,40(2):397-400.
[46]Nichols D S,Hart P,Nichols P D,et al.Enrichment of the roti-fer Brachionus plicatilis fed an Antarctic bacterium containing polyunsaturated fatty acids[J].Aquaculture,1996,147(1-2):115-125.
[47]Gelin F,Volkman J K,De Leeuw J W,et al.Mid-chain hydroxy long-chain fatty acids in microalgae from the genus Nannochlo-ropsis[J].Phytochemistry,1997,45(4):641-646.
[48]Volkman J K,Dunstan G A,Jeffrey S W,et al.Fatty acids from microalgae of the genus Pavlova[J].Phytochemistry,1991,30(6):1855-1859.
[49]Mansour M P,Volkman J K,Holdsworth D G,et al.Very-long-chain(C28)highly unsaturated fatty acids in marine dinoflagel-lates[J].Phytochemistry,1999,50(4):541-548.
[50]寇秀颖,于国萍.脂肪和脂肪酸甲酯化方法的研究[J].食品研究与开发,2005,26(2):46-47.
[51]Koven W,Barr Y,Lutzky S,et al.The effect of dietary arachi-donic acid(20:4n-6)on growth,survival and resistance to han-dling stress in gilthead seabream(Sparus aurata)larvae[J].Aquaculture,2001,193(1-2):107-122.
[52]Wahlen B D,Willis R M,Seefeldt L C.Biodiesel production by simultaneous extraction and conversion of total lipids from mi-croalgae,cyanobacteria,and wild mixed-cultures[J].Biore-source technology,2011,102(3):2724-2730.
[53]刘建国,刘伟.雪藻高密度连续培养中生物量和花生四烯酸的高产率[J].海洋与湖沼,2002,33(5):499-508.
[54]Mohan S V,Devi M P,Mohanakrishna G,et al.Potential of mixed microalgae to harness biodiesel from ecological water-bodies with simultaneous treatment[J].Bioresource Technolo-gy,2011,102(2):1109-1117.
[55]Mclarnon-Riches C J,Rolph C E,Greenway D L A,et al.Effects of environmental factors and metals on Selenastrum capri-cornutum lipids[J].Phytochemistry,1998,49(5):1241-1247.
[56]Jaschinski S,Brepohl D,Sommer U.The trophic importance of epiphytic algae in a freshwater macrophyte system(Potamogeton perfoliatus L.):stable isotope and fatty acid analyses[J].A-quatic Sciences-Research Across Boundaries,2011,73(1):91-101.
[57]de Lange H,Van D.Effects of UVB‐irradiated algae on life history traits of Daphnia pulex[J].Freshwater Biology,1997,38(3):711-720.
[58]Caers M,Coutteau P,Sorgeloos P.Incorporation of different fatty acids,supplied as emulsions or liposomes,in the polar and neutral lipids of Crassostrea gigas spat[J].Aquaculture,2000,186(1-2):157-171.
[59]Freites L,Labarta U,Fernandez-Reiriz M J.Evolution of fatty acid profiles of subtidal and rocky shore mussel seed(Mytilus galloprovincialis,Lmk.).Influence of environmental parame-ters[J].Journal of Experimental Marine Biology and Ecology,2002,268(2):185-204.
[60]冯福应,杨文,林伟,等.海洋微藻Chlorellasp.1061的脂类特征与分类探讨[J].海洋与湖沼,2004,35(5):453-458.
[61]Levine R B,Costanza-Robinson M S,Spatafora G A.Neochloris oleoabundans grown on anaerobically digested dairy manure for concomitant nutrient removal and biodiesel feedstock production[J].Biomass and Bioenergy,2011,35(1):40-49.
[62]Wang L,Li Y,Chen P,et al.Anaerobic digested dairy manure as a nutrient supplement for cultivation of oil-rich green microal-gae Chlorella sp[J].Bioresource Technology,2010,101(8):2623-2628.
[63]Viso A C,Marty J C.Fatty acids from28marine microalgae[J].Phytochemistry,1993,34(6):1521-1533.
[64]Gressler V,Yokoya N S,Fujii M T,et al.Lipid,fatty acid,protein,amino acid and ash contents in four Brazilian red algae species[J].Food Chemistry,2010,120(2):585-590.
[65]Nanton D A,Lall S P,Mcniven M A.Effects of dietary lipid lev-el on liver and muscle lipid deposition in juvenile haddock,Mel-anogrammus aeglefinus L.[J].Aquaculture Research,2001,32(s1):225-234.
[66]Nanton D A,Castell J D.The effects of dietary fatty acids on thefatty acid composition of the harpacticoid copepod,Tisbe sp.,for use as a live food for marine fish larvae[J].Aquaculture,1998,163(3-4):251-261.
[67]Copeman L A,Parrish C C,Brown J A,et al.Effects of docosa-hexaenoic,eicosapentaenoic,and arachidonic acids on the early growth,survival,lipid composition and pigmentation of yel-lowtail flounder(Limanda ferruginea):a live food enrichment experiment[J].Aquaculture,2002,210(1-4):285-304.
[68]Napolitano G E,Heras H,Stewart A J.Fatty acid composition of freshwater phytoplankton during a red tide event[J].Bio-chemical systematics and ecology,1995,23(1):65-69.
[69]Volkman J K,Jeffrey S W,Nichols P D,et al.Fatty acid and lipid composition of10species of microalgae used in mariculture[J].Journal of Experimental Marine Biology and Ecology,1989,128(3):219-240.
[70]Berge J P,Gouygou J P,Dubacq J P,et al.Reassessment of lip-id composition of the diatom,Skeletonema costatum[J].Phyto-chemistry,1995,39(5):1017-1021.
[71]Xu X Q,Beardall J,Hallam N D.Modification of fatty acid com-position in halophilic antarctic microalgae[J].Phytochemistry,1998,49(5):1249-1252.
[72]Mooney H M,Cooney J J,Baisden C M,et al.Fatty acids of Dunaliella tertiolecta and Skeletonema costatum grown in the presence of Phenyltin compounds[J].Botanica marina,1995,38(1-6):423-430.
[73]Bao X,Ohlrogge J.Supply of fatty acid is one limiting factor in the accumulation of triacylglycerol in developing embryos[J].Plant physiology,1999,120(4):1057.
[74]Renaud S M,Parry D L,Thinh L V,et al.Effect of light inten-sity on the proximate biochemical and fatty acid composition of Isochrysis sp.and Nannochloropsis oculatafor use in tropical aq-uaculture[J].Journal of Applied Phycology,1991,3(1):43-53.
[75]Wikfors G H,Patterson G W,Ghosh P,et al.Growth of post-set oysters,Crassostrea virginica,on high-lipid strains of algal flagellates Tetraselmis spp.[J].Aquaculture,1996,143(3-4):411-419.
[76]卢丽娜,孙利芹,田焕玲,等.32株海洋微藻总脂含量及其脂肪酸组成的研究[J].中国油脂,2009,34(11):68-73.
[77]孙利芹,杨林涛.环境因子对球等鞭金藻脂肪酸含量和组成的影响[J].食品研究与开发,2006,27(5):11-13.
[78]Xu X Q.Fatty acids of six Codium species from southeast Aus-tralia[J].Phytochemistry,1998,48(8):1335-1339.
[79]张秋会,马莺.气相色谱法测定海洋微藻中二十碳五烯酸和二十二碳六烯酸[J].色谱,2004,22(6):662.
[80]张秋会,马莺.高产EPA和DHA藻株的筛选[J].西部粮油科技,2003,28(6):51-53.
[81]Krohn B J,Mcneff C V,Yan B,et al.Production of algae-based biodiesel using the continuous catalytic Mcgyan(R)process[J].Bioresource technology,2011,102(1):94-100.
[82]Okuyama H,Morita N,Kogame K.Occurrence of octadecapen-taenoic acid in lipids of a cold stenothermic alga,prymnesiophyte strain B1[J].Journal of phycology,1992,28(4):465-472.
[83]Liang Y,Beardall J,Heraud P.Effects of nitrogen source and UV radiation on the growth,chlorophyll fluorescence and fattyacid composition of Phaeodactylum tricornutum and Chaetoceros muelleri(Bacillariophyceae)[J].Journal of Photochemistry and Photobiology B:Biology,2006,82(3):161-172.
[84]Liang Y,Beardall J,Heraud P.Changes in growth,chlorophyll fluorescence and fatty acid composition with culture age in batch cultures of Phaeodactylum tricornutumand Chaetoceros muelleri(Bacillariophyceae)[J].Botanica Marina,2006,49(2):165-173.
[85]Liang Y,Beardall J,Heraud P.Effect of UV radiation on growth,chlorophyll fluorescence and fatty acid composition of Phaeodactylum tricornutum and Chaetoceros muelleri(Bacillariophyceae)[J].Phyco-logia,2006,45(6),605-615.
[86]Lepage G,Roy C C.Improved recovery of fatty acid through di-rect transesterification without prior extraction or purification.[J].Journal of Lipid Research,1984,25(12):1391.
[87]Lin Q,Lin J.Effects of nitrogen source and concentration on bio-mass and oil production of a Scenedesmus rubescens like microalga[J].Bioresource Technology,2011,102(2):1615-1621.
[88]廖启斌,李文权,陈清花,等.营养盐对三角褐指藻脂肪酸含量与百分组成的影响[J].海洋环境科学,2000,19(2):6-9.
[89]蔡阿根,郑爱榕,李文权,等.海洋微藻中脂肪酸的气相色谱分析[J].海洋技术,1998,17(4):64-68.
[90]Nitsan Z,Mokady S,Sukenik A.Enrichment of poultry products withω3fatty acids by dietary supplementation with the algaNan-nochloropsis and mantur oil[J].Journal of agricultural and food chemistry,1999,47(12):5127-5132.
基本信息:
DOI：10.16441/j.cnki.hdxb.2013.04.003
中图分类号:Q946
引用信息:
[1]梁英,石伟杰,田传远.微藻脂肪酸组成测定方法概述[J],2013,43(04):21-28.DOI:10.16441/j.cnki.hdxb.2013.04.003.
基金信息:
“十二五”国家科技支撑计划项目(2011BAD14B01);; 国家自然科学基金项目(40976076)资助