Journal of the American Oil Chemists' Society

, Volume 89, Issue 2, pp 189–198 | Cite as

Optimization of an Analytical Procedure for Extraction of Lipids from Microalgae

  • Eline RyckeboschEmail author
  • Koenraad Muylaert
  • Imogen Foubert
Original Paper


An optimized procedure for extraction of total and non-polar lipids from microalgae is proposed. The effects of solvent, pretreatment (lyophilization, inactivation of lipases, and addition of antioxidants) and cell-disruption (liquid nitrogen, sonication, and bead beating) on total lipid content, lipid class, and fatty acid composition were examined. Chloroform–methanol 1:1 was shown to be the best solvent mixture for extraction of total lipids from microalgae. When performing this extraction, lyophilized algae can be used, no pretreatment with isopropanol to inactivate the lipases is needed and addition of antioxidants is not necessary. Furthermore, cell-disruption is not essential, although in that case two extractions must be performed in series to ensure that, irrespective of the microalgal species, all lipids are extracted. Determination of non-polar lipid content should be performed by separation of the total lipid extract on an SPE column. Extraction using petroleum ether is only appropriate when a bead beater is used for pretreatment.


Algae Lipid extraction Solvent mixture Pretreatment Cell-disruption Fat content 



The research presented in this paper was financially supported by the Institute for the Promotion of Innovation by Science and Technology—Strategic Basic Research (IWT-SBO) project Sunlight and K.U.Leuven Kulak. We acknowledge IS-X (Interscience, Louvain-la-Neuve, Belgium) for the use of the GC–MS at their demolab.


  1. 1.
    Guschina IA, Harwood JL (2006) Lipids and lipid metabolism in eukaryotic algae. Prog Lipid Res 45:160–186CrossRefGoogle Scholar
  2. 2.
    Folch J, Lees M, Stanley GHS (1957) A simple method for the isolation and purification of total lipides from animal tissues. J Biol Chem 226:497–509Google Scholar
  3. 3.
    Bligh EG, Dyer WJ (1959) A rapid method of total lipid extraction and purification. Can J Biochem Physiol 37:911–917CrossRefGoogle Scholar
  4. 4.
    Iverson SJ, Lang SLC, Cooper MH (2001) Comparison of the Bligh and Dyer and Folch methods for total lipid determination in a broad range of marine tissue. Lipids 36:1283–1287CrossRefGoogle Scholar
  5. 5.
    Guckert JB, Cooksey KE, Jackson LL (1988) Lipid solvent systems are not equivalent for analysis of lipid classes in the micro eukaryotic green alga, Chlorella. J Microbiol Methods 8:139–149CrossRefGoogle Scholar
  6. 6.
    Lee SJ, Yoon B-D, Oh H-M (1998) Rapid method for the determination of lipid from the green alga Botryococcus braunii. Biotechnol Tech 12:553–556CrossRefGoogle Scholar
  7. 7.
    Grima EM, Medina AR, Giménez AG, Pérez JAS, Camacho FG, Sanchez JLG (1994) Comparison between extraction of lipids and fatty acids from microalgal biomass. JAOCS 71:955–959CrossRefGoogle Scholar
  8. 8.
    Matyash V, Liebisch G, Kurzchalia TV, Shevchenko A, Schwudke D (2008) Lipid extraction by methyl-tert-butyl ether for high-throughput lipidomics. J Lipid Res 49:1137–1146CrossRefGoogle Scholar
  9. 9.
    Nielsen SS (2003) Food Analysis, 3rd edn. Kluwer Academic/Plenum, New YorkGoogle Scholar
  10. 10.
    Ausborn M, Nuhn P, Schreier H (1992) Stabilization of liposomes by freeze–thaw––and lyophilization techniques: problems and opportunities. Eur J Pharm Biopharm 38:133–139Google Scholar
  11. 11.
    Christie WW (2003) Lipid Analysis: isolation, separation, identification and structural analysis of lipids, 3rd edn. The Oily Press, BridgewaterGoogle Scholar
  12. 12.
    Wrolstad RE, Acree TE, Decker EA, Penner MH, Reid DS, Schwartz SJ, Shoemaker CF, Smith DM, Sporns P (2005) Handbook of food analytical chemistry. Wiley, HobokenGoogle Scholar
  13. 13.
    Han D, Walde P, Luisi PL (1990) Dependence of lipase activity on water content and enzyme concentration in reverse micelles. Biocat Biotrans 4:153–161CrossRefGoogle Scholar
  14. 14.
    Li H-B, Cheng K-W, Wong C-C, Fan K-W, Chen F, Jiang Y (2007) Evaluation of antioxidant capacity and total phenolic content of different fractions of selected microalgae. Food Chem 102:771–776CrossRefGoogle Scholar
  15. 15.
    Lee J-Y, Yoo C, Jun S-Y, Ahn C-Y, Oh H-M (2010) Comparison of several methods for effective lipid extraction from microalgae. Bioresour Technol 101:S75–S77CrossRefGoogle Scholar
  16. 16.
    Guillard RRL, Lorenzen CJ (1972) Yellow-green algae with chlorophyllide c. J Phycol 8:10–14Google Scholar
  17. 17.
    Borowitzka MA, Volcani BE (1978) The polymorphic diatom Phaeodactylum tricornutum: ultrastructure of its morphotypes. J Phycol 14:10–21CrossRefGoogle Scholar
  18. 18.
    Grossi V, Blokker P, Damste JSS (2001) Anaerobic biodegradation of lipids of the marine microalga Nannochloropsis salina. Org Geochem 32:795–808CrossRefGoogle Scholar
  19. 19.
    Takeda H (1995) Cell wall sugars of some Scenedesmus species. Phytochem 42:673–675CrossRefGoogle Scholar
  20. 20.
    Corre G, Templier J, Largeau C, Rousseau B, Berkaloff C (1996) Influence of cell wall composition on the resistance of two Chlorella species (Chlorophyta) to detergents. J Phycol 32:584–590CrossRefGoogle Scholar
  21. 21.
    Takeda H (1991) Sugar composition of the cell wall and the taxonomy of Chlorella (Chlorophyceae). J Pycol 27:224–232Google Scholar
  22. 22.
    van Eykelenburg C (1977) On the morphology and ultrastructure of the cell wall of Spirulina platensis. Antonie van Leeuwenhoek 43:89–99CrossRefGoogle Scholar
  23. 23.
    van Eykelenburg C (1978) A glucan from the cell wall of the cyanobacterium Spirulina platensis. Antonie van Leeuwenhoek 44:321–327CrossRefGoogle Scholar
  24. 24.
    Chen G-Q, Jiang Y, Chen F (2007) Fatty acid and lipid class composition of the eicosapentaenoic acid-producing microalga Nitzschia laevis. Food Chem 104:1580–1585CrossRefGoogle Scholar
  25. 25.
    Kangani CO, Kelley DE, DeLany JP (2008) New method for GC/FID and GC–C-IRMS analysis of plasma free fatty acid concentration and isotopic enrichment. J Chrom B 873:95–101CrossRefGoogle Scholar
  26. 26.
    Spanner S (1973) Separation and analysis of phospholipids. In: Ansell GB, Hawthorne JN, Dawson RMC (eds) Form and function of phospholipids. Elsevier, Amsterdam, pp 43–65Google Scholar
  27. 27.
    Hanahan DJ (1960) Lipide Chemistry. Wiley, New YorkGoogle Scholar
  28. 28.
    Hamilton S, Hamilton RJ, Sewell PA (1992) Extraction of lipids and derivative formation. In: Hamilton RJ, Hamilton S (eds) Lipid analysis, a practical approach. Oxford University Press, New York, pp 13–64Google Scholar
  29. 29.
    Lee CM, Trevino B, Chaiyawat M (1996) A simple and rapid solvent extraction method for determining total lipids in fish tissue. J AOAC Int 79:487–492Google Scholar
  30. 30.
    Henderson RK, Baker A, Parsons SA, Jefferson B (2008) Characterisation of algogenic organic matter extracted from cyanobacteria, green algae and diatoms. Wat Res 42:3435–3445CrossRefGoogle Scholar

Copyright information

© AOCS 2011

Authors and Affiliations

  • Eline Ryckebosch
    • 1
    Email author
  • Koenraad Muylaert
    • 2
  • Imogen Foubert
    • 1
  1. 1.K. U. Leuven Kulak, Research Unit Food and Lipids, Department of Molecular and Microbial SystemsLeuven Food and Nutrition Research Centre (LFoRCe)KortrijkBelgium
  2. 2.K. U. Leuven Kulak, Lab Aquatic BiologyBiology DepartmentKortrijkBelgium

Personalised recommendations