American Journal of Biochemistry and Biotechnology 7 (2): 70-73, 2011 ISSN 1553-3468 © 2011 Science Publications Corresponding Author: Robert D. Long, Department of Physical Sciences, Eastern New Mexico University, Station 33, Portales, NM, 88130, USA 70 Mixed-Polarity Azeotropic Solvents for Efficient Extraction of Lipids from Nannochloropsis Microalgae Robert D. Long and Elseddik Abdelkader Department of Physical Sciences, Eastern New Mexico University, Station 33, Portales, NM, 88130, USA Abstract: Problem statement: The extraction of lipids from microalgae for biofuel production is a significant problem due to the intractability of their cell wall. The aim of this study of lipid extraction from Nannochloropsis microalgae was to examine the relative efficiencies of some commonly used solvent systems and test the use of co-boiling solvents in Soxhlet extraction as an improved method. Approach: We conducted a series of trials of lipid extraction from dried Nannochloropsis microalgae powder by conventional methods and compared the results to low boiling point mixed-polarity azeotropic solvent mixtures in Soxhlet extraction (a technique which has not previously been reported). Results: We found that in conventional room temperature solvent extractions, the chloroform/methanol protocols gave higher yields of lipids from Nannochloropsis than a hexane/isopropanol mixture. In testing Soxhlet extractions, we found that superior results could be obtained with hydrocarbon/alcohol azeotropic mixtures compared to hydrocarbon only solvents (i.e., hexane, cyclohexane), especially with a cyclohexane/butanol (9:1 ratio) mixture. Conclusion: The latter results indicate the potential for performing efficient microalgae lipid extractions with non- halogenated solvent systems in the future (a greener alternative). Key words: Lipid extraction, algal oil, soxhlet extractions, solvent extractions, solvent systems, extraction protocols, azeotropic solvent, soxhlet extraction, nannochloropsis microalgae, algal biomass INTRODUCTION Biodiesel is an important renewable transportation fuel that can be made from vegetable oils, animal fats, used cooking oil, or algal oil (Demirbas, 2007; Pienkos and Darzins, 2009). Microalgae are efficient photosynthetic factories that have the ability to convert atmospheric carbon (existing as CO 2 ) to lipids, especially to Triacylglycerols (TAG’s) (Hu et al., 2008). The extraction of these lipids from algal biomass is a necessary first step in their conversion to biodiesel or other biofuels (Sharif Hossain et al., 2008; Schenk et al., 2008; Chisti, 2007). Extraction of oils from microalgae is a much more difficult problem than extraction of oils from oilseed. Microalgae are single-cell organisms with extremely tough cell walls that can be difficult to disrupt (Sheehan et al., 1998). Thus, techniques other than those used for oilseeds involving ultrasonication, microwaves, electrochemical cell wall disruption, electromechanical cell wall rupture and supercritical fluid extraction are all currently being studied relative to microalgae extraction, in addition to the more conventional solvent extraction methods (Lee et al., 2010; Samorì et al., 2010). Apart from these newer methods, the most common techniques for lipid extraction from microalgae in current use involve chloroform/methanol mixtures or hexane in solvent or Soxhlet extraction. We performed a comparison of different solvent systems using these traditional extraction techniques in order to judge their relative effectiveness. We also wanted to evaluate whether other non- chlorinated solvent systems could be effective, thus eliminating environmental concerns and down-stream processing problems associated with chlorinated solvents. For this purpose, we examined the use of mixed-polarity (hydrocarbon/alcohol mixture) solvent systems in ratios equivalent to the azeotropic mixture expected to be formed from them in Soxhlet extractions as an alternative technique for lipid extraction. MATERIALS AND METHODS Preparation: Two batches of frozen Nannochloropsis microalgae paste were donated by Center of Excellence