Enzyme-Assisted Aqueous Extraction of Oleosomes from Soybeans (Glycine max) VIRGINIE N. KAPCHIE,* DAIJING WEI,CATHERINE HAUCK, AND PATRICIA A. MURPHY Department of Food Science and Human Nutrition and Center for Crops Utilization Research, Iowa State University, Ames, Iowa 50011 Oleosomes, with their unique structural proteins, the oleosins, are known to be useful in cosmetics and other emulsion applications. A procedure to fractionate intact oleosomes to produce soybean oil without the use of organic solvents was investigated. Process parameters, enzyme treatment, filtration, cell lysis, and centrifugation, were studied. Successive extractions of the residue, eliminating the filtration step, pressurization, or ultrasonication of soybean flour prior to enzyme treatment and enzyme treatment on the residue, were the key steps. A mixture of Multifect Pectinase FE, Cellulase A, and Multifect CX 13L in equal proportion gave 36.42-63.23% of the total soybean oil from oleosomes, respectively, for 45 and 180 s of blending time, compared to the conventional method with lower yields (34.24 and 28.65%, respectively, for 45 and 180 s of blending time). Three successive extractions of the residue increased the oil yield to a maximum of 84.65% of the total soybean oil recovered in intact oleosomes. The percentage of lipid in the supernatant fraction decreased to a minimum value of 0.33% with the use of the enzymes at a 3% dosage. The results are considered to be useful for developing large-scale and efficient extraction of oleosomes from soybean. KEYWORDS: Soybean oleosomes; pectinase; cellulase; hemicellulase; oil recovery INTRODUCTION Oleosomes or oil bodies are intracellular plant organelles of about 0.2–2.0 µm in diameter for most oilseeds and 0.2–0.5 µm in the case of soybean (1–3). The oleosomes possess a structure consisting of a triacylglycerol core (94–98% of the dry weight) surrounded by a monolayer of phospholipids (0.6–2% w/w) embedded with oleosins and some minor proteins of higher molecular mass (0.6–4% w/w) (4, 5). Oleosomes are known to be useful in cosmetics and other emulsion applications (6, 7). Oleosomes can be isolated from hydrated seed homogenates by flotation with centrifugation in a sucrose buffer, as their density is similar to that of oil. The conventional method of extracting oleosomes, involving hydrating intact soybean seeds or cotyledons, grinding, filtering, and centrifuging, is frequently used to fractionate intact oleosomes (5, 8–10). Our preliminary investigation determined that the conventional method resulted in intact oleosomes yields of <45% of the total soybean oil. It is obvious that a major contributing factor to the low yield of oleosomes is the inability of the treatment involved to adequately rupture the cellular structure without shearing the oleosome membranes. Therefore, more efficient extraction procedures needed to be developed. Scanning electron microscopy analysis showed that oleosomes from soybeans are enmeshed in a cytoplasmic network com- posed of proteins (11). The spaces between protein bodies in cotyledon cells are then filled with lipid body and the cytoplas- mic network (1, 12). Unlike the cytoplasmic features, which are characterized by the presence of protein and lipid, the walls that surround the plant cell are primarily composed of cellulose, hemicellulose, and lignin in addition to pectin. This suggests that enzyme preparations capable of attacking cell walls must contain cellulases, hemicellulases, and pectinases (13, 14). Rosenthal et al. (15, 16) have reviewed the main aspects relating to aqueous and enzymatic processes for oil extraction and have reported that the disruption of the cell wall of soybean cotyledon during milling operation resulted in an increase in oil and protein extraction yields. Therefore, it could be possible to adapt an enzyme-assisted technique to improve the yield of oleosomes. The objective of this work was to explore the use of enzymes that hydrolyze cell wall components to improve the release of intact oleosomes from soybean. We compared a conventional method of extracting oleosomes, involving hydrating, grinding, filtering, and centrifuging (5, 10), with enzyme-assisted extrac- tion with various modifications in the process to determine the preferable extraction procedure that would enable the release of oleosomes from soybeans. MATERIALS AND METHODS Soybeans. Full-fat soybean flour (commodity soybeans of 2006 crop) was procured from the Center for Crops Utilization Research (CCUR), * Author to whom correspondence should be addressed [telephone (515) 294-4010; fax (515) 294-8181; e-mail vkapchie@iastate.edu]. 1766 J. Agric. Food Chem. 2008, 56, 1766–1771 10.1021/jf0721390 CCC: $40.75  2008 American Chemical Society Published on Web 02/06/2008