Food Lipids: Chemistry, Nutrition, and Biotechnology

polar solvents, such as alkanols, for their extraction. Therefore, use of a single universal
solvent for extraction of lipids from tissues is not possible. During solvent
extraction van der Waals and electrostatic interactions as well as hydrogen bonds are
broken to different extents; however, covalent bonds remain intact.
Neutral lipids are hydrophobically bound and can be extracted from tissues by
nonpolar solvents, whereas polar lipids, which are bound predominantly by electrostatic
forces and hydrogen bonding, require polar solvents capable of breaking such
bonds. However, less polar neutral lipids, such as TAGs and cholesterol esters, may
also be extracted incompletely with nonpolar solvents, probably due to inaccessibility
of a significant part of these lipids to the solvents. Lipids that are covalently bound
to polypeptide and polysaccharide groups will not be extracted at all by organic
solvents and will remain in the nonlipid residue. Therefore, a hydrolysis step may
be required to release covalently bound lipids to render them fully extractable.
1. Properties of Solvents and Their Mode of Extraction
The type of solvent and the actual method of lipid extraction depend on both the
chemical nature of the sample and the type of lipid extract (e.g., total lipids, surface
lipids of leaves) desired. The most important characteristic of the ideal solvent for
lipid extraction is the high solubility of lipids coupled with low or no solubility of
proteins, amino acids, and carbohydrates. The extracting solvent may also prevent
enzymatic hydrolysis of lipids, thus ensuring the absence of side reactions. The
solvent should readily penetrate sample particles and should have a relatively low
boiling point to evaporate readily without leaving any residues when recovering
lipids. The solvents mostly used for isolation of lipids are alcohols (methanol, ethanol,
isopropanol, n-butanol), acetone, acetonitrile, ethers (diethyl ether, isopropyl
ether, dioxane, tetrahydrofuran), halocarbons (chloroform, dichloromethane), hydrocarbons
(hexane, benzene, cyclohexane, isooctane), or their mixtures. Although solvents
such as benzene are useful in lipid extraction, it is advisable to look for alternative
solvents due to the potential carcinogenicity of such products. Flammability
and toxicity of the solvent are also important considerations to minimize potential
hazards as well as cost and nonhygroscopicity.
Solubility of lipids in organic solvents is dictated by the proportion of the
nonpolar hydrocarbon chain of the fatty acids or other aliphatic moieties and polar
functional groups, such as phosphate or sugar moieties, in their molecules. Lipids
containing no distinguishable polar groups (e.g., TAGs or cholesterol esters) are
highly soluble in hydrocarbon solvents such as hexane, benzene, or cyclohexane and
in more polar solvents such as chloroform or diethyl ether, but remain insoluble in
polar solvents such as methanol. The solubility of such lipids in alcoholic solvents
increases with the chain length of the hydrocarbon moiety of the alcohol; therefore,
they are more soluble in ethanol and completely soluble in n-butanol. Similarly, the
shorter chain fatty acid residues in the lipids have greater solubility in more polar
solvents (e.g., tributyrin is completely soluble in methanol whereas tripalmitin is
insoluble). Polar lipids are only sparingly soluble in hydrocarbon solvents unless
solubilized by association with other lipids; however, they dissolve readily in more
polar solvents, such as methanol, ethanol, or chloroform [4].
2. Extraction Methods with Single Organic Solvent
Diethyl ether and petroleum ether are the most commonly used solvents for extraction
of lipids. In addition, hexane and sometimes pentane are preferred to obtain
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