Food Lipids: Chemistry, Nutrition, and Biotechnology

the ionic groups of lipids. Thus, a polymer with fixed cations will bind anionic lipids
from a mixture, provided that the pH of the solvent mixture allows ionization of the
anionic groups. At the same time, the concentration of nonlipid anions in the solvent
mixture should not compete for all of the fixed ions [84]. Some of the ion exchange
chromatographic materials commonly used for lipid analysis are given in Table 2.
Diethylaminoethyl (DEAE) cellulose is used for separation of lipid classes, and triethylaminoethyl
(TEAE) cellulose is useful for separation of lipids having only ionic
carboxyl groups (e.g., fatty acids, bile acids, gangliosides) or phosphatidylethanolamine
from ceramide polyhexosides [83]. In polar lipid analysis, DEAE cellulose in
the acetate form is the most frequently used anion exchange material. It is most
effective in the pH range 3–6, and often separation of polar lipid is achieved by
stepwise elution with ammonium acetate buffer in water–ethanol. The cation exchanger
carboxymethylcellulose (CMC) as its sodium salt has been used occasionally
over the same pH range for separation of phospholipids [84].
Complexing the adsorbent material with silver nitrate enables separation of
lipid mixtures according to the number, position and cis and trans isomerism of
double bonds in unsaturated fatty acids and their derivatives. Use of borate treatment
of the column material complexes the compounds containing hydroxyl groups on
adjacent carbon atoms and assists the separation of glycolipids [84]. Complexing of
adsorbent materials is discussed in detail in the section on thin-layer chromatography.
Nowadays commercial columns are prepacked with a variety of solid stationary
phases, which are available for separation of lipid classes and may be referred to as
solid phase extraction (SPE) columns. SPE consumes less time, solvent, and packing
material than does classical column chromatography [85]. SPE can be used for isolation,
concentration, purification, and fractionation of analytes from complex
mixtures [86,87]. Aminopropyl-bonded phase has been used for separation of total
lipids in lipid classes obtained from different sources [88–91].
Immunoaffinity column chromatography has been used for isolation and purification
of apolipoproteins. Ligand molecules (antibody; antibody to apolipoprotein
or antigen; purified apolipoprotein) are immobilized on the solid support (matrix)
and bind to corresponding target molecules (antigen or antibody) in a mixture of
macromolecules. The bound target molecule (e.g., apolipoprotein) can then be desorbed
from the ligands and eluted in the purified form using appropriate buffers
[92].
Table 2 Ion Exchange Chromatography Materials Used in Lipid Analysis
Ionizing group Commercial classification Analytical use
—(CH 2) 2N � H(C 2H 5) 2
(diethylaminoethyl)
—(CH 2) 2N � (C 2H 5) 3
(triethylaminoethyl)
—CH2COO— (carboxymethyl)
Source: Adapted from Ref. 84.
Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.
DEAE (anionic exchanger) Anionic lipids (phospholipids,
sulfolipids, sialoglycolipids,
fatty acids)
TEAE (anionic exchanger) Anionic lipids
CM (cationic exchanger) Phospholipid mixtures