Food Lipids: Chemistry, Nutrition, and Biotechnology

Figure 1 Diagrammatic representation of globules (particles) in various types of emulsions.
In the ball-and-stick model of amphiphilic (surfactant) molecules, the ball represents the polar
head group and the stick the hydrophobic region of the molecule.
et al. (37) and Sanchez-Ferrer and Garcia-Carmona (38) can be consulted for more
information on microemulsions.
B. Normal Phase Emulsions
Most lipase assays for hydrolytic reactions are carried out in normal phase emulsions
whereby the water-insoluble oil substrate is emulsified by sonication into a buffered
aqueous enzyme preparation. The emulsion is stabilized with an emulsifier and the
aqueous phase often contains Ca 2� . The specific ingredients, buffer types, pH, and
relative amounts of components vary widely with the author and specific lipase being
assayed. An example of such an assay is as follows: 2 mL 0.2 M Tris maleate–
NaOH buffer (pH 8.2), 1 mL 0.03 M Ca2Cl, 5 mL distilled water, 1 mL olive oil,
and 1 mL enzyme solution. The emulsion is incubated at 30�C for 60 minutes or an
appropriate time depending on lipase activity, and the reaction is stopped with 20
mL of acetone ethanol (1:1). Common emulsifiers used in lipase assays include
polyvinyl alcohol and gum arabic at 1–2% by volume of assay mixture. Hydrolytic
activity is most often determined by titration of the fatty acid products of the reaction
with NaOH. Other methods and assay conditions have been reviewed by Jensen (39).
C. Invert Emulsions
Using lipases from several fungi, Mozaffar and Weete (40) reported a reaction mixture
containing 5 mL olive oil, 0.1 mL 520 mM taurocholic acid in 50 mM sodium
phosphate buffer (pH 7.5), and 0.1 mL enzyme preparation. Final concentrations of
taurocholic acid and water in the 5.2 mL of reaction mixture were 10 mM and 4%.
The mixture was emulsified by vortexing for about 30 seconds and incubated at 45�C
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