To confirm the lipolytic activity, the titrimetric method
with tributyrin (Sigma) as the substrate was also used
according to the method of Stuer et al. (1986). Enzyme
solution (100 mL or about 40 mg of protein) was added
to 20mL of substrate emulsion [10mM of substrate in 2%
(w/v) gum arabic in distilled water and 0.1M CaCl2]. The
reaction mixture was incubated at 37 1C and the release of
free fatty acids was monitored continuously by titration with
10mM NaOH to maintain pH 8.0 with a pH stat (Mettler).