The esterification degree (ED) represents the percentage of initial FFAs
consumed by methylation. ED was determined by acid-base titration after checking
that the results were similar to those obtained by TLC and GC, which take longer. The
samples were diluted with acetone (ratio 1:1 v/v) and phenolphthalein was used as
indicator. The samples from esterification of microalgal FFAs were washed with
alumina, diluted with acetone (fatty acid/acetone ratio 1:20 v/v) and titrated, using
thymol blue as indicator. Previously the acidity of the initial reaction mixture had
been determined under the same conditions in which the acidity was measured,
once the reaction had finished. ED was calculated by the equation