Although the reaction velocities and conversions obtained using
the immobilized catalyst were smaller than with the nonimmobilized
catalyst, experiments were carried out to determine
the lipase stability of the immobilized cells within BSPs. A lipase
batch was used to catalyze two successive transesterification reactions,
each of which lasted 96 h. Fig. 2B shows that, after the first
use of the immobilized lipase, an appreciable diminution of reaction
velocity and transesterification conversion occurred (from
around 70% to 43% at 96 h in only two reaction cycles). In this
respect, Li et al. (2008) observed that a considerable amount of
reaction products (FAMEs and glycerol) accumulated inside the cell
during the repeated use of BSPs even using t-butanol as the reaction
medium, decreasing the enzymatic activity of the whole cell
immobilized within BSPs with the number of uses. Hama et al.
(2007) also observed that the FAME conversion decreased sharply
(from 90% to 10% in the tenth cycle) with the increasing numbers
of batch cycles in the methanolysis of soybean oil catalyzed by
immobilized R. oryzae within BSPs in a shaken bottle. Therefore,
the accumulation of polar lipids along with the accumulation of
glycerol and FAMEs on the BSP surface could be the reason for
the low reaction velocities, conversions and lipase stability
observed in this work using microalgal oil (Fig. 2).
Given that the conversion to FAMEs was higher using the nonimmobilized
whole-cell catalyst (81.8%) rather than the immobilized
one (70.3%) (Fig. 2A) and that the stability of the immobilized
whole-cell lipase was not as high as could be expected (Fig. 2B),
free R. oryzae cells were selected as the catalyst in order to obtain
higher conversions. This choice also reduces the process costs by
eliminating the immobilization of the R. oryzae cells.