Fig. 4 shows the final spectra used to determine the relative
activity for urease in solution or immobilized in solid supports
under several conditions. The UV–vis spectra shows the relative
magnitude of the band centered at 588 nm after 30 min of reaction
due to the effect on the indicator bromocresol purple, which
changes in color as a consequence of the pH increase. To better
compare the exact evaluation of the relative activity, Table 1 is
shown. The relative activity is measured after a lag time (which
was negligible for all of the systems) and is determined through
the linear increase of the absorbance at 588 nm with time. There
is no enzyme activity for the LB film without urease, as expected.
Urease casted directly onto the solid support presents some activity,
and urease immobilized in a DODAB LB film presents activity
approximately two-fold higher than urease immobilized as a casted
film, which may result from a certain amount of urease molecules
immobilized on the solid support during contact with the aqueous
subphase. When EPS are present in the LB film, the relative activity
is five times higher than that when urease is immobilized in the
DODAB LB film without EPS. The relative activity in this case is even
higher than that of the enzyme in solution, as shown in Table 1. This
result is remarkable because it reveals a high affinity of the enzyme
for the EPS-DODAB monolayer, therefore serving as a good matrix
for the incorporation of urease and enhancing its activity. The relative
activities obtained after 1 week retained a similar comparative
trend among the systems, and less than 10% of the activity was
lost for the urease-EPS-DODAB LB film, in contrast to 65% for the
urease-DODAB LB film.