ed for use with the laboratory experiments employed in
analyzing bioadditive performance. In other cases, no
obvious or universal assay will be available and thus a
method must be designed to match the specific requirements of the specific test conditions. This is especially the
case where a dry, cast film must be tested for its enzymatic activity against the target molecule of interest (e.g.,
in this case, grease, cooking oils, natural fats, etc.) In
either case, it is very important to consider the sensitivity and availability of substrates.
For our study of lipolytic enzymes, a visual method was
selected based on previous literature reports of its effectiveness in quickly assessing the specificity of lipase enzymes.
Our colleagues at the University of Southern Mississippi
developed a rapid colorimetric method to detect degreasing
of a surface using pH indicators embedded in a gel matrix
containing the oil or grease of interest. When the indicator
material was applied to surfaces coated with leading floor
sealants (five mil thickness) containing DeGreez, a color
change was noted within 10 minutes, with the assay completed in 30 minutes (Figure 3). The indicator material is initially dark green and changes to yellow in the presence of
active DeGreez additive in the coating being tested. No color
change is observed in control surfaces that do not contain
any DeGreez. This method enables in situ visual evaluation
and real-time observation of the catalytic activity of the
biobased additive immobilized in the applied coating.
STEP 4: SCREEN FOR COMPATIBILITY
WITH POLYMER SYSTEMS
After confirming that catalytic activity of the selected bio-
compound candidates can be correctly assayed in the coat-
ing as a cast film, the next step in the development process
is to determine if the candidates are compatible with the
wide range of resin systems commonly used in commercial
paints and coatings. Several questions must be answered
at this stage: a) is the bioactive stable in the polymer
matrix? b) does the biocatalyst function properly when
blended with the polymer? and c) is the activity level affect-
ed by the presence of the resin system? An understanding
of the role that the physical and chemical properties of the
polymer and the interplay between actives and polymer
and the chemicals being reacted each control the efficiency
of the reactive coating. To optimize the activity of embed-
ded biocatalysts, selection criteria of the solid phase poly-
mer type are as important as those of the biocatalyst.
Although molecular interaction with polymeric materials
may alter some of the enzymatic characteristics through
activation or enhanced specificities, the fundamental prop-
erties must remain unchanged in a successful functional
coating. If chemical interactions take place between the
resin system and the biological molecule, the activity of the
bioadditive can be reduced or eliminated.
Table 2: Enzyme Classifications
Group
Reaction catalyzed
Typical reaction
Enzyme example(s)
with trivial name
EC 1
Oxidoreductases
To catalyze oxidation/reduction reactions;
transfer of H and O atoms or electrons from
one substance to another
AH+B A+BH
(reduced)
A + O AO (oxidized)
Dehydrogenase, oxidase
EC 2
Transferases
Transfer of a functional group from one sub-
stance to another. The group may be methyl-,
acyl-, amino- or phosphate group
AB+C A+BC
Transaminase, kinase
EC 3
Hydrolases
Formation of two products from a substrate by
hydrolysis
AB + H2O AOH + BH
Lipase, amylase, pepti-
dase
EC 4
Lyases
Non-hydrolytic addition or removal of groups
from substrates. C-C, C-N, C-O or C-S bonds
may be cleaved
RCOCOOH RCOH +
CO2 or [x-A-B-Y] [A=B
+ X-Y]
Decarboxylase
EC 5
Isomerases
Intramolecule rearrangement, i.e. isomerization
changes within a single molecule
AB BA
Isomerase, mutase
EC 6
Ligases
Join together two molecules by synthesis of
new C-O, C-S, C-N or C-C bonds with simul-
taneous breakdown of ATP
X + Y+ ATP XY + ADP +
Pi
Synthetase
