Enzyme Immobilization on Chitosan-Based Supports - Chitosan Based Hydrogels: Functions and Applications - page 358

in this manner has almost no activities. The functionalization of chitosan with DE hydro-
chloride allows good adsorption and activity retention. Spagna et al. reported that the
immobilization of Rha on supports chitin, chitosan and derivatized chitosan and DE-chitosan
contained in a commercial preparation is used in the winemaking industry. In particular
on DE-chitosan, the Rha was adsorbed and cross-linked with various bifunctional agents
(GA, diepoxyoctane, suberimidate, and EDC), whose best results (immobilization yields
and activity) were obtained with EDC that allowed a reduction in the involvement of the
enzyme amine groups that are probably important in the catalytic mechanism. In addition,
the use of rhamnose and a succinimide (NHS) during cross-linking enhanced the action of
the EDC and so increased the immobilization yield and activity. This biocatalyst allowed an
increase in the aroma in a model wine solution containing glycosidic precursors with a
marked reduction in specificity toward tertiary monoterpenols as compared with the free
enzyme. The results obtained seem to indicate that the amino group plays an essential role
in the catalysis of Rha from A. niger [68].
8.4.1.3 Amino Acid-Modified Chitosan
Amino acids, the monomeric units of proteins and bio-origin materials, are chiral molecules
with a relatively low molecular weight and have various properties due to their various
side chains (acidic, basic, hydrophobic, etc.). They have also been used as pseudobiospecific
affinity ligands for proteins. Therefore, chitosan beads with various properties could be
easily prepared by introducing amino acids into their polymer backbone.
Yi et al. consider that amino acids are suitable materials for the preparation of modi-
fied chitosan beads with increased lipase-compatibility. Besides, the preparation pro-
cess is simple and economical, and some amino acids are expected to improve the
catalytic performances of the immobilized lipases due to the increased compatibility
with the immobilized lipase. Therefore, it is considered that the development of amino
acid-modified chitosan beads might be valuable for industrial applications of lipase.
Various amino acids (Gly, Glu, Lys, Leu, Ala, Phe, Ser, Tyr)-modified chitosan beads
(CBs) for immobilization of lipases from C. rugosa were prepared by activation of a chi-
tosan backbone with ECO followed by amino acid coupling. The immobilized lipase on
unmodified chitosan beads showed the highest immobilization yield (92.7%), but its
activity was relatively low (10.4%). However, in spite of low immobilization yields
(15-50%), the immobilized lipases on the amino acid-modified chitosan beads showed
activities higher than that of unmodified chitosan beads, especially on Ala- or Leu-
modified chitosan beads with 49% activity for Ala-CB and 51% for Leu-CB. Moreover,
the immobilized lipases on amino acid-modified chitosan beads showed good thermal
stability, storage stability, and reusability and, therefore, are suitable for industrial
application [69].
Xiao et al. reported the synthesis and properties of a novel cross-linked chitosan resin
modified by l-lysine (LMCCR). It is known that l-lysine is a natural alkaline amino acid
and owns three reactive groups (a-carboxyl group, a-amino group, and e-amino group),
making the chemical modification possible. In order to synthesize the novel resin, an
acylation reaction between a-carboxyl group in l-lysine and primary amino group in
chitosan has to be conducted besides a cross-linking reaction between GA and chitosan.
At the same time, N , N -dicyclohexylcarbodiimide, an efficient peptide coupling reagent,
is used to induce the acylation reaction. Micrographs and SEM images show that LMCCR
is spherical in shape with macroporous structure. Due to its cross-linked structure and
side chains, LMCCR is amorphous but more rigid and stable, and no longer dissolved in