Chitosan Derivatives - Chitosan Based Hydrogels: Functions and Applications - page 56

respectively, to prepare N-sulfated-chitosan. Compared to chlorosulfonic acid, the sulfur
trioxide-organic solvent complexes are mild and less destructive.
We aimed to generate compounds having lowest toxicity for determining the pharmaco-
logical structure-function relationships among different backbone structures and differ-
ently arranged functional groups compared to those of heparin and heparan sulfate. Solely
or in combination, N -sulfo, O -sulfo, N -acetyl, and N -carboxymethyl groups were intro-
duced into chitosan with the highest possible regioselectivity and completeness and
defined distribution along the polymer chain [132].
N -Sulfonation: N-Sulfonation of chitosan or 6- O -sulfochitosan in a homogeneous aque-
ous solution using the trimethylamine-SO 3 complex according to Holme and Perlin [118]
resulted in almost complete N -acetyl- N -sulfochitosan without sulfation at C-3 and without
a decrease in the original NAc content.
6-O -Sulfonation: Focher et al. [133] described a method for regioselective 6-O-sulfonation
of chitosan. These workers used a copper complex for simultaneous intermediate protec-
tion of the amino groups at C-2 and the OH group in the 3-position of the saccharide back-
bone. Application of this method to chitosan (DD 0.86) was highly regioselective and
complete without changes in NAc content and without reaction at C-3.
3,6 -di- O -sulfonation: Because chitosan (DD 0.86) or carboxymethylated chitosan (0.86) is
insoluble in DMF, the phthalimido group as the intermediate protecting group was intro-
duced to solubilize the polymer. 3,6-O-disulfonation was carried out with the SO 3 -pyridine
complex, which when followed by deprotecting the amino group with hydrazine hydrate
resulted in a highly sulfated chitosan derivative with a sulfate DS at C-3 of 0.97 and at C-6
of 0.79 [125].
3-O- Sulfonation: In heparin chemistry, there is a known sulfato transfer reaction for spe-
cific 3-O-sulfonation from glucosamine- N -sulfonate units, which is not applicable to chito-
san. Therefore, Yao et al. [125] first completely sulfated both the 3- and 6-position of the
glucosamine moiety, and then tried to use specific 6-O-desulfonation reactions from hepa-
rin chemistry to obtain 3-O-sulfonation chitosan [134].
Generally, the chemical derivatization reactions of chitin or chitosan were conducted
under heterogeneous or semiheterogeneous conditions due to chitin's poor solubility in
common organic solvents. Consequently, the attainment of a high degree of sulfation was
difficult, with poor selectivity for the site of sulfation (C-6, C-3, or N-2 positions), inevitably
resulting in multisubstituted derivatives unless tedious preprotection and deprotection
steps were taken [111,124]. The uncertainty regarding degree of sulfation at the individual
positions led to structure-activity relationship ambiguities. Finally, heterogeneous reac-
tions are known for their poor reproducibility, limiting industrial production and practical
applications. Zou and Khor [129] and Baumann and Faust [132] presents carefully prepared
and characterized sulfated-chitins whose anticoagulant reactivity can be specifically
related to the structure of the material. 6-O and 3,6-O-sulfated-chitins with degree of
sulfation ranging from 0.53 to 1.91 were prepared under mild and homogeneous condi-
tions, in a controllable manner, in a 5% LiCl/DMAc solvent system. Sulfation at room
temperature yielded only monosubstituted 6-O-sulfated-chitins, whereas elevated tem-
peratures gave 3,6-O-disulfated-chitins. Sulfation at the two positions resulted in different
effects on the structural features of sulfated-chitins, the C-3 position being more subject to
structural variation than the 6-O position. At low degree of sulfation, both the 6-O and
3,6-O-sulfated-chitins showed structural heterogeneity that eased as degree of sulfation
increased, becoming more homogeneous and uniform.
Chitosan sulfates have been shown to possess anticoagulant and hemagglutination inhi-
bition activities due to structural similarity to heparin [105, 106, 135-138]. By sulfation of