Advances in Chitin and Chitosan Modification through Graft...

Biopolymer chitin, the most abundant natural amino polysaccharide, and its mostimportant derivative, chitosan, are recently considered as the subjects for exten-sive worldwide academic and industrial research. In spite of potential applications

of chitin and chitosan, it is necessary to establish efficient appropriate modifications toexplore fully their high potential. A variety of chemical modifications are employed to mod-ify these carbohydrate polymers. The present article provides a comprehensive review onone of the most promising approaches to modify chitin and chitosan, i.e., graft copolymer-ization, with an emphasis on the synthetic aspects. Both chemically- and radiation-initiat-ed graft copolymerization of various vinyl monomers onto the trunk polymers are investi-gated. Meanwhile, the limited cases of polycondensation and oxidative coupling are pre-sented as the non-vinyl graft copolymerization methods. Then, the ring-opening graftcopolymerization is described and the cases of the cyclic monomers α-aminoacid N-car-boxy anhydrides and ε-caprolactone are investigated. An extensive description of the“grafting onto” approach is provided. The preformed polymers discussed here for graftingonto chitin/chitosan include living poly(2-alkyl oxazolines), poly(ethylene glycol)s, blockpolyethers, poly(ethylene imine)s, poly(2-hydroxyalkanoate)s, polyurethanes, poly(dimethylsiloxane)s, and dendrimer-like hyperbranched polymers. Chitin/chitosan multi-ple modification including graft copolymerization is also investigated. Regioselectivegrafting using derivatives such as 6-iodo-, mercapto-, deoxy(thiosulphato)-chitins, and N-trichloroacetyl chitosan are described as suitable approaches to achieve chitin/chitosangraft copolymers with well-known structures.

M. Jalal Zohuriaan-Mehr*

Superabsorbent Hydrogel Division, Iran Polymer and Petrochemical InstituteP.O. Box: 14965-115, Tehran, I.R. Iran

Received 22 April 2004; accepted 14 September 2004

Advances in Chitin and Chitosan Modificationthrough Graft Copolymerization:

A Comprehensive Review

chitin;chitosan;polysaccharide;graft copolymerization;modification.

A B S T R A C T

Key Words:

INTRODUCTIONGRAFT COPOLYMER SYNTHESISVinyl Graft Copolymerization

(a) Chemically Initiated Vinyl Polymerization(b) Radiation-initiated Vinyl Polymerization

Non-Vinyl Graft Copolymerization(a) Graft Copolymerization via Polycondensation

CONTENTS

Iranian Polymer Journal

14 (3), 2005, 235-265

(*) To whom correspondence should be addressed.E-mail: [email protected]

236

Advances in Chitin and Chitosan Modification ... Zohuriaan-Mehr M.J.

Iranian Polymer Journal / Volume 14 Number 3 (2005)

(b) Graft Copolymerization via Oxidative CouplingCyclic Monomer Graft Copolymerization: Ring-Opening Method Preformed Polymer Grafting: �Grafting Onto� MethodMultiple Modification of Chitin/Chitosan

(a) Further Modification of the Graft Copolymers(b) Grafting onto Pre-modified Chitin/Chitosan

CONCLUSIONACKNOWLEDGEMENTSREFERENCES

INTRODUCTION

Cellulose and chitin as biopolymers are the most abun-dant organic compounds in Nature and estimated to beat levels approaching 1011 tons annually [1]. Chitin hasbeen a major structural component of animal exoskele-ton since the Cambrian Period, more than 550 millionyears ago. The total amount of chitin harvestable with-out imbalancing the marine ecosystem is estimated tobe 1.5*108 kg/year [2], mostly from the shells of crus-taceans such as crab, shrimp and krill. Although cellu-

lose has been studied extensively, only limited attentionhas been paid to chitin, principally from its biologicalproperties [1,3]. Despite its huge annual production andeasy accessibility, chitin still remains an unutilized bio-mass resource primarily because of its intractable bulkstructure [1]. However, as Khor has stated [3], the 21stcentury can be the century of chitin taking a place as anextraordinary material, because chitin and its deriva-tives have exhibited high potential in a wide variety offields including medical, pharmaceutical, cosmetics,bio-related science and technology, food industry, agri-culture, and environmental protection [4-9]. RaviKumar has emphasized on the pharmaceutical applica-tions of these biopolymers in his recent reviews [9-11].Some reviews are published on fibre- and film-formingcapabilities of chitin and chitosan [12,13]. In a veryfascinating field i.e., gene therapy, chitosan and itsappropriate derivatives are recently found to be excel-lent candidates for controlled gene delivery [14-18].Overall, as exhibited in Figure 1, either scientific [3] orpatent [7] literatures reveal a considerable growing inthe field of chitin/chitosan science and technology frommid 1980s.

Chitin is structurally similar to cellulose, but it hasacetamide groups at the C-2 positions instead ofhydroxyl groups. So it is a nitrogen (amido/amino) con-taining polysaccharide, with repeating units of 2-acetamido/amino-2-deoxy-(1 4)-β-D-glucopyra-nose (Scheme I). In addition to its unique polysaccha-ride architecture, the presence of a little amino groups(5-15%) in chitin [4,19] is highly advantageous for pro-viding distinctive biological functions and for conduct-ing modification reactions [7,20,21]. Chitosan is the N-deacetylated derivative of chitin, though this N-deacetylation is almost never complete [20,21]. Actual-ly, the names �chitin� and �chitosan� correspond to afamily of polymers varying in the acetyl content.

Figure 1. Annual number of chitin/chitosan related reports;(a) patents, 1966-2000 [7], (b) scientific research articles inthe 1990s as obtained from ScienceDirect [3].

Scheme I. Structural repeating units of chitin (DD= 5-15%)and its deacetylated product, chitosan (DD 40%). ≥

Therefore, the degree of acetylation (DA) determineswhether the biopolymer is chitin or chitosan. �Chi-tosan� is the term used for the considerably deacetylat-ed chitin that is soluble in dilute acetic acid (degree ofdeacetylation, DD 70%) [1].

In spite of potential applications of chitin and chi-tosan, it is necessary to establish efficient appropriatemodifications to explore fully the high potential ofthese biomacromolecules. Chemical modifications ofchitin are generally difficult owing to the lack of solu-bility, and the reactions under heterogeneous conditionsare accompanied by various problems such as the poorextent of reaction, difficulty in selective substitution,structural ambiguity of the products, and partial degra-dation due to severe reaction conditions. Therefore,with regard to developing advanced functions, muchattention had been paid to modification of chitosanrather than chitin.

Chitin and chitosan have been modified via a vari-ety of chemical modifications. Some authors havereviewed the methods [1,4] and Roberts have explainedthe modification reactions in his source-book ChitinChemistry [20]. Of the various possible modifications(e.g., nitration, phosphorylation, sulphation, xanthation[14], acylation, hydroxyalkylation [4], Schiff’s baseformation and alkylation [4,20-22]), graft copolymer-ization is expected to be one of the most promisingapproaches to a wide variety of molecular designs lead-ing to novel types of hybrid materials, which are com-posed of bio- and synthetic-polymers [23]. This modi-fication technique, as foreseen by Kurita [1], will like-ly find new applications in some fields including watertreatment, metal cation adsorption, toileries, medicine,agriculture, food processing and separation. Thismethod has not been explored extensively, so that theRobertsí famous sourcebook comprises only less thanone page on this topic. A literature review showed that,except a very short article published in 1996 by Kuritain the Polymeric Materials Encyclopedia [24], there isno review article on the graft polymerization ontochitin and chitosan.

During recent years, various research-teams as wellas the author and coworkers [25-33] have focusedremarkably on graft copolymerization as a versatilemethod of chitin and chitosan modification. This fasci-nating technique may be considered as an approach toachieve novel chitin/chitosan-based materials with

improved properties including all the expected useful-ness of these biomaterials [4-11]. As concluded in thepresent article, chitin will not be a �biomaterial in wait-ing� any more.

GRAFT COPOLYMER SYNTHESISA graft copolymer is a macromolecular chain with oneor more species of block connected to the main chain asside chain(s) [34]. Thus, it can be described as havingthe general structure shown in Scheme II, where themain polymer backbone poly(A), commonly referredto as the trunk polymer, has branches of polymer chainpoly(B) emanating from different points along itslength. The common nomenclature used to describethis structure, where poly(A) is grafted with poly(B), ispoly(A)-graft-poly(B), which can be further abbreviat-ed as poly(A)-g-poly(B).

Grafting of synthetic polymer is a convenient

Advances in Chitin and Chitosan Modification ...Zohuriaan-Mehr M.J.

Iranian Polymer Journal / Volume 14 Number 3 (2005) 237

Scheme II. General free radical mechanism of graft copoly-merization of vinyl monomer B onto trunk polymer poly(A) toform the graft copolymer poly(A)-g-poly(B).

method to add new properties to a natural polymer withminimum loss of the initial properties of the substrate.Chitin and chitosan possess aforesaid useful propertiesthat render them interesting starting materials for thesynthesis of graft copolymers. Most of the copolymersare prepared through graft polymerization of vinylmonomers onto the biopolymer backbone [31]. But thisis not the only approach for synthesizing a graftedproduct. Because the chemistry of grafting vinylmonomers is quite different from that of grafting othermonomers (or performed side chains), this section isdivided into two subsections, the first dealing withgrafting of vinyl monomers, the second reviewing othertypes of grafting methods.

Vinyl Graft CopolymerizationGrafting of polyvinylic and polyacrylic synthetic mate-rials on the polysaccharides are mainly achieved byradical polymerization. Graft copolymers are preparedby first generating free radicals on the biopolymerbackbone and then allowing these radicals to serve asmacroinitiators for the vinyl (or acrylic) monomer(Scheme II). Mino and Kaizerman firstly reported thisapproach in 1958 for graft copolymer preparation usinga ceric ion redox initiating system [36]. Then, thechemistry and technology of the radical graft copoly-merization technique [23,37] was developed especiallyin the case of cellulose [38] and starch [34,39]. Gener-ally, free radical initiated graft copolymers have medi-um to high molecular weight branches that are infre-quently spaced along the polysaccharide backbone[38]. The copolymerizations can also be initiatedanionically by allowing monomer to react with an alka-li-metal alkoxide derivative of polysaccharide. Howev-er, this method has not been progressed due to difficul-ty of the process and the low molecular weight of thegrafted branches [39]. The properties of the resultinggraft copolymers may be controlled widely by the char-acteristics of the side chains including molecular struc-tures, length, number, and frequency.

One of the most important features of graft poly-merization is unwanted formation of homopolymer,homopoly(B), that is not chemically bonded to the sub-strate poly(A). Homopolymer can result if the initiatorused is one that produces free radicals in solution (inthe presence of vinyl monomer B initiating homopoly-merization) before creating the macroradicals. Once a

grafted chain has been initiated and begins to propa-gate, chain transfer from the growing grafted chain endcan occur with some species in the medium to yieldfree radicals that could initiate the growth ofhomopoly(B) chains [23].

To evaluate the efficiency of the graft copolymer-ization, the homopolymer is extracted with an appropri-ate solvent. Then, the homopolymer percentage (Hp%)and other various grafting compositional parametersare calculated. Although there are no unified definitionsfor calculating the parameters, the most frequentlyreported expressions for all kinds of graft copolymer-izations are as follows [23,34,40]:

Graft yield (G%) = 100 (W3-W0) / W0 (1)

Add-on (Ad%) = 100 (W3-W0) / W3 (2)

Hp% = 100 (W2-W3) / W2 or,Hp% = 100 W4/ W1 (3)

where W0, W1, W2, W3 and W4 designate the weight ofthe original substrate, monomer charged, total product(i.e., copolymer and homopolymer), pure graft copoly-mer, and homopolymer, respectively.

The various initiating systems employed to graftcopolymerize different vinyl monomers onto chitin orchitosan can be categorized to two main classes, i.e.chemical initiation and radiation initiation that areinvestigated in the next sections.

(a) Chemically-initiated Vinyl PolymerizationAmong the variety of chemical reagents reported forinitiating the vinyl monomer graft copolymerizationonto chitin/chitosan, ceric ion initiation and Fenton’sinitiation are the most important systems.

Cerium in its tetravalent state is a versatile oxidiz-ing agent used most frequently in the graft copolymer-ization of vinyl monomers onto cellulose and starch[34-40]. It forms a redox pair with the anhydroglucoseunits of the polysaccharide to yield the macroradicalsunder slightly acidic conditions. As with cellulose andstarch, the ceric ion has been a useful initiation methodfor graft copolymerizing chitin and chitosan with typi-cal vinyl monomers [27-32] due to the similarities inthe chemical structures of these polysaccharides.

The mechanism of initiation for chitosan is

238



believed to begin with a complex formation of Ce4+

with the primary amine and the hydroxyl groups at theC-2 and C-3 positions, respectively. The radicalsresponsible for the initiation of grafted copolymerchains using vinyl monomer are produced from thecomplex dissociation. A general mechanism for thereaction proposed recently by the author and coworkers[28] is shown in Scheme III. At higher temperatures(e.g., 90oC), it is proposed that the imine moiety shownin Scheme III is further hydrolyzed in the aqueousacidic conditions to the corresponding aldehyde,whereby oxidation to an acyl radical gives another site

capable of initiating a grafted polymer chain [23].Moderate to excellent yields accompanied various

amounts of homopolymer are reported. Table 1 pro-vides the grafting conditions and compositional param-eters of the synthetic polymer grafted chitin/chitosanprepared using the ceric-saccharide initiating system.According to the recent study by the author andcoworkers [27,28], acrylonitrile was optimally graftcopolymerized onto chitosan in a homogeneous phasewhile a very low level of homopolyacrylonitrile (2 %)was formed (the first row of Table 1). Based upon thestudy, the optimum conditions for achieving the maxi-mum grafting were determined to be as: chitosanamount 0.20 g, acetic acid 2% w/w, reaction tempera-ture 50oC; AN 1.60 g, ceric ammonium nitrate (CAN)concentration 0.006 M, time 2 h. The grafting efficien-cy was recognized to remain almost unchanged withthe reaction time. The grafting-time independence canbe attributed to a decrease in concentration for both ini-tiator and monomer and also to a reduction of the num-ber of sites on the chitosan backbone accessible forgrafting as the reaction proceeds. Empirical rate of thepolymerization showed a first-order dependence on themonomer concentration and a half-order dependenceon the initiator concentration. An overall activationenergy of 44.9 kJ/mol was determined for this graftpolymerization reaction [28]. The hydrophobicallymodified chitosan exhibited higher thermal stabilitythan chitosan itself [30].

Kim et al. [41] reported the ceric-induced graftcopolymerization of N-isopropylacrylamide (NIPAM)onto chitosan at 25oC to prevent a high level ofhomopolymer formation (Table 1). The maximumgrafting yield (48%) was obtained at 0.5 M ofmonomer concentration, 0.002 M of CAN initiator and2 h of the reaction time. They found a decreased per-cent of grafting when the initiator concentration washigher than 0.002 M. The grafting loss was attributed tolower macroradical formation at the expense of produc-ing more homopoly(NIPAM). The excessive CAN mayalso be consumed for oxidation of the polysaccharidebackbone leading to decreased molecular weight of thegraft copolymer product.

Vinyl acetate (VAc), a less reactive monomer thanacrylates, was also recently graft copolymerized ontochitosan by CAN in dispersion medium at 60oC[42,43]. With an addition of 0.5-7.5 g of chitosan based



Scheme III. General mechanism for ceric-initiated graftcopolymerization of a typical vinyl monomer, acrylonitrile(AN), onto chitosan [28]. The opening of the pyranose ringshown above is very rarely occurred along a chitosan chain,so the initial structure of the trunk polymer is not actuallydestroyed.

240



on 50 g VAc, the monomer conversion was found to bebetween 70 and 80% after 2 h of reaction. The experi-mental results indicated that the chitosan macromole-cules not only took part in the copolymerization, butalso served as a surfactant providing the stability of thedispersion particles. The particulate membrane chi-tosan-g-PVAc formed after drying exhibited highertoughness and lower water-absorption compared tonon-grafted chitosan. The copper ion adsorption by thepoly(VAc) grafted chitosan was also studied [43].

The monomer 4-vinylpyridine (4VP) is anothernon-acrylic monomer graft polymerized onto chitosan[44] under homogeneous conditions. Percent graftingwas increased with the amount of the monomer, show-ing a tendency to level off at a 4VP concentration of0.53 M. As given in Table 1, maximum graft yield(331%) was achieved in lieu of a very high CAN con-sumption (0.087 M). According to the authors, theirused system has a drawback since it requires the pre-cipitation of the obtained product in a basic medium,resulting in the coprecipitation of the excess ceriumsalts that cannot be separated from the grafted product.The purified graft copolymers showed swellabilitybehaviour in 1:1 and 1:2 acetic acid:ethanol solventmixtures and insolubility in ethanol, dimethylfor-

mamide (DMF), dimethylsulphoxide (DMSO) andtetrahydrofurane (THF).

Acrylic and methacrylic acids were graft polymer-ized onto chitosan by Shantha et al. [45] to tailor drugcarriers. They have reported an ambiguous procedureof polymerization without giving the CAN concentra-tion used. Although they did not achieve a high graft-ing percent (Table 1), they utilized graft copolymers forpreparing functionalized chitosan beads by a polymerdispersion technique. The drug sulphadiazine wasentrapped in the microspheres and the in vitro drugrelease profiles were established in either simulatedgastric and intestinal fluids. A sulphobetain methacrylicmonomer, N,Ní-dimethyl-N-methacryloxyethyl-N-(3-sulphopropyl) ammonium, was recently reported to begraft polymerized onto chitosan by ceric ion initiation[46]. A maximum percentage of grafting about 50%was obtained under an optimized condition (0.5 g chi-tosan in 50 mL of 2 wt% acetic acid aqueous solution,CAN 0.0182 M, monomer 0.1434 M, 60oC, 2 h). Ther-mal properties of the graft copolymer were found to beslightly different from the original chitosan.

Chemically modified chitosan microspheres weresynthesized by graft copolymerization of a bifunction-al macromolecular monomer, poly(ethylene glycole)

Table 1. Preparative reaction conditions and grafting parameters of synthetic polymer grafted chitin/chitosan synthesized via ceric-

induced graft copolymerization.

(a) Acrylonitrile, (b) Not quantified, (c) N-Isopropylacrylamide, (d) Vinyl acetate, (e) 4-Vinyl pyridine, (f) Acrylic acid, (g) Not given, (h) Methacrylic acid, (i) N,N-Dimethyl-N-

methacryloxyethyl-N-(3-sulphopropyl) ammonium, (j) Acrylamide, (k) Methyl methacrylate.

Substrate MonomerCopolymerization conditions

Grafting parameters

(%) Ref.

Ce4+ (mM) Monomer concentration Temp. (oC) Time (h) G Ad HpChitosan

Chitosan

Chitosan

Chitosan

Chitosan

Chitosan

Chitosan

Chitin

Chitin

Chitin

Chitin

ANa

NIPAMc

VAcd

4VPe

AAf

MAAh

DMSAi

AA

AA

AMj

MMAk

6

2

2.16

87

NGg

NG

18.2

8.0

3.45

8.8

4.25

6.6 wt%

0.5 M

9.6 wt%

1.22 M

NG

NG

0.1434 M

1.17 M

0.959 M

0.42 M

0.0199 M

50

25

60

70

50-55

50-55

60

60

60

60

40

2

2

2

1

3

3

2

2

3

2

5

NQb

48

236

331

57

NQ

50

200

45

243

~300

81

NQ

NQ

76.8

NQ

NQ

NQ

NQ

31.3

70.8

NQ

2

7

NQ

NQ

10

10

NQ

NQ

NQ

NQ

NQ

27,28

41

42,43

44

45

45

46

47

48

47

49



diacrylate onto chitosan backbone using CAN [47].The products were fully characterized by spectral, ther-mal and morphological techniques. Since the prepara-tive reaction resulted in cross-linked networks, thegrafting parameters could not be determined.

Regarding chitin, limited reports have been pub-lished about the ceric initiated grafing. Kurita et al. [48]have reported an efficient and reproducible procedurefor graft copolymerization of acrylamide (AM) andacrylic acid (AA) onto powdery chitin. As a solvent,water proved to be superior to conventional nitric acidsolution (except the reactions with a small amount ofthe initiator CAN). In spite of the heterogeneous condi-tions, under optimized conditions (0.10 g chitin, 10 mLwater, 0.8-1.6 g CAN, 0.30-0.43 g monomer, 60oC, 2h),around 240% and 200% graft yields were achieved forAM and AA, respectively. The resulting graft copoly-mers showed improved affinity for solvents and hygro-scopicity compared to the original chitin.

Acrylic acid was graft copolymerized onto chitinusing CAN by other workers as well [49]. Theyachieved a low grafting (45%) under conventionallyacidified (1 M nitric acid) reaction conditions (chitin1.00 g, water 45 mL, CAN 0.00345 M, AA 0.959 M,60oC, 1 h). The chitin-g-poly(AA) was used to studythe effect of carboxylic group of the graft copolymer onthe metal binding ability of calcium ions in aqueousmedium as a function of pH, contact time and the metalconcentration. The maximum adsorption of the graftcopolymer and the original chitin were found to be 0.50and 0.19 mmol Ca2+ g-1, respectively.

Ren et al. [50] modified chitin via graft polymeriza-tion of methyl methacrylate (MMA) by CAN. Theyfound the optimized grafting conditions to be: chitin0.50 g, water 220 mL, nitric acid 0.0383 M, CAN0.0425 M, monomer 0.0199 M, 40oC, 5 h. Grafting per-centage of about 300% was obtained under the condi-

tions. Solubility of the highly poly(MMA) graftedchitins in various organic solvents showed significantchanges, and a gel-like mass swelling was resulted.Fine films could be made from the gel-like materialespecially in DMF or dimethylacetamide, when a smallamount of lithium chloride was used. Complementarystudies revealed that the amphiphilic comb-like chitinderivatives containing PMMA side chains were able toform stable monolayers with high collapse pressure[51].

Fenton�s reagent is another frequently used initiatorfor graft copolymerizing vinyl monomers ontochitin/chitosan [52,53]. The reagent involves a redoxreaction between the ferrous ion and hydrogen perox-ide, providing hydroxyl radicals (Scheme IV). Theseradicals are believed to be responsible for creating themacroradicals on the polysaccharide backbone, bymeans of hydrogen abstraction, that initiate the growthof grafted chains with various monomers. AlthoughH2O2 alone could be an adequate initiator for thecopolymerization, there are reasons why reducingagents such as Fe2+ are used for grafting onto chitosan.In addition to the higher yield of radical production atmuch lower temperatures via the redox reaction, thechelating properties of chitosan with metal ions tend topromote OH radical formation in the vicinity of the chi-tosan in order to increase macroradical yields ratherthan homopolymer formation initiation.

MMA was graft copolymerizaed onto chitosan withgrafting pecentages of 400-500% with homopolymeryields of around 20-30% [52]. Methyl acrylate (MA)has also been grafted with yields of 250-300% whilehomopoly(MA) was produced in the range of 15-20%[53].

Scheme IV. Mechanism for hydroxy radical formation bymeans of Fentons’s reagent in aqueous media.

Scheme V. Mechanism for macroradical formation on thebackbone of chitin by means of ferrous ion-persulphate redoxsystem in aqueous media.

Since chitin in less reactive than chitosan, itschelating properties may be enhanced through the addi-tion of thiocarbonate sites along the chitin backbonevia the xanthate process, i.e. treatment with concentrat-ed aqueous sodium hydroxide and carbon disulphide[54]. After treating the chitin thiocarbonate derivativewith ferrous ion, the complex (Chitin-O-CS2

-)2Fe2+ isformed. The decomposition of the complex leads tofree Fe2+, which is believed to react with hydrogen per-oxide as shown in Scheme IV to produce OH radicalsthat subsequently create chitin macroradicals uponhydrogen abstraction. Grafting percentages on theorder of 80 and 40% were obtained with this processusing acrylonitrile and acrylic acid, respectively. Inboth cases, homopolymer was obtained but not quanti-fied.

As an alternative method for grafting, a variation ofFenton’s reagent has been investigated. Thus, potassi-um persulphate (KPS) and ferrous ammonium sulphateare combined in a redox reaction that ultimately pro-duces hydroxy radicals being able to form chitinmacroradicals (Scheme V). MMA was graft polymer-ized onto chitin with this system at grafting percentagesof 300-500%, where 40-50% of the monomer washomopolymerized [55]. Potassium persulphate alonewas only able to achieve around 80-90% grafting whileproducing similar yields of homopoly(MMA).

When a non-acrylic monomer, i.e. N-vinyl pyrroli-done, was employed to graft copolymerize onto chi-tosan using KPS alone, graft yields of 200-300% wereobtained accompanied by 10-20% homopolymer yields[56]. After the grafting, the solubility of chitosan wasmarkedly reduced either in common organic solventsor in dilute organic or inorganic acids. However, thesolubility of the grafted chitosan substantiallyimproved after adsorption of copper ions, becomingcompletely soluble in dilute hydrochloric acid.

Chitosan has been subjected to graft copolymeriza-tion, comparatively with MA and MMA monomersusing KPS alone and KPS coupled with various reduc-ing agents [57]. The results are summarized in Table 2.No reference was made to a mechanism where the per-sulphate reacts specifically with the chitosan, so it isassumed the general mechanism proceeds as in SchemeV, where the other reducing agents (MnCl2, ammoniumtartrate, etc.) may be substituted for the ferrous ion.

Hsu et al. [58] found that chitosan was degraded by

KPS in aqueous media via a free-radical mechanism.This is an important point that should be taken intoaccount in all the persulphate containing initiating sys-tems. The same authors reported recently the synthesisof chitosan-modified PMMA by emulsion polymeriza-

242



Monomer Initiator system G (%) Hp (%) Ref.

MMA

MMA

MMA

MMA

MMA

MMA

MA

MA

MA

MA

MA

MA

VP

MMA

AN

AM

AA

AMPS

MA

VAc

MA

MMA

AN

MMA

MMA

MMA

KPS

KPS-FASa

KPS-CuCl2

KPS-MnCl2

KPS-AOXb

KPS-ATAc

KPS

KPS-FAS

KPS-CuCl2

KPS-MnCl2

KPS-AOX

KPS-ATA

KPS

KPS

KPS

KPS

KPS-FAS

KPS

DPICd

AIBN

AIBN

AIBN

AIBN

APS

H2O2

TBBe

268

300-350

497

489

397

388

281

80-90

48

335

511

339

200-300

276

249

220

52

180

650

10

20

65

10

300

300

40

37

40-50

29

16

29

31

63

40-50

87

22

4

40

10-20

-

-

-

-

-

-

-

-

-

-

50

50

50

57

55

57

57

57

57

57

55

57

57

57

57

56

60

60

61

62

63

64

65

65

65

65

23

23

66

Table 2. Various initiating systems employed for graft copoly-

merization of different vinyl monomers onto chitosan. This

table does not include the ceric- and Fenton's-based initiating

systems.

(a) Ferrous ammonium sulphate, (b) Ammonium oxalate, (c) Ammonium tartrate,

(d) Potassium diperiodatocuprate (III), (e) Tributyl borane.



tion [59]. They verified that chitosan played multipleroles in the emulsion. In addition to a surfactant role,KPS can appreciably be deactivated by low molecularweight chitosans produced by the persulphate-induceddegradation. Therefore, long reaction times will notfavour the formation of high copolymers when they areinitiated by a persulphate system.

Most recently, KPS-initiated graft copolymeriza-tion of acrylonitrile (AN) and MMA onto chitosan wasreported [60]. Maximum graft yield of chitosan-g-PAN(249%) was obtained with 0.12 M of AN and 0.00074M of KPS at 65oC for 2 h for 1% chitosan solution. Forchitosan-g-PMMA, 0.14 M of MMA at 65oC gavemaximum grafting (276%). No residual monomerswere found by HPLC in the graft copolymers. Thinmembranous films could be prepared by thermopress-ing the modified chitosans.

Yazdani-Pedram et al. have recently studied theeffect of reaction variables on KPS-initiated graftcopolymerization of acrylamide onto chitosan in thepresence of N,Ní-methylenebisacrylamide (MBA)cross-linker [61]. They found the optimum reactionconditions (0.8 g chitosan, 50 mL acetic acid 2%, 2.4 gAM, 0.25% MBA, 0.002 M potassium persulphate,60oC, 30 min.) to achieve high grafting percentage(~220%). Acrylic acid has also been graft copolymer-ized onto chitosan by the same research group [62].They employed KPS-ferrous ammonium sulphate(FAS) redox initiating system to achieve low efficiencyof grafting (52%) under optimized conditions (chitosan0.3 g, water 50 mL, acrylic acid 2 mL, KPS 0.01 M,FAS 0.00006 M, 70oC, 2 h). All the grafted productswere insoluble in water and in dilute acid solutions.The insolubility is the main difference between thegraft copolymers and the chitosan itself which readilydissolves in slightly acidic media. The grafted chitosansamples, however, were swelled in these media. Theswelling properties of the highly swollen poly(acrylicacid) grafted chitosan hydrogels were also studied [62].

Optimized synthesis of a graft copolymer based onchitosan and 2-acrylamido-2-methylpropane sulphonicacid (AMPS) was reported by Najjar et al. [63] usingKPS in homogeneous solution. The maximum percent-age of grafting of ~180% was achieved under the opti-mum conditions (1% v/v acetic acid, 50oC reactiontemperature, 10 min. chitosan-KPS mixing period, 0.37mmol of KPS, and 28.96 mmol AMPS).

Most recently, a novel redox system, Cu(III)-chi-tosan, was employed to initiate the graft copolymeriza-tion of MA onto chitosan in alkali aqueous media [64].A maximum grafting of around 650% was achievedunder the optimum conditions concluded: 0.3 g chitosan(DD 0.82, mesh 60), 2.6 mL MA, pH 11.6 (by addingKOH solution), Cu(III) 0.00192 M, 35oC, 1 h. Accord-ing to a proposal of the authors, a single electron trans-fer from -NH2 of chitosan to Cu(III) occurs to producea radical-cation converted subsequently to -NH radicalin alkaline medium. The -NH radical is thought to initi-ate the graft copolymerization (Scheme VI(a)). Thetrivalent copper, as the salt potassium diperioda-tocuprate (III), was easily prepared from the cheap cop-per salt, CuSO4.5H2O. Since the grafting reaction canbe carried out at a mild temperature of 35oC using acheap initiator, the initiating system was recognized tobe superior to conventional initiators [64].

A redox system based on a supernormal valencetransition metal, i.e. Ni(IV)-chitosan, was recentlyreported in the Chemical Journal on Internet [65].Thus, acrylonitrile was graft polymerized onto chitosan(MW 2-3*105, DD>0.82) using potassium diperioda-tonickelate (IV). Under the optimized copolymeriza-

Scheme VI. Single electron transfer mechanism for initiatinggraft copolymerization of vinyl monomers onto chitosan using(a) Cu(III) and (b) Ni(IV) proposed recently by Liu et al.[64,65].

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tion conditions (Ni(IV) 2.25 mM, monomer/chitosanwt. ratio 5.3, 32oC, 2 h) for treating 0.3 g of chitosan, agrafting percentage of 110% was achieved. Based onthe FTIR spectral N-H zigzag bands and the previousreports, the authors verified the process involving atwo-step single electron transfer mechanism for the ini-tiation of the polymerization (Scheme VI(b)).

Azobisisobutyronitrile (AIBN), ammonium persul-phate (APS), and hydrogen peroxide (H2O2) are com-monly employed to graft copolymerize vinylmonomers onto chitin and chitosan [23,66]. Underheating (or irradiation), the first radicals produced bythese systems occur from homolytic bond scissions ofthe initiator, whereby these radicals subsequently reactwith the monomer to initiate the polymerization. Fortypical graft copolymerization, these radicals providedby the initiator, in addition to reacting directly withvinyl monomer, abstract hydrogens from chitin or chi-tosan creating macroradicals that are capable of initiat-ing a grafted chain with vinyl monomers. Vinyl acetate,AN, MA, and MMA are graft polymerized onto chi-tosan using this kind of initiation system. The typicalresults are summarized in Table 2.

Tributylborane (TBB) was also utilized for initiat-ing the grafting onto chitosan. According to Kojima etal. [67], the alkylborane-initiated polymerizations ofvarious vinyl monomers in the presence of oxygenoccur by means of a free-radical mechanism, prompt-ing the investigation of the initiator for the grafting ofchitin with MMA. Based on the proposed mechanism,a solvated chitin-TBB complex produces radicals onthe chitin backbone that in turn, initiate the graft poly-merization. The system produces homopoly(MMA)value as high as 50% and low level of graft yield (about40%).

(b) Radiation-initiated Vinyl PolymerizationBoth high- and low-energy radiation may be used forgraft copolymerization of vinyl monomers onto poly-saccharides. The radiation-induced grafting onto cellu-losics has been discussed in the chapter 3 of the source-book of Hebeish and Guthrie [38].

Employing high-energy radiation (e.g., β, γ, X-ray)is an efficient basic method for initiating radical graftpolymerization onto polysaccharides. The initiationmethod with the highest graft efficiency seems to bepre-irradiated of the polysaccharides with γ-radiation

followed by the �activated� polysaccharide with vinylmonomers under suitable reaction conditions [35,38].Grafting efficiency means in this connection not only ahigh number of grafted branches with high molecularweights, it means in particular a low level ofhomopolymer formed. Although the radiation-basedgrafting is �cleaner� and more efficient in this regardthan chemical initiation methods, they are harder tohandle under technical conditions [35]. This is why thenumber of researches on �irradiation� methods havebeen considerably smaller that that of the �chemical�methods.

Irradiation of γ-ray on powdery chitin initiates thegraft polymerization of styrene, as in the case of cellu-lose, but the grafting percentage is low (i.e., 64%).Styrene was also graft copolymerized onto chitosanpowders or films. Water was recognized to be essentialfor the both grafting reactions. The chitosan-g-poly-styrene adsorbed bromine better than chitosan, and thecopolymer films showed less swelling and higher elon-gation in water than what the chitosan films did [1,68].

Pengfei et al. [69] recently reported the γ-radiation-induced graft copolymerization of styrene onto chitinand chitosan powder. The reaction was promoted in thepresence of methanol, and atmospheric oxygen delayedthe reaction but did not inhibit it completely. Molecularweight of the grafted polystyrene did not change obvi-ously with the dose of radiation, but it was increasedwith the increase of the concentration of the monomercharged. The polydispersity index of the grafted chainswas measured to be basically between 1 and 2.

Singh and Ray graft copolymerized 2-hydrox-yethylmethacrylate (HEMA) onto chitosan films using60Co gamma radiation to improve their blood compati-bility [70]. They found that the level of grafting couldbe controlled by the grafting conditions, namely sol-vent composition, monomer concentration, dose rate,and total dose. They achieved a maximum graft yield of108% under the conditions of solvent water-methanolvolume ratio 1:1, HEMA concentration 20 vol%, doserate 90 rad/s and total dose 0.216 Mrad. The swellingof this PHEMA-grafted film in phosphate buffer (pH7.4, 0.1 M) at 37oC was 58% compared to that of theoriginal chitosan film, i.e. 110%.

Chitosan films have also been subjected to gammaradiation-induced graft copolymerization of the vinylmonomer N,Ní-dimethylaminoethyl methacrylate [71].



The reaction variables affecting on the grafting percent-age have been studied. The grafting of 70% is beingachieved under the conditions of solvent water-methanol volume ratio 1:1, the monomer concentration15 vol%, dose rate 90 rad/s and total dose 0.216 Mrad,and irradiation time 80 min. The garfted chitosan wasfully characterized by swelling and tensile measure-ments, and FTIR, DSC, TGA and XRD methods. Thedegree of swelling, crystallinity, and tensile strengthwere decreased by 51, 43, and 37%, respectively, at agraft level 54%, whereas the modified films showedimproved thermal stability [71].

Low energy photons may also initiate the polymer-ization of a vinyl or acrylic monomer if the irradiationis carried out in the presence of an activator (photosen-sitizer). Such a sensitizer must become active on expo-sure to the particular wavelength range of the incidentradiation [38]. Irradiation with low energy radiation,i.e. visible or ultraviolet (UV) lights, usually in thepresence of a photosensitizer such as benzophenone orazo compounds, is a rarely used method for graftingonto chitin/chitosan. UV-initiated graft copolymeriza-tion of MMA onto chitosan has been reported [23,72].The grafting percentage was decreased in order:

Photo-initiation (by the light of 253 nm) without acatalyst (G 300%, Hp 30-40%) > photoinduced methodwith photosensitizer AIBN (G 150%, Hp 60%) > pho-toinduced method with photosensitizer benzophenon(G 140%, Hp 40%).

Under the noncatalytic photoinduced initiation con-ditions (i.e., the irradiation of only chitosan, solventand monomer), it was proposed that the amine group ofchitosan be removed by the photolysis [23]. When amixture of acrylonitrile and styrene was used, almostalternation copolymers were introduced [24].

Photo-induced initiation was also applied to thegraft copolymerization of MMA onto chitin or oxy-chitin (oxidized chitin). A small amount of dimethylfor-mamide reduced the induction period and increasedeither the grafting percentage or the apparent numberof grafted chains. Non-catalytic photo-induced graftpolymerization was smooth and obtained higher graftyield (G 150%, Hp 50%), compared to the graftingphoto-sensitized with H2O2 (G 70%, Hp 50%) or AIBN(G 60%, Hp 90%) [23,72].

Some pre-designated chitin/chitosan derivativeshave also been subjected to irradiation-induced graft

copolymerization to yield indirectly the correspondinggrafted products. They are explained in the next sectionentitled �grafting onto pre-modified chitin/chitosan�.

Non-vinyl Graft Copolymerization(a) Graft Copolymerization via PolycondensationCondensation polymerization has not been widely usedfor preparing graft copolymers of polysaccharides usu-ally due to susceptibility of the saccharide backbone tohigh temperature and harsh conditions of the typicalpolycondensation reactions.

However, lactic acid (LA) was successfully graftcopolymerized onto chitosan through condensationpolymerization of D,L-lactic acid in absence of a cata-lyst [73]. So, the bio-active and compatible polymerpolylactic acid (PLA) is conjugated with the biopoly-mer chitosan through an amide linkage.

The degrees of substitution (DS) ranged from 6 to18 were measured using both elemental analysis andsalicylaldehyde methods. Degree of polymerization(DP) of the PLA side chains was determined, by 1HNMR analysis, to be in the range of 0.9-4.4. The graftcopolymers were found to be physically cross-linkedduring the polymerization leading to pH-sensitive chi-tosan-PLA hydrogels. Water uptake of the hydrogelswas investigated as a function of values of DP, DS, pHand salt concentration. The structural change andswelling mechanism of the hydrogels were also inves-tigated [74]. A study on cytocompatibe chitosan-g-PLAcopolymer film has recently been reported [75]. Theresults showed that the cell growth rate on the film wasobviously faster than chitosan itself.

(b) Graft Copolymerization via Oxidative CouplingWith a view to prepare conductive polymers, polyani-line was grafted onto chitosan [24]. On the treatment ofchitosan in aqueous acetic acid solution with aniline inthe presence of APS, polyaniline side chains wereintroduced at the amino groups. Chitosan-g-polyanilinewas fabricated into films and fibres, but the propertiesvaried according to the ratio of amino group to anilinein the grafting reaction. With a ratio of 1/1 to 1/5, theproducts were sturdy and flexible, while those with a1/6 to 1/10 ratio were brittle. Optical microscopicobservations indicated that the products prepared at aratio below 1/5 were homogeneous, but those of 1/6ratio had crystalline regions. The graft copolymers

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were deep blue and became dark green when treatedwith hydrochloric acid. The conductivity could beraised from less than 10-7 to 0.01 S/cm upon protonicdoping with HCl [1,24].

Cyclic Monomer Graft Copolymerization: Ring-opening MethodIn general, four groups of cyclic monomers have beenmainly used for graft copolymerization onto polysac-charides: oxiranes (epoxides), lactons, α-aminoacid N-carboxy anhydrides (NCAs) and 2-alkyl oxazolines[35]. However, the various NCAs have been the maintype of cyclic monomer used to graft onto chitin andchitosan.

Chitin-g-oligo(caprolactone) was recently reportedto be synthesized via ring opening graft polymerizationof ε-caprolactone (CL) to partially deacetylated chitincatalyzed by tin(II) 2-ethylhexanoate in the presence ofwater as a swelling agent [76]. Thus, N-substitutedgraft copolymer with 40 wt% oligo(CL) (averagedegree of polymerization ~4) was obtained by the reac-tion conditions: 0.20 g chitin (DD 0.51), catalyst 0.17mol%, water 130 mol%, 100oC, 20 h. The structure of

the copolymer was fully characterized by IR, NMR andXRD methods.

Partially deacetylated chitin have been grafted withD,L-alanine NCA, γ-methyl L-glutamate NCA, and L-alanine NCA [23,24,77]. NCA ring can undergo nucle-ophilic attack to open and polymerize with evolution ofCO2 to yield a polypeptide chain [78]. As shown inScheme VII, the free amine of the deacetylated chitin isbelieved to initiate the graft copolymerization bymeans of attack upon carbonyl, ultimately creating thegrafted chitin derivative. The advantages of thismethod are low level of homopolymer formation andthe possibility of the side chain length control by theregulation of the NCA concentration under proper con-ditions. Degree of polymerization (DP), however, is notusually higher than 20 [23]. The resulting copolymersare new types of hybrid materials composed of both apolysaccharide and polypeptides.

Chitosan or water-soluble chitin failed to initiatethese reactions in organic solvents due to low extents ofswelling and rapid hydrolysis of NCAs [1]. A water-soluble chitin (50% deacetylated chitin), however,exhibited high reactivity in aqueous solution. It wastreated with an ethyl acetate solution of γ-methyl L-glu-tamate NCA [24]. The ring-opening graft polymeriza-tion of the NCA proceeded smoothly at 0oC to givechitin-g-poly(γ-methyl L-glutamate). Although NCAsare very susceptible to hydrolysis, the grafting efficien-cy was surprisingly high, up to 91%, and no homopoly-merization was observed. On alkaline hydrolysis, sidechain ester groups were transformed into carboxylategroups. Graft copolymerization of NCAs onto partiallydeacetylated chitins is also possible in DMSO, but thegrafting efficiency was moderate because of the hetero-geneous reaction conditions [1]. Chitin-g-poly(γ-methyl L-glutamate) copolymers have shown varyingdegrees of solubility in common polar solvents depend-ing on the side chain length [1,24].

Poly(L-alanine) side chains were also introduced tochitin in a similar manner [77]. The grafted poly(L-ala-nine) chains could be regarded as spacer arms having aterminal free amino group which can be used for fur-ther modifications. They were utilized to immobilizedihydronicotinamide groups (the active site of coen-zyme NDAH) (Scheme VIII). The resulting bioconju-gates were utilized for asymmetric reduction of aketone while the peptide spacer arms regulated the

Scheme VII. Synthesis of chitin/chitosan-polypeptide biocon-jugates via ring-opening graft copolymerization of α-aminoacid N-carboxy anhydrides (NCAs) onto partiallydeacetylated chitin [24,78].



reduction process [78].

Preformed Polymer Grafting: �Grafting Onto�MethodGrafting with telechelic polymers provides an alterna-tive method, commonly referred to as �grafting onto�,for synthesizing hybrid branched architectures [79].Telechelic polymers have been defined as those �con-taining one or more functional end groups that have thecapacity for selective reaction to form bonds withanother molecule� [80]. Unlike the classic graftingtechniques where the grafted chain is grown from thetrunk polymer by the continual addition of monomer to

Scheme VIII. Immobilization of dehydronicotinamide group(coenzyme NADH active site) on partially deacetylated chitin[77,78].

Scheme IX. General representation of the “garfting onto”method for modification of trunk polymer poly(A) with pre-formed reactive (telechelic) polymer, poly(B) [79]. Reaction offunctional group x with y leads to a covalent linkage.

Scheme X. Synthesis of living poly(2-alkyl-2-oxazoline),PAO, followed by its use to graft onto partially deacetylatedchitin [81].

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the growing chain end, �grafting onto� connects a pre-formed polymer chains (poly(B)) and the trunk poly-mer (poly(A)) by covalently bonding the chain end ofthe poly(B) with a particular site on poly(A)’s backbone(Scheme IX).

Living poly(2-alkyl oxazolines) telechelic poly-mers have been grafted onto partially deacetylatedchitins by Aoi et al. [81] in DMSO. In this solvent, thewater-soluble chitin swells to some extent, thus theamino group was used to terminate the living polyoxa-zolines synthesized by cationic ring-opening polymer-ization of the corresponding oxazolines with methyl tri-fluoromethanesulphonate (Scheme X). Full degrees ofN-substitution of available amine groups have beenobtained with grafted chain DPs ranging from 8 to 33units [82]. The number of side chains was controlled bythe amount of living polymers employed. This methodenabled the introduction of monodispersed side chains.

The grafted derivatives were miscible with PVC tovarying degrees [83,84]. They were soluble in water,DMF, and DMSO, and partially soluble in chloroform,acetonitrile, and methanol. Living poly(2-methyl-2-oxazoline) and poly(isobutylvinyl ether) cation wassuccessfully terminated by surface amino groups onchitosan powder to give the corresponding polymer-grafted chitosan [85].

According to a pioneering report published in 1999,the same research group [86] synthesized chitin deriv-atives with well-defined block copolymer side chains.In fact, they reported the first example of introductionof living block copolymer to a polysaccharide. Thegraft copolymers having monodisperse amphiphilicpoly(2-oxazoline) block copolymer side chains exhibit-ed associative behaviour and complexation withhydrophobic substances depending on the chemicalstructure. Among the derivatives, chitin-graft-[poly(2-

Scheme XI. Approaches for PEGylating chitosan, an outstanding case of grafting of a preformed polymer onto chitosan. The chi-tosan-g-PEG graft copolymer is often referred to as “PEGylated chitosan”. mPEG= methoxyterminated PEG, PNP= paranitro-phenyl, WSC= water-soluble carbodiimide, BtOH= hydroxybenzotriazole.



methyl-2-oxazoline)-block-poly(2-phenyl-2-oxazo-line)] formed cylindrical aggregates (diameter 40 nm,length 80-200 nm) in aqueous solution.

Poly(ethylene glycol) (PEG) have appeared to be avery important synthetic macromolecule in bio-scienceand technology [87,88]. The term �PEGylation� is usu-ally referred to a process involving the conjugation ofPEG with a substrate. The conjugation of PEG to drugs,especially protein drugs, is well known to enhance thesolubility and stability of the protein in solution, to alterbioavailability, pharmacokinetics, immunogenic prop-erties, and biological activities, and also to protect itfrom recognition by the immune system, prolonging itscirculation time and efficacy in vivo [89]. Severalmethods have been reported on the PEGylation ofchitn/chitosan using PEGs with various terminal reac-tive groups (Scheme XI). Harris et al. [90] have report-ed the synthesis and characterization of various func-tional PEGs useful for PEGylation.

PEGylated chitosans may be especially suitable ascarriers for delivery of anionic drugs, such as proteins,glycosaminoglycans, and DNA plasmides or oligonu-cleotides [89]. Methoxy PEG p-nitrophenol carbonate(MW 5000) was used to link PEG to chitosan througha urethane linkage [89]. The synthetic route includingthe reaction of the chitosan free amine groups withmethoxy PEG p-nitrophenol carbonate is shown inScheme XI. Grafting yields were 80-90% based on theweight of chitosan, where grafted derivatives were sol-uble in aqueous solutions at pH 6.5, contrary to highlydeacetylated chitosan.

Methoxy PEG acid (Mn = 5000) was activated witha carbodiimide/hydroxylbenzotriazole technique usedin peptide synthesis, to subsequently acylate chitosanfree amine groups [91]. The product PEGylated chi-tosan chains were obtained with degrees of N-substitu-tion (DS) ranging from 0.02 to 0.55, where the copoly-mers having DS greater than 0.10 were observed to bewater-soluble after ultrasonication. Other workers [92]reported another approach for immobilization of pro-teins with PEG tresylates and chitosan tresylates. Theyfound that the tresylate reacts by an unexpected mech-anism with this regard, however, to give a sulphonate-amide linkage as the major product rather than simpleN-alkylation. PEG tresylate also PEGylates the thiols[88], and this may sometimes be a disadvantage.

PEGylation of chitosan via reductive alkylation of

the amine group of the chitosan was firstly reported in1984 by Harris et al. [90]. This facile one-pot syntheticmethod includes the reaction of the -NH2 with -CHO ofan aldehyde-terminated PEG to form a Schiff-base. Theimine unsaturation (-CH=N-) is then reduced by sodi-um cyanoborohydride (NaBH3CN) to produce a stableN-PEGylated chitosan (Scheme XI). This approach wasthen employed by other workers to efficiently preparethe chitosan-PEG copolymeric hybrids [90,91,93].Sugimoto et al. [92,93] optimized the process to obtainpurified chitosan-PEG hybrids with DS values substan-tially higher than those reported by the pioneers (DS0.74 νs. 0.06). They then treated the hybrids with aceticanhydride to prepare chitin-PEG hybrids. The solubili-ty of the graft copolymers in water was reported to bedependent on the PEG molecular weight, the weightratio of PEG in the hybrids, DS, and degree of acetyla-tion [93]. The modification with the higher molecularweight PEG improved water-solubility of chitosankeeping the main skeleton intact. The bioactivities ofthe PEGylated chitosans were studied as well [94,95].PEG side chains with very low molecular weight (i.e.,oligo(ethylene glycol) pendant chains) were also pre-pared via the same way to achieve comb-shaped chi-tosan derivatives [96]. The tri- and tetra(ethylene gly-col) monosubstituted derivatives were characterized byhigh affinity for organic solvents as well as water insharp contrast to the original chitosan. They showedsignificant adsorption capacity toward metal cations. Itshould be pointed out that the above-described reduc-tive amination is also a potentially useful method forconjugation of any NH2 containing drugs to PEG.

The reductive amination is also possible by usingPEG propionaldehyde or PEG acetaldehyde [97] toprovide a direct amine link to PEG with retention ofbasic properties. However, the ease of polymerizationof the PEG acetaldehyde in preparation has presentedproblems in application of these methods [98]. It hasalso been found that air oxidation of PEG acetaldehydeoccurs readily and low-temperature storage under aninert atmosphere is thus necessary [99]. Bentley et al.came over these problems using a simple and reliablemethod for preparation and use in reductive aminationof PEG acetaldehyde hydrate generated in situ byhydrolysis of PEG acetaldehyde diethylacetal. Theydemonstrated the application of their approach inPEGylation of lysozyme and chitosan (Scheme XI) to

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form water-soluble methoxy PEG (mPEG) derivativesand PEG-chitosan hydrogels [99].

In a three-step synthetic approach for chitosanPEGylation reported by Ouchi et al. [91], the C-6hydroxyl group of chitosan was firstly protected bymeans of triphenylmethylation. The 6-O-triphenyl-methyl chitosan derivative was then coupled withmPEG acid (Mn 5000) in the presence of a water-solu-ble carbodiimide and a hydroxybenzotriazole. The pro-tected hydroxyl group was finally deprotected by treat-ing with acetic acid solution to yield chitosan-g-PEG inwhich the PEG has been grafted onto chitosan via anamide linkage (Scheme XI). The degree of PEG intro-duction was estimated by colloidal titration to be 25%.The article has focused on the aggregation phenome-non of the PEG-grafted chitosan in aqueous solution[91].

To control the elastic modulus of chitosan forapplying in resorbable small diameter vascular grafts,polyethylene glycol was grafted onto chitosan usingsuccinimidyl-propionate-PEG to produce PEG-chi-tosan graft copolymer [100] (Scheme XI). The graftingof PEG chains increased the distance between the poly-mer chains, thus reducing the crystallinity and resultingin a lower elastic modulus.

Triblock polyethers PEO-PPO-PEO, known asPluronic polyols or poloxamers, were grafted onto chi-tosan and poly(acrylic acid) by A.S. Hoffman et al.[101]. They first activated one OH end group of thepolyol with p-nitrophenyl (pNP) chloroformate andthen conjugated directly with chitosan (MW 750000,DD 0.8) (Scheme XII) at various pHs. Dilute solutions(2-3 wt%) of the hybrid product, chitosan-g-[poly(eth-ylene oxide)-b-poly(propylene oxide)-b-poly(ethyleneoxide)], formed gels upon warming from 4 to 34oC.The viscosity buildup increased significantly with thedegree of grafting and decreases sharply with increas-ing shear rates.

Poly(ethylene imine) (PEI), a branched or linearcommercially available polyamine, has also been graft-ed onto chitosan (Scheme XII). Kawamura et al. [102]cross-linked chitosan beads with ethylene diglycidylether and then reacted the derivative with epichlorohy-drin. The epoxidized chitosan beads were finally react-ed with PEI to achieve polyaminated highly porous chi-tosan chelating resin useful for meat ion adsorption.The authors investigated the adsorption of mercury (II)

on the resin in detail [103]. Another approach was fol-lowed by Ruiz et al. [104] to prepare highly aminatedchitosan-based chelating resins. Chitosan beads (fromspecially controlled coagulation of a chitosan solutionin a NaOH solution) were brought into contact indimethylacetamide and then rinsed PEI-impregnatedparticles were reacted with glutaraldehyde. The cross-linked imine unsaturation-containing material washydrogenated by sodium borohydride to stabilize theresin. Sorption of platinum and palladium [104], andosmium and iridium [105] has been studied by thesePEI-grafted chitosans. It was shown that the secondaryand tertiary amine functions increased the reactivity ofthe sorbent for metal ions. After re-hydration, the PEIgrafts maintained the structure of the polymer and pre-vented the collapse of the porous network andimproved diffusion properties.

Poly(β-hydroxyalkanoate)s, PHAs, the highly crys-talline, optically active, biocompatible aliphatic poly-esters elaborated by a wide variety of microorganisms,have also been conjugated with carbohydrate polymersthrough the grafting onto method. The microbial poly-ester poly(β-hydroxybutyrate), PHB, was grafted ontochitosan firstly by Yalpani et al. [106]. Thus, the chi-tosan solution in dilute acetic acid was treated withreduced molecular weight PHB (in the presence ofdimethylsulphoxide at ambient temperature for 1-5days) to afford the corresponding amide conjugates(Scheme XII). The degree of substitution of the chi-tosan amine functions in the bioconjugate was very low(DS 0.02-0.03), varying insignificantly over the rangeof PHB-chitosan ratios examined. The molecularweight of the PHB branches was around 10000.

Most recently, the synthesis of a polyurethane-grafted chitosan was reported by Silva et al. [107].They firstly prepared the urethane prepolymer by con-densation reactions of PEG of two different molarmasses and isophorone diisocyanate. In a DMF/aceticacid (1:1) medium, the NCO terminated prepolymerwas then grafted with chitosan backbone through a urealinkage (Scheme XII). The DS values varied from 0.03to 0.6 depending on the reaction conditions.

Poly(dimethylsiloxane) (PDMS)-grafted chitosanwas prepared and characterized [108]. Thus, PDMSprepolymer was synthesized by ionic ring-openingpolymerization of octamethylcyclotrisiloxane using n-butyl lithium. The tensile strength and elongation of



chitosan-g-PDMS copolymer were mostly constantregardless of the grafting percentage. While criticalsurface energy of chitosan is about 0.032 N/m, that ofthe copolymer was a little decreased to 0.025-0.029N/m by grafting PDMS onto chitosan.

Epoxy-terminated PDMS was grafted onto chitosanusing UV irradiation at room temperature withoutusing a catalyst. The product was a pH-sensitive hydro-gel without a chemical cross-linking occurrence. Infact, the PDMS substituents provided the basis forhydrophobic interactions that contribute the formationof the hydrogel network. The hydrogels exhibited highequilibrium water content in the range of 82-92%

[109].Chitosan miniemulsions were used for the synthe-

sis of epoxy particles by polyaddition. As chitosanbears amine and alcohol functions, it can react with theepoxide and can be grafted onto the particles, which areobtained by polyaddition reaction. This turned out to bea convenient technique to modify or graft the water-sol-uble chitosan with water-insoluble reaction partners,thus resulting in new and not accessible chitosan deriv-atives [110]. Here, also other biodegradable polymer(i.e., a protein) was added for hybrid formation(Scheme XII). Finally, nanocapsules consisting ofhybrid polyaddition polymers as shell and a hydropho-

Scheme XII. Various synthetic routes to chitosans conjugated with different macromolecular pendant groups through “graftingonto” method. PEI= polyethyleneimine, PHB= poly(3-hydroxybutyrate), PEO= poly(ethylene oxide). PPO= poly(propylene oxide),pNP= para-nitrophenyl, G-APG= Gluadin APG (a partially hydrolyzed wheat gluten protein, MW ~5000), PPG= poly(propylene gly-col), BPA= bisphenol A residue, PDMS=poly(dimethyl siloxane), PEG= poly(ethylene glycol), IPDI= isophorone diisocyanate (3-isocynatomethyl-3,5,5-trimethyl-cyclohexyl isocyanate).

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bic part as core were elaborated. These biocompatibleand biodegradable capsules promise applications indrug delivery.

In the course of the reaction for grafting a pre-formed polymer onto a substrate, the preformed poly-mer may be converted to another polymer with totally

different, but desirable, properties. Most recently,Zohuriaan-Mehr and coworkers have achieved a one-step reaction to prepare lightly cross-linked poly(sodi-um acrylate-co-acrylamide) grafted chitosans throughhydrolytic treatment of chitosan/polyacrylonitrileblends. According to FTIR spectroscopy, no cyanide

Scheme XIII. Chitosan modification via dendronization [112,113]. TEG= triethylene glycol, R=H.



functional group was detected in the grafted chitosan.This chitosan-based copolymeric hydrogel exhibitedsuper-swelling properties and ampholytic behaviour[111].

Dendrimer-like hyperbranched polymers, a newclass of topological macromolecules, have recentlybeen grafted onto chitosan. Tsubokawa et al. [112]reported the surface modification of chitosan powder bygrafting of hyperbranched dendritic polyamidoamine.They found that the polyamidoamine was propagatedfrom the surface of chitosan by repetition of twoprocesses: (1) Micheal addition of methyl acrylate to thesurface amino groups and (2) amidation of the resultingesters with ethylenediamine to give polyamidoaminedendrimer grafted chitosan powder (Scheme XIII).

Sashiwa et al. [113] synthesized a dendronized chi-tosan-sialic acid hybrid using convergent grafting ofpre-assembled dendrons built on gallic acid and tri(eth-ylene glycol) (TEG) backbone. Thus, sialic acid den-drons bearing a focal aldehyde end group were synthe-sized by a reiterative amide bond strategy. Polyamine-ending trivalent (1st generation: G1) and nona-valent(2nd generation: G2) dendrons having gallic acid asbranching unit and TEG as spacer arm were preparedand initially attached to a sialic acid p-phenylisothio-cyanate derivative. The focal aldehyde sialodendronswere then convergently grafted onto chitosan by reduc-tive amination in 76-80% yields. The DS of the sialo-dendrimer in the hybrid were 0.13 (G1) and 0.06 (G2).The water solubility of these novel hybrids was furtherimproved by N-succinylation of the remaining aminefunctionality.

During recent years, Sashiwa and Aiba (from theGreen Biotechnology Research Group of The SpecialDivision for Human Life Technology, National Inst.Adv. Ind. Sci. Tech., Osaka, Japan) have started exten-sive research on chitin and chitosan with collaborationof other academic/industrial centers. They often pub-lish their results as original papers under continuedtitles: �studies on chitin and chitosan� and �chemicalmodification of chitosan�. Several articles among theseseries (including the parts 3, 6, and 8-11) have beenfocused on chitosan-dendrimer hybrids (CDHs). In thesixteenth part of the latter title, for example, the synthe-sis of polypropyleneimine dendrimer-chitosan hybridhas been reported [114]. The hybrids were prepared in80-90% yield and DS of 0.11 (G1), 0.042 (G3), and

0.037 (G4). CDHs having carboxyl, ester, and PEG andvarious generations were also prepared using den-drimer acetal by reductive N-alkylation. The syntheticprocedure could be accomplished by one-step reactionwithout organic solvent [115]. The DS of the den-drimers was 0.13-0.18. Perfectly or partially water-sol-uble CDHs could be obtained. Good biodegradationwas observed in these hybrids.

Multiple Modification of Chitin/ChitosanThere are some cases in which: (i) the reactivity ofchitin/chitosan itself is insufficient to participate in thedesired reaction, (ii) the modified chitin/chitosan doesnot possess the desired properties or (iii) some site(s) ofchitin/chitosan must be protected (and finally depro-tected) to sustain during the modification reaction(s). Insuch instants, there are two general approaches viachemical modification when the graft polymerization isa certain pathway to achieve desired characteristics: (a)in-situ- and/or post-treatment of the graft copolymer,(b) graft copolymerization onto a previously modifiedchitin/chitosan. So, the products may generally bereferred to as �multiply modified� materials.

(a) Further Modification of the Graft CopolymersRecently, a hydrophobically modified chitosan, chi-tosan-g-polyacrylonitrile copolymer prepared througha ceric-induced graft polymerization [28,29], was post-treated under alkaline conditions to achieve novelpolyampholytic smart superabsorbing hydrogels withpH-reversibility and on-off switching behaviour[30,31]. The superabsorbents so called as to H-chi-toPAN, showed low salt sensitivity when the add-onvalue of the initial chitosan-polyacrylonitrile graftcoplymer was low [31]. The both modified chitosansexhibited enhanced thermal stability over chitosanitself [30]. According to the authors, the sharp pH-responsiveness behaviour of the hydrogels is expectedto be a promising factor of their possible application inmany technologies such as controlled delivery ofbioactive agents.

Another case is an in situ modification ofpoly(NIPAM) grafted chitosan in the course of the syn-thetic reaction. When the grafting reaction was con-ducted in the presence of glutaraldehyde (GA) as across-linker, thermo- and pH-sensitive interpenetratingpolymer network (IPN) hydrogels were obtained. The

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hydrogels were comparatively studied with some chi-tosan/poly(NIPAM) blend IPHs with the same compo-sition ratios [41]. The equilibrium water content of allthe IPN samples dropped sharply at pH 6 and tempera-ture higher than 30oC. Therefore, the IPN hydrogelsexhibited swelling/deswelling changes in response toexternal stimuli such as pH and temperature were rec-ognized to be useful as modulation systems in biomed-ical fields.

A similar approach was reported recently by Don etal. [43] in the case of graft copolymerization of VAconto chitosan in the presence of GA. They obtainedparticulate membranes of chitosan-g-PVAc formedafter drying the suspension mixture of the polymeriza-tion reaction [42]. The membranes were then subjectedto copper ion adsorption experiments. The mechanicalstrength of the wet membranes was improved by the in-situ cross-linking with GA. The copper ion adsorptionquantity of the membranes, however, was disfavouredby the strengthening [43].

Recently, microspheres of chitosan-g-polyacry-lamide cross-linked with GA were prepared to use forencapsulating indomethacin, a nonstreoidal anti-inflammatory drug [116]. The microspheres were char-acterized for drug entrapment efficiency, particle size,and water transport into the polymeric matrix as well asfor the drug-release kinetics.

(b) Grafting onto Pre-modified Chitin/ChitosanA few chitosan derivatives are reported to be graftcopolymerized with some vinyl monomers via the con-ventional vinyl graft copolymerization. Hyroxypropylchitosan was prepared, characterized, and subjected tograft copolymerization with methacrylic acid (MAA)using APS as an initiator [117]. The same monomerwas recently graft copolymerized onto carboxymethylchitosan (CMCTS) by APS. An optimized grafting con-ditions (for CMCTS 8 g/L) was reported to be APS 8mmol/L, MAA 2.4 mol/L, 60-70oC, 2h. Grafting per-centage as high as 1500% was reported under the opti-mal conditions [118].

Sodium salts of acrylic acid (AA) and MAA weregraft polymerized onto carboxymethyl chitosan usingAPS initiator [119]. Under a tentative conditions(CMCTS 0.02 g, monomer 1.2 M, APS 0.4 mM, 70oC,2 h), grafting percentages 875 and 933% were achievedfor CMCTS-g-PAA and CMCTS-g-PMAA, respective-

ly. The antibacterial activity of the doubly modifiedchitosans against S. aureus and E. coli was explored bythe viable cell counting method.

O-Acetyl-chitin was synthesized and subjected tograft copolymerization with MMA via a photo-inducedgrafting by Morita et al. [120]. They conducted thenon-catalytic reaction in a rotary photochemical reactorirradiating UV light with a 160 W low-pressure mercu-ry lamp at distance of 75 mm. A grafting percentage~490% was achieved under optimized conditions: O-acetyl-chitin (DS 0.81) 0.15 g, H2O 10 mL, MMA2 mL, 50oC, irradiation time 4 h.

As mentioned before, since chitin is less reactivethan chitosan, its reactivity may be enhanced throughthe addition of thiocarbonate sites along the chitinbackbone via the xanthate process, i.e. treatment withconcentrated aqueous sodium hydroxide and carbondisulphide [54]. After treating the chitin thiocarbonatederivative with ferrous ion, the complex chitin xan-thate-Fe(II) is formed. The decomposition of the com-plex leads to free Fe(II), which is believed to react withhydrogen peroxide as shown in Scheme IV to produceOH radicals that subsequently create chitin macroradi-cals upon hydrogen abstraction. Grafting percentageson the order of 80 and 40% were obtained with thisprocess using acrylonitrile and acrylic acid, respectively.

Chitosan was graft copolymerized with hydrox-yethyl methacrylate (HEMA) via a thiocarbonated chi-tosan-potassium bromate redox initiation approach[121] (Scheme XIV). For a chitosan thiocarbonatedunder the optimized conditions NaOH 1%, CS2 2%,material-to-liquor (M/L) ratio 1:20, 30oC, and 1 h, totalconversion of the monomer ranged up to 75% and amaximum grafting of 38% was achieved under the

Scheme XIV. Thiocarbonation-bromate redox initiation [121]of graft polymerization of a vinyl monomer onto chitosan (X=NH or O).



Scheme XV. Macromolecular architecture of graft copolymeric chitins with well-known structures from tosyl-, iodo-, mercapto-, anddeoxy(thiosulphato)-chitin derivatives [122-126]. Ac= acetyl, Me= methyl, Ph= phenyl, DMSO= dimethylsulfoxide.

optimized grafting conditions (KBrO3 0.6 mmol/g chi-tosan, formic acid 2 g/g chitosan, HEMA 75%, M/Lratio 1:30, 40oC, 2 h). The polymerization composite

product (including the graft copolymer, homopoly(HEMA) and unreacted chitosan) was easily cast asfilms which were featureless in the scanning electron

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microscopy.Most of the vinyl graft copolymerizations (e.g., the

above case, pesulphate- and ceric-intiating systems) arecomprised a simple manner, but the initiating site andhence the structures of the resulting copolymers are notwell-defined. Kurita et al. showed that with iodo-, tosyl-,and mercapto-chitin derivatives, graft copolymers hav-ing well-defined structures could be prepared, because itwould produce initiating species only at the positioncarrying the certain functional group [1, 122-125].

Tosylation (treatment with excess p-toluenesulphonyl chloride) was carried out on alkali chitin. 6-Tosylated chitins were then treated with sodium iodidein DMSO. The reaction proceeded smoothly to give 6-iodo-chitins that exhibited good solubility in the sol-vent [122]. On addition of a Lewis acid such as SnCl4to iodo-chitin in nitrobenzene, reactive carbeniumspecies were formed, and styrene was efficiently graftcopolymerized by a cationic mechanism in a swollenstate (Scheme XV). The grafting percentage (G%) wasreported to be up to 800%. The Mn and PDI of thegrafted polystyrene were measured to be 58000 and1.5, respectively. Irradiation of UV light on iodo-chitinin DMSO solutions gave rise to the homolysis of the C-I bonds to form carbon free radicals, and styrene wasgraft copolymerized by a radical mechanism. The graft-ing percentage is low, however, no appreciable amountof homopolymer was detected [1].

6-Mercapto-chitin is another candidate for themacroradical initiator to prepare well-defined graftcopolymers. Altough mercapto-chitin is insoluble, it isexpected to efficiently graft copolymerization owing tothe presence of the readily dissociating mercaptogroups and to swelling in organic solvents. Actually,

styrene was graft copolymerized onto chitin efficientlyin DMSO at 80oC, and the resulting G% reached almost1000 [123]. The Mn and PDI of the grafted polystyrenewere measured to be 974000 and 2.62, respectively.The G% and Mn values indicated that 4% of the mrcap-to groups were used for the graft copolymerization, anda polystyrene chain was attached on average to every45 pyranose units. MMA was also graft polymerizedonto mercapto-chitin under similar conditions toachieve chitin-g-PMMA copolymers [124] (SchemeXV). The G% value was enhanced with the amount ofthe monomer and reached above 1200 under appropri-ate conditions. The resulting graft copolymers exhibit-ed remarkable affinity for various common organic sol-vents.

Tosyl-chitin and iodo-chitin can also serve as poly-meric initiators for ring-opening polymerization of 2-methyl-oxazolines to give poly(N-acetylethyl-eneimine)s in dimethylacetamide solution at 80oC[125] (Scheme XV). Tosyl-chitin was recognized to bemore suitable than iodo-chitin judging from the G%values [1, 125]. Mn of the side chains isolated from thecopolymer of 160% grafting was 2700, indicating that18% of the tosylate groups were actually utilized forinitiating the graft copolymerization.

6-Deoxy(thiosulphato)chitin (S2O3-chitin, DS0.49) was synthesized and introduced as a precursor fornon-catalytic photo-induced graft copolymerization ofMMA, AN, AA, and AM [126] (Scheme XV). Chitinwas first tosylated and subsequently transformed intoS2O3-chitin. UV irradiation (at a fixed temperature50oC) easily proceeded the polymerization. The photol-ysis of the S2O3 groups (confirmed by IR spectra) car-ried only in quartz, not in a Pyrex tube. The monomerMMA and AN showed good activities. In the case ofMMA, under optimized conditions (S2O3-chitin 0.15 g,water 10 mL, MMA 2 mL, 1h), G% value reachedaround 600.

Chitosan powder was grafted with methyl acrylate(MA) utilizing the trichloroacetyl/Mn2(CO)10 pho-toinitiating system by Jenkins and Hudson [127]. First,highly deacetylated chitosan was N-trichloroacetylatedby trichloroacetic anthydride. The trichloroacetyl chi-tosan powder was then graft copolymerized heteroge-neously with MA using the manganese carbonyl co-ini-tiator photoactivated with 436 nm light at room temper-ature (Scheme XVI). Graft percentages greater than

Scheme XVI. Mechanism for graft copolymerization of chi-tosan (R-NH2) with methyl acrylate using a newly reportedtrichloroacetyl-manganese carbonyl co-initiator photoactivat-ed system [127]. Trichloroacetyl chitosan derivative (R-NH-COCCl3) is necessarily used in this system.



600% were obtained, while 20-30% of the poly(MA)formed was homopolymer.

In a multi-step synthetic approach for PEGylatingchitosan [91], the C-6 hydroxyl group of chitosan wasfirstly protected be means of triphenylmethylation. The6-O-triphenylmethyl chitosan derivative was then cou-pled with mPEG acid (Mn 5000) in the presence awater-soluble carbodiimide and a hydroxybenzotria-zole. The protected hydroxyl group was finally depro-tected by treating with acetic acid solution to yield chi-tosan-g-PEG in which the PEG has been grafted ontochitosan via an amide linkage (Scheme XI). The degreeof PEG introduction was estimated by colloidal titra-tion to be 25%.

Unlike the most of approaches to PEGylate chi-tosan leading to N-substituted chitosan-PEG graft

copolymer as illustrated in Scheme XI, most recently,chitosan-O-PEG graft copolymers were synthesized viaetherification of N-phthaloyl chitosan by PEGmonomethyl ether (mPEG) iodide in DMF in the pres-ence of silver oxide, and finally, deprotection of the N-phthaloylated chitosan amino group [128]. Varying theratio of mPEG iodide to chitosan, different degree ofO-substitution of mPEG to monosaccharide residue ofchitosan (5-197%) was obtained. The graft copolymerswere soluble in water and aqueous solutions of widepH range. Reduced viscosity of aqueous solutions ofthe copolymers was extremely low and similar to thatof mPEG 2000.

Ohya et al. [129] reported a novel 6-step syntheticstrategy (Scheme XVII) to tailor a new chitosan-g-polystyrene with well-known structure and amphiphilic

Scheme XVII. A 6-step synthetic strategy for synthesizing a chitosan-g-polystyrene copolymer with well-known structure havingamphiphilic properties [129]. DCC= dicyclohexylcarbodiimide, DMF= dimethylformamide, Py= pyridine, Ph= phenyl.

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properties. Thus, the amino group of chitosan was first-ly protected by phthalic anhydride. The primary alco-hol group of the N-phthaloylchitosan was protected bytritylchloride. 6-O-tritylchitosan was then preparedusing hydrazine-deprotection of the amino group. 6-O-tritylchitosan was coupled with 4-4í-azobis(4-cyanova-leric acid) (ACVA). Graft polymerization of styrene

onto the 6-O-tritylchitosan was carried out in DMFemploying the pendant ACVA moiety as an initiator.After deprotection of trityl groups of the polymeriza-tion products, chitosan-g-polystyrene graft copolymerwas obtained. Under the reported conditions (6-O-tritylchitosan (DS 24 mol%) 0.2 g, styrene-to-ACVAmole ratio 810, 60oC, 8 h), the weight percentage and

Scheme XVIII. The synthetic pathway to a novel gene delivery agent; galatosylated chitosan-graft-poly(vinyl pyrrolidone) [133].Ac= acetyl, EDC= 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride, NHS= N-hydroxysuccinimide.



Mn of polystyrene content of the copolymer was meas-ured to be around 96% and 51000, respectively. Thegraft copolymer showed a micro-phase separated mor-phology.

In another approach to modification, a polymeriz-able group is firstly introduced to the chitosan back-bone and the reactive derivative is then (co)polymer-ized. For example, treatment of chitosan with maleicanhydride led to O- and N-maleinated chitosan. Thisfully substituted derivative was then copolymerizedwith acrylamide by APS initiator to yield cross-linkedcopolymers. The products were characterized byremarkable swelling in water with a volume increase of20-150 times [130]. Using NIPAM, a similar strategywas followed to prepare pH/temperature-sensitivehydrogels [131].

Most recently, Tanodekaew et al. [132] reported thepreparation of acrylic grafted chitin for wound dressingapplication. Acrylic acid (AA) was first linked to chitinvia esterification (H2SO4, 70oC, 1 h). The acrylic dou-ble bonds acted as the active grafting sites on the sub-strate that was further polymerized with AA monomer(KPS, 65oC, 4 h) to form a network structure. Theswelling behaviour and gel strength were found todepend upon the monomer feed content. Chitin-poly(AA) 1:4 yielded optimal swelling and gelstrength. The overall results of the cellular behaviouron the modified chitin film suggested that the materialhas a potential for biomedical applications particularlyas temporary skin substitute.

In the field of gene delivery, a galactosylated chi-tosan-g-poly(vinyl pyrrolidone) was recently preparedto employ as hepatocyte-targeting DNA carrier [133].Chitosan was coupled with lactobionic acid via anactive ester intermediate using 1-ethyl-3-(3-dimethyl-aminopropyl)-carbodiimide hydrochloride (EDC).Through a �grafting onto� method, a preformed syn-thetic water-soluble polymer, i.e. monocarboxylic ter-minated poly(vinyl pyrrolidone) (PVP), was thenchemically conjugated with galactosylated chitosan(GC) utilizing EDC and N-hydroxysuccinimide (NHS)as an activator (Scheme XVIII). The complex forma-tion of GC-g-PVP/DNA complexes was confirmed byagarose gel electrophoresis. The morphology of thecomplex that observed by atomic force microscopy hada compact and spherical shape, around 40 nm particlesizes at a charge ratio of 3. The DNA-binding property

of the graft copolymer mainly depended on the molec-ular weight of chitosan and composition of the synthet-ic part of the bioconjugate.

CONCLUSION

The science and technology of chitin and chitosan areadvancing quite rapidly as a result of expanding inter-est in these biopolymers, which have unique character-istics. With regard to developing advanced functions,much attention has been paid to modification of chitinand chitosan. Graft copolymerization is considered tobe one of the most promising approaches to a widevariety of molecular designs leading to novel type oftailored hybrid materials.

The present review deals with the chemistry ofmodification of chitin and chitosan via graft copoly-merization with an emphasis on synthetic approaches.The article includes the majority of published papers inthe field. Overall, the main concluding remarks may besummarized as follows:

- Direct grafting of vinyl monomers onto the sub-strate via radical copolymerization, being a simple anduseful method of modification, have classically beenplagued by a lack of control over the mechanism. Here,there are many different reactions occurring simultane-ously, namely, initiation, propagation, etc. This compli-cated reaction system results in ill-defined structuresand non-desired homopolymer. Major efforts to reducehomopolymer formation have been attempted; howev-er, it is still generated more or less.

- Graft copolymerization of vinyl monomers ontothe substrate activated by high-energy radiation (pre-irradiation technique) can solve the homopolymer for-mation problem in some extent.

- Living polymerization systems may be consideredas a good solution of the homopolymer problem, asmentioned in the case of grafting of living poly(2-alkyloxazolines) or living poly(isobutylvinyl ether) ontopartially deactylated chitin.

- Grafting onto special derivatives of the biopoly-mers (e.g. tosyl-, iodo- or mercapto-chitin) is a certainapproach to achieve graft copolymers with well-defined structures.

- By employing the �grafting onto� method, a highlevel of control on the macromolecular structure is pos-

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sible without suffering from the above mentioned prob-lems. This multi-step approach, however, is time con-suming and cost non-effective, especially when a cer-tain functional group of the trunk polymer is necessaryto be protected (and then deprotected).

- While the unique structure and properties of chi-tosan (e.g., biological and cationic polymer characteris-tics) are mainly originated from its free amino group,the most modification strategy lead to N-substituted chi-tosan copolymers, in which the majority of the aminogroups are blocked. Therefore, retaining these function-al groups is an important challenge for preserving theintrinsic properties of the biopolymer in the hybridmaterial. Protection of the NH2 group, usually asphthaloyl group, grafting treatment(s), and deprotectionof the amino groups may be considered as an appropri-ate sequence result in desirable regioselectivity.

- Chitosan and partially deacetylated chitin havebeen more frequently subjected to the grafting ratherthan chitin itself. The reason is mainly related to insol-ubility of natural chitin in most of possible reactionmedia. Therefore, the intractability of chitin has yetremained as a challenge for the researchers who insistto modify chitin. Anyhow, it should be pointed out thatthe harsher the reaction conditions, the higher the poly-saccharide molecular weight loss will be.

This contribution is also intended to stimulate fur-ther research on chitin and chitosan modification inorder to use these precious renewable biomaterialsinstead of the fossil-based materials used in bio-scienceand technology. Fast-growing academic/industrialactivities in this regard, as obviously exhibited in Fig-ure 1, ensures that chitin will not be a �biomaterial inwaiting� any more.

ACKNOWLEGEMENTS

The author is very grateful to Professor A. Pourjavadiand the PhD students G.R. Mahdavinia and H. Hossein-zadeh from Chemistry Department of Sharif Universi-ty. Technology, for their main role in our chitin/chi-tosan-related collaborations. The author’s specialthanks are due to the IPPI Director, Professor H.Mirzadeh, for his continued support of the Superab-sorbent Hydrogel Division and its recent orientationtoward the biopolymer-based hydrogels. His moralsupport was a major driving force to write this paper.

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