Appendix A: Future Directions
A.7.3 Example for the Interaction of Structural, Aerodynamic,
10.1 Chitosan-Based Gels
10.1.2 Supramolecular Interactions and Gel Formation
sites, for example, reacting with glutaraldehyde (GA) [2,3], glyoxa [4], proanthocyanidin [5], 1-ethyl-3-(3-dimethyl aminopropyl) car- bodiimide (EDC), or genipin [6] to form imine cross-linking between linear chitosan chains that lead to gel formation.
Other components can be added, such as additional polymers to form a hybrid polymer network (HPN) or a semi- or full-IPN (interpenetrating polymer network). Auxiliary molecules can also be used to initiate reactions during the preparation of the network. An IPN is defi ned as a combination of two cross-linked polymers; at least one of them synthesizes or cross-links in the presence of the other. If one of the polymers is linear (without being cross-linked), a semi-IPN results. Th e IPN technique can be used to improve the properties of chitosan-based gels.
Lee and Chen [7] examined GA-cross-linked chitosan–
isopropylacrylamide(chitosan/PNIPAAm) semi-IPN and IPN hydrogels. Th ey found that the swelling ratios of IPN gels are lower than those of semi-IPN gels and the reason is that the addition of cross-linked chitosan in the PNIPAAm hydrogel makes the net- work structure of the gel denser and more hydrophobic. Th e swell- ing ratios of PNIPAAm/chitosan gels slightly decrease with the addition of chitosan, but the swelling ratios are not signifi cantly aff ected by the amount of chitosan added to the PNIPAAm gel.
Because of the existence of many –NH2 groups on chitosan chains, hydrogels composed of chitosan are also used as pH- sensitive materials [8]. A novel pH-sensitive hydrogel composed of N,O-carboxymethyl chitosan and alginate cross-linked by genipin (NOCC/alginate) and the swelling ratio of this hydrogel at pH 1.2 was about 2.5 within 30 min subsequent to swelling, while it was approximately 6.5 at pH 7.4 (as shown in Figure 10.1). Th is fi nding may contribute to the fact that, at low pH, the swelling ratio was limited due to formation of hydrogen bonds between NOCC (–COOH and –OH) and alginate (–COOH and –OH). However, at higher pH 7.4, the carboxylic groups on the genipin-cross-linked NOCC/alginate hydrogel became progressively
ionized (–COO–). In this case, the hydrogel swelled more sig- nifi cantly due to a large swelling force created by the electrostatic repulsion between the ionized acid groups. Th e swelling ratio of the hydrogel with alginate was signifi cantly lower than that without alginate due to the formation of hydrogen bonds between NOCC (–NH2 and –OH) and alginate (–OH) [9].
Yao et al. [10] examined GA cross-linked chitosan–poly- (propylene glycol) (PPG), semi-IPN as a pH-sensitive hydrogel.
Chitosan–PPG semi-IPN shows pH-dependent swelling; the highest swelling degree (SD) is at pH 4.0 and the lowest at pH 7.0 (Figure 10.2). Th e structure features of chitosan–PPG are reversible (e.g., –NH3 to –NH2) when the gels are transferred from a pH 1.0 to a pH 7.8 buff er [11], because it is a hydrophobic and water-insoluble polymer and the incorporated PPG is expected to decrease the equilibrium SD in a low pH medium.
In addition, linear PPG in the network can enhance the fl exi- bility of a semi-IPN. For the same reason, at a higher pH it may not fully ionize all the –NH2, the pH-sensitive, biodegradable, semi-IPN chitosan–polyvinyl alcohol (PVA) hydrogel cross- linked with GA and shows low-bound water concentrations (CBW, mg/mg dry hydrogel). On the other hand, if the pH goes too low, the high concentration of H+ will actually impair the swelling, so the maximum CBW, mg/mg dry hydrogel would appear at pH 3 medium (Figure 10.3) [12,13].
Th e SD of hydrogels based on [N-(2-carboxybenzyl) chitosan, CBCS] (CBCSG) was signifi cantly increased with the raise of the pH in the range of pH 5.0–9.0 and the lowest swelling occurred almost always at pH 5.0 among CBCSGs, as shown in Figure 10.4 [14]. Th is maybe depends mainly on the osmotic pressure diff er- ence between the inside of the gel and the surroundings caused by the redistribution of mobile ions [15], and along with decreas- ing pH. Th e amount of –COO is gradually reduced inside the hydrogels, which leads to a decrease in osmotic pressure and makes the SD of the hydrogels smaller. When the pH was reduced to 1, the –COO was completely inhibited, and the dominant ionic groups were –NH3
+, but the number is small because the majority of the –NH2 groups were linked, therefore the CBCSGs swelled sluggishly as pH diminished in the range of pH 1.0–5.0.
0 0 4 2 6 8 10 12 14 16
50 100 150 200
(n = 5)
250 300
Time (min)
Swelling ratio
NOCC:Alginate = 1:0, 0.75mM Genipin, pH 1.2 NOCC:Alginate = 1:1, 0.75mM Genipin, pH 1.2 NOCC:Alginate = 1:0, 0.75mM Genipin, pH 7.4 NOCC:Alginate = 1:1, 0.75mM Genipin, pH 7.4
FIGURE 10.1 Swelling characteristics of the NOCC and NOCC/
alginate hydrogels cross-linked with 0.75 mM genipin at pH 1.2 or 7.4 at 37°C.
01 10 20 30 40 50 60
2 3 4 5 6 7
pH
Equilibrium degree of swelling
8 9 10 11 12 13
FIGURE 10.2 Equilibrium degree of swelling vs. pH for the semi-IPN, synthesized from diff erent amounts of cross-linking agent. Molar ratios of –CHO:–NH2 are 2.21° and 4.83°, respectively.
Collagen is known to be the most promising of biomaterials and has been found diverse applications, which can form a com- plex with chitosan [16]. Th e collagen–chitosan complex can be used to mimic the components of the native extracellular matrix (ECM). It has also been reported that the hybrids of collagen–
chitosan manufactured by cross-linking [17]. Gelatin is a denatured form of collagen, composed of glycine, praline, hydroxyproline, arginine, and other amino acids. Th e amphiphilic protein (iso- electric point = 4.96) can provide amino groups for cocross-linking with chitosan via GA thus preparing a chitosan–gelatin (CG) HPN. Th e pH-sensitive swelling behavior of HPN gel is displayed in Figure 10.5. Th e data show that the degree of swelling declines
sharply at pH 7.0 and this can be explained by the fact that the hydrogen bonds within the chirosan–gelatin HPN dissociate in an acidic medium [18].
Th e properties of many chitosan-based hydrogen bond complexes depends on not only pH but also temperature. Carmen et al. investigated the swelling behavior of diff erent crosslink chitosan–poly (N-isopropylacrylamide) interpenetrated networks (CS/PNIPA). Th e SD of all the CS/PNIPA IPN decrease with the enhancement of temperature, and the SD at low pH 3 is higher than that at high pH 8 for any temperature (as shown in Figure 10.6) [19]. Temperature/pH-sensitive poly(2-ethyl-2-oxazoline)/
chitosan(PEtOz/CS) IPN hydrogels exhibited considerable shrink- age over the temperature range of 35°–45°, and thermoresponsive behavior has decreased with increasing chitosan content [20].
Th e electrical fi eld is another important factor, which infl u- ences the properties of chitosan-based hydrogels. Th e response of electrosensitive hydrogels is generally exhibited in the form of either swelling and shrinking or bending behaviors. Figure 10.7 shows the degree of bending of the chitosan–poly(hydroxyethyl methacrylate) semi-IPN hydrogel as a function of the various applied voltages in an aqueous 1.0% by weight NaCl solution.
Th e degree and the increase in the speed of bending as well as an increase in the voltage applied across the hydrogel indicates that the bending is induced by the electric current, however, the bending did not occur in pure water [21]. Th e chitosan–polyal- lylamine IPN hydrogel exhibited the same characteristic [22].
10.1.2.2 PEC
PEC are generally obtained either by the reaction of polycations and polyanions or by polymerizing monomers that have suitable functional groups onto polymeric templates of known structures.
PECs have numerous applications such as membranes, medical prosthetic materials, environmental sensors, and chemical detectors. Chitosan, a cationic polysaccharide, has been com- plexed with anionic polymers. Th e properties of PEC are mainly 1
0.0 1.0 2.0 3.0 4.0
2 3 4 5 6 7
pH Bound water concentration (CBW) g/g dry gel
8 9 10 11 12
FIGURE 10.3 Bound water concentration in the chitosan–PVA hydrogel at diff erent pH values (chitosan:PVA molar ratio 1:10, GA concentration 33.3 M).
0 0 2 4 6 8 10 12 14
4 8 12 16 Time (h)
Swelling ratio
pH 9.0 pH 5.0 pH 1.0
pH 7.4 pH 3.0
20 24
FIGURE 10.4 Swelling behavior of CBCSG in diff erent buff er solutions.
01 2 4 6 8 10
2 3 4 5 6 7
pH
Equilibrium degree of swelling
8 9 10 11 12 13
FIGURE 10.5 Swelling behavior of CG hybrid network specimen with a –CHO:–NH2 molar ratio of 10 in solutions of diff erent pH with ionic strength I= 0.1 at 37°C.
0 20 Degree of bending (%)30 40 50 60 70 80 90
5 10 15
Voltage (V)
20 25
FIGURE 10.7 Degree of bending of chitosan/PHEMA semi-IPN in response to various voltages in a 1.0 wt % NaCl solution at T = 35°C.
determined by the degree of interaction between the individual polymers. Th is latter condition depends essentially on their global charge densities and determines their relative composition in the PEC. Th e lower the charge density of the polymer, the higher is the polymer proportion in the PEC, since more polymeric chains are required to react with the other polymers. A PEC can be formed via two ionizable polymers with opposite charges individually. Th is means that a PEC-forming reaction can only occur at pH values in the vicinity of the pKa interval of the two polyelectrolytes.
CS is a pH-sensitive polymer (pKa is ca. 6.5) whose amino groups can be protonated depending on the pH of the environ- ment. When CS is mixed with ampholytic gelatin (Gel, its pHiso
is approximately 4.7), which in the case of pH of medium above the isoelectric point of Gel, and where a net charge is negative one, there occurs electrostatic interaction between the ammo- nium ions of CS salts and carboxylate groups of Gel. Yao et al.
[23] found that there was strong interaction between Gel and CS in the aqueous medium that was enough to form PEC in situ, and the CS/Gel PEC only yielded at pH value larger than 4.7 and above pH 6.2 CS could precipitate from the solution.
Pectin is an acidic polysaccharide that has a repeating unit of α-(1,4)-l-rhamnose units. A PEC has been formatted from anionic pectin and cationic chitosan. Th e PEC swells obviously at pH < 3 and pH > 8, and does not swell in the range of 3 < pH < 8.
Moreover, its degree of swelling in an acidic medium is by far higher than that in an alkaline medium. Th e swelling of the PEC correlates with its composition and is also aff ected by the degree of deacetylation (DDA) and the methoxy level of pectin [24].
Alginate comprises a linear chain of (1,4)-linked β-d-mannuronate and α-l-glucuronate residue arranged blockwise. A gel-like chitosan/
alginate PEC was at one time formed with coexisting polyions of opposite charges (–NH3+ and –COO−). Polysaccharides (chitosan and alginate) have bulky pyranose rings and a highly stereoregular confi guration in their rigid linear backbone chains. At a given pH, the composition of the PEC shift ed to a lower alginate content as the degree of N-acetylating of chitosan increased and the higher the pH, the lower the alginate content of the PEC for a given chitosan sample [25]. W. Argüelles-Monal et al. [26] found that the degree of polymer- ization of chitosan and the chemical composition of alginic acid have shown to have an infl uence on the PEC formation.
Yoshitsune et al. [27,28] found that the chitosan/polyalkylene- oxide–maleic acid copolymer (CS/PAOMA) PEC fi lms swelled at a low pH, shrunk at pH between 4.8 and 6.5 and their swelling enhanced aft er pH 6.5. Th is swelling behavior can be attributed to the electrostatic interaction between the protonation of amino groups in CS and carboxyl groups in the PAOMA. Th e pKa of CS is 6.5 and the pKa of PAOMA is 4.8. In diff erent pH medium corresponding groups (e.g., NH3+ and COO−) play diff erent roles between intermolecules.
10.1.2.3 Grafted and Block Network
Many polymers, such as phosphatidylethanolamine, poly- (ethylene glycol) (PEG), poly(vinyl acetate), poly(vinyl alcohol), poly(3-hydroxyalkanoate), linoleic acid, poly(l-lactic acid), and 0
20 40 60 80 100
0 20
20 25 30 35 40 45
Temperature (⬚C) pH 8 (Phosphate buffer)
pH 3 (Glycine buffer)
40 60 80 100
Degree of swelling (%)Degree of swelling (%)
FIGURE 10.6 Dependence of the degree of swelling (%) of PNIPA/CS IPNs on the pH and temperature of the aqueous medium; IPN-0 (O), IPN-1 (), IPN-3 (), IPN-5 (▼), and IPN-7 (▼). PNIPA/chitosan-I IPNs exhibited a similar pattern. (IPN-X, where X denotes the concentration (Vol.%) of GA.
so on, have been graft ed onto the chitosan in order to obtain some type of special performance.
PEG is a water-soluble polymer that has low toxicity and immunogenicity. Hydrophilic PEG can form a diff use layer con- taining water molecules, which has protein resistance when PEG is adsorbed on a surface. Many researchers have turned their attention to graft copolymers of chitosan and PEG, as shown in Figure 10.8, because of their high biocompatibility [29–31].
Hydrogen bonds exist between the hydrogen of the amino groups of chitosan and the oxygen of the polyether in alkaline regions, whereas the hydrogen bonds dissociate at acidic pH. Additionally, the SD is very sharp at a high pH 6 [32], and the polymers were soluble in water over a wide pH range within a short time and the viscosities of the solution were very low. Th e degradation rate of the copolymer declined with the enhancing degree of substi- tution of PEG to chitosan, which was probably due to the relative stability of PEG to lysozyme [33].
Poly(α-hydroxy acids) generates acidic degradation prod- ucts at the implanted site, which evokes undesirable tissue reaction [34]. Th e acid by-product may lead to local distur- bances due to poor vascularization in the surrounding tissue.
Chitosan may be combined with acid-producing biodegradable polymers, so that local toxicity due to the acid by-products can be alleviated. Yao et al. obtained the cytocompatible poly(chitosan-g-l-lactic acid) through graft ing oligo(l-lactic acid) onto the amino groups in chitosan without a catalyst.
Th ese graft copolymers had a higher strength than chitosan and their swelling behavior was infl uenced by pH (as shown in Figure 10.9). When a pH < 2 and with an increase in the buff er pH, the concentration of the charged ionic groups in the fi lms also increased. Th e swelling of the samples were shown to increase due to the enhancement of the osmotic pressure and charge repulsion. While at a higher pH, the degree of ioniza- tion was reduced due to the deprotonation of the amino units of chitosan and the swelling of the fi lms decreased. In addi- tion, the hydrophobic side-chain aggregation and hydrogen bonds in the copolymers were much stronger, which also lead to lower swellability of the samples [35,36].
10.1.2.4 Self-Assembly
Th e technique of self-assembly layer by layer (LbL) PEC multi- layer fi lms opens up many new opportunities for us to achieve the
ideal model surface whose properties are controllable [37]. Th e self-assembly polyelectrolyte systems are sensitive to the fabrica- tion orders in terms of the orders of static electric interactions. It has long been known that one of the challenges of the materials construction is the controllable properties of the materials, including shape, size, morphology, thickness, roughness, chemical and transport properties, degree of interpenetration [38], and surface evidence associated with the mechanisms of the absorp- tion and formation of polyelectrolytes on charged surfaces [39].
Poly(acrylic acid) (PAA) is a synthetic anionic electrolyte that has been synthesized for use in transmucosal drug delivery and self-curing membranes with CS. CS-PAA with a nanofi brous structure have been formed by self-assembling. Th e nanofi bers are of diff erent lengths ranging from 1 μm to several microns, but the widths of the nanofi bers are on a nanoscale from 50 to 150 nm. In the CS/PAA system, ionic interactions occur between the two oppositely charged macromolecules forming a polycom- plex that precipitates as an insoluble aggregate because the charged groups, which are responsible for solubility (carboxylic groups of PAA in the form of COO− and amino groups of CS in the form of NH3+), are involved in the complex [40].
Th e formation of nanoparticles by self-assembly is an energy- saving process. Self-assembling nanoparticles can be formed spontaneously under mild conditions. Figure 10.10 shows the OH OH OH
HO HO HO
NH NHAc NH2
X = Linker
X z x y
n
O O
O O O
O O O O
OMe
FIGURE 10.8 Structure of chitosan-g-PEG.
0 0 4 8 12 16 20 24
2 4 6
pH
CL-5 CL-3 CL-1
Swelling degree
8 10 12 14
FIGURE 10.9 Eff ect of the LA/CS feed ratio and pH value on the equilibrium water uptake of the CL fi lms (CL-1, CL-3, and CL-5 denote that LA/CS = 0.5, 2.0, and 4.0(w/w), respectively).
formation process for chitosan/carboxymethyl cellulose (CS–
CMC) nanoparticles. Th e size of distribution was relatively narrow and the mean particle size decreased from 226 to 165 nm with the decreasing molecular weight of chitosan hydrolysate from 9.5 to 6.8 kDa. Moreover, with a decreasing pH, the aver- age size was enhanced however; the size was not aff ected by low temperature [41].
10.1.2.5 Coordination Complexes
Chitosan has one amino group in addition to hydroxyl groups in the repeating unit. So chitosan can coordinate strongly with transition-metal ions [42] where the precious metal and rare metal ions form coordination complexes [42]. Th ey may be used as absorbents for metal ions and separation membranes. Th e order for the stability contents of these chitosan–transition- metal complexes is Mn <Fe<C<Ni<Cu<Zn, which is known as the Irving-Williams order. Th e diff erence in the stability of com- plexes may cause a variance in the degree of swelling of a com- plex membrane whose metal/glucosamine ratio ranges from 1:8 to 1:64 [43]. Domard et al. [44] confi rms that in a weak alkaline medium, the free amine function is the most favorable site for the coordination with one copper II ion, leading to a stable pend- ing complex.