In Situ Crosslinkable Hydrogel Formed from a Polysaccharide-Based

Biomacromolecules 2009, 10, 959–965

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In Situ Crosslinkable Hydrogel Formed from a Polysaccharide-Based Hydrogelator Fei Song, Li-Ming Zhang,* Nan-Nan Li, and Jun-Feng Shi Laboratory for Polymer Composite and Functional Materials, Institute of Polymer Science, School of Chemistry and Chemical Engineering, Sun Yat-Sen (Zhongshan) University, Guangzhou 510275, China Received December 30, 2008; Revised Manuscript Received February 2, 2009

In situ crosslinkable hydrogel formed from an amphiphilic amylopectin-based hydrogelator in aqueous solution was investigated with respect to its viscoelasticity, structure as well as protein encapsulation and release. Different from the physical hydrogel formed from an aqueous amylopectin system of sufficiently high concentration, such a hydrogel could be formed rapidly at room temperature and exhibit enhanced viscoelastic properties, mechanical strength, and shear thinning behavior. In addition, it has a more complex network structure with a higher fractal dimension due to intermolecular hydrophobic interactions and macromolecular chain entanglements. By circular dichroism analyses and in vitro release experiments, this hydrogel material was found to have a great potential as new matrix for the entrapment and sustained release of bovine serum albumin.

Introduction In recent years, there has been increasing interest in water gelation by amphiphilic organic molecules with hydrophobic groups to promote aggregation and hydrophilic groups to provide solubility.1,2 Such a hydrogelation can occur usually under mild conditions (room temperature and physiological pH) without the help of any chemical crosslinking by radical polymerization, chemical reactions of complementary groups, or high energy irradiation. Therefore, it is especially suitable for the in situ encapsulation of bioactive molecules such as pharmaceutical proteins, living cells, and drugs.3 Up to now, some small organic amphiphiles have been reported as effective hydrogelators for the formation and applications of some supramolecular hydrogels. For example, Friggeri et al.4 used N,N′-dibenzoyl-L-cystine as the hydrogelator to form the supramolecular hydrogel for the entrapment and release of quinoline derivatives; Cao et al.5 entrapped salicylic acid into the supramolecular hydrogels formed from L-phenylalanine derivatives as the hydrogelators; Hamachi and co-workers6 reported the thermally controlled release of DNA from supramolecular hydrogels formed by glycosylated R-amino acids; Heeres and co-workers7 studied the supramolecular hydrogels as the drug carriers, which could be formed by cyclohexane-based hydrogelators; and Jayawarna and co-workers8 fabricated the supramolecular hydrogels for three-dimensional cell culture through the self-assembly of fluorenylmethoxycarbonyl-dipeptides. In contrast, few studies have dealt with water gelation by high-molecular-weight organic amphiphiles as the polymeric hydrogelators. As a kind of hydrophilic polymers, water-soluble polysaccharide derivatives have broad importance in many applications where the environmental considerations, biodegradability, and biocompatibility take a growing concern.9-28 Their functions depend greatly on the properties they impart to solutions or hydrogels. For specific applications, optimizing the final properties by a suitable control of the chemical structure and physical properties is necessary. In this context, the incorporation of hydrophobic groups onto hydrophilic polysaccharide chains * To whom correspondence should be addressed. Tel./Fax: (+86) 2084112354. E-mail: [email protected].

provides amphiphilic polysaccharides with adaptive properties. In our previous studies, we carried out the hydrophobic modification of sodium carboxymethylcellulose (NaCMC) by the graft copolymerization with diallyllaurylamine or dimethyloctyl(2-methacryloxyethyl)ammonium bromide and found that the resultant polysaccharide amphiphiles could exhibit enhanced salt-, temperature-, and shear-tolerant properties in aqueous solutions when compared with the unmodified NaCMC.29,30 Recently, we prepared novel conjugates of some hydrophilic polysaccharides such as dextran, maltoheptaose, alginate, or carboxymethylcellulose with hydrophobic aliphatic polyesters or cholestanol and found that these amphiphlic polysaccharide derivatives could self-assemble in aqueous solutions into nanosize polymeric micelles suitable for controlled drug delivery.31-36 In this work, we synthesize a new polymeric hydrogelator by grafting lauryl chains onto the amylopectin backbone and find that such a polysaccharide amphiphile could self-assemble rapidly in water to form physically crosslinked hydrogel networks. To understand the properties and structure characteristics of the resultant hydrogel, dynamic and steady rheological properties have been studied as a function of the concentration of this polysaccharide-based hydrogelator, and a fractal analysis has been carried out by relating the viscoelastic properties to a scaling model. In addition, the suitability of using this polysaccharide-based hydrogelator for the in situ encapsulation and controlled release of a model protein has been examined by means of circular dichroism analyses and in vitro release experiments.

Experimental Section Materials. Amylopectin (amylose-free) was purchased from Kasei Kogyo Co. Ltd. Lauryl bromide (98%, Laboratory) was supplied by Alfa Aesar Co. Pyrene and bovine serum albumin (BSA) were purchased from Sigma. Phosphate buffered saline (PBS) was provided by Guangzhou Chemical Company in China. All other chemicals and reagents used were of analytical grade. Preparation of Amylopectin-Based Hydrogelator and Its Structural Characterization. In a typical experiment, 2.5 g amylopectin was dissolved in dry dimethylsulfoxide (50 mL) at 70 °C under vigorous

10.1021/bm801500w CCC: $40.75  2009 American Chemical Society Published on Web 03/10/2009

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stirring. Then, 3.7 mL of LaB and 1.25 mL of pyridine were added into the reaction system. After the reaction at 50 °C for 24 h, the reaction mixture was poured into ethanol. The resultant amylopectin derivative, namely, La-amylopectin, was filtered off, washed several times by ethanol, and finally dried in a vacum at 40 °C for 48 h. La-amylopectin was obtained with the yield of 88% and used as the hydrogelator. For the structural characterization of La-amylopectin, 1H NMR spectrum was recorded on a Varian Mercury-Plus 300 NMR spectrometer (Varian, U.S.A.) by using dimethyl sulfoxide as the internal standard, and the X-ray diffraction (XRD) analysis was performed with the help of a Rigacu D/MAX 2200 VPC diffractometer (Japan) using a monochromatized X-ray beam with nickel filtered CuKR radiation. For a comparison, the native amylopectin was also investigated for its structure characterization by means of 1H NMR and XRD analyses. Detection of Hydrophobic Microdomains. The hydrophobic microdomains in aqueous solutions of La-amylopectin were detected by fluorescence spectroscopy. For this purpose, pyrene was initially dissolved in acetone and then diluted in deionized water. The sample solutions were prepared by mixing aqueous pyrene solution with aqueous La-amylopectin solution, and were kept at 4 °C overnight before the measurements. The pyrene concentration was kept to be 1.0 × 10-6 mol/L in all samples. The fluorescence spectra were recorded on a RF-5301PC luminescence spectrometer (Shimadzu Co., Japan). The excitation wavelength was 330 nm and the fluorescence emission spectra were recorded in the range from 350 to 500 nm. For a comparison, aqueous solution of the native amylopectin was also investigated by fluorescence spectroscopy. Preparation of Aqueous Polysaccharide Systems and Their Rheological Charaterization. For the rheological experiments, aqueous systems of La-amylopectin and the native amylopectin were prepared in deionized water, and were kept at 4 °C overnight before the masurements. All rheological measurements were performed with the help of an advanced rheometric extended system (ARES, TA Co.), fitted with a parallel plate geometry (25 mm diameter). Four kinds of rheological analyses were carried out: (1) dynamic strain sweep tests for the investigation of the system structure. The applied strain ranged from 0.08 to 500% (1 rad/s and 25 °C). Over a certain strain, a drop in the modulus was observed, which corresponded to the breakdown of the hydrogel structure. The critical strain value (γ0) was determined from the storage-strain profiles of the system; (2) dynamic frequency sweep tests for the investigation of the viscoelastic properties and the determination of the gel point. The frequency applied to the sample system increased from 0.1 to 100 rad/s (25 °C). Depending on the viscoelastic properties of each sample, a suitable strain was used to ensure the linearity of dynamic viscoelasticity; (3) stress relaxation tests for the investigation of the hydrogel strength and relaxation behavior (10.0% strain and 25 °C); (4) steady shear tests for the investigation of the flow behavior under shear stress. The applied shear rate increased from 0.1 to 100 s-1 (25 °C). Scaling and Fractal Analyses of Viscoelastic Properties. For aqueous systems of La-amylopectin and the native amylopectin, the fractal analyses were carried out by relating the viscoelastic properties to a scaling model developed by Shih et al.,37 in which an aggregate hydrogel may be considered as closely packed fractal flocs with the elastic property. The strong-link or weak-link character of the hydrogel is dependent on the strength of the interfloc links relative to that of those within the floc. In the strong-link regime, the dependence of the hydrogel elastic constant, G0, and the upper limit (the critical strain) of the hydrogel linear viscoelastic region, γ0, on the particle (in this study, La-amylopectin or native amylopectin) concentration, φ, could be described as

G0 ∼ φ(3+x)/(3-Df)

(1)

γ0 ∼ φ-(1+x)/(3-Df)

(2)

where Df is the fractal dimension of the flocs (Df e 3), and x is the backbone fractal dimension of the flocs. In the weak-link regime, the dependence of G0 and γ0 on φ could be expressed as

G0 ∼ φ1/(3-Df)

(3)

γ0 ∼ φ1/(3-Df)

(4)

Characteristically, γ0 increases with φ for weak-link hydrogels, whereas an inverse dependence is found for strong-link hydrogels. For each sample system, the maxmium value of the elastic modulus in the linear region of the strain sweep measurement was collected and taken as the G0 value,37 and the strain amplitude at which the elastic modulus begins to decrease by 5% from its maxium value was taken as the γ0 value.38 Scanning Electron Microscopy Observation. For scanning electron microscopy (SEM) observation, the hydrogel samples were allowed to dry in a vacuum to a constant weight, and the resultant dried gels were then coated with a thin layer of gold. The SEM images were obtained by using a Hitachi S-520 scanning electron microscope (Hitachi Co., Japan). Protein Encapsulation and Circular Dichroism Experiments. For the in situ encapsulation of BSA, aqueous BSA solution was first prepared in distilled water, and 1.0% La-amylopectin was then added to induce water gelation at room temperature (25 °C). Circular dichroism (CD) spectroscopy was used to measure the conformation change of BSA before and after the encapsulation. For this purpose, the hydrogel samples were manually crushed with a glass rod and suspended in 0.01 mol/L phosphate buffer (pH 7.4). The CD spectra were recorded on a J-810 circular dichroism spectrometer (JASCO, Japan) with a 1.0 cm path length rectangular quartz cell controlled by a thermoelectric cell holder. Data were collected from 195 to 260 nm, at 0.2 nm intervals with 20 nm/min scan speed, 2 nm bandwidth, and 16 s response. To investigate the thermal stability of the native BSA and encapsulated BSA, the changes of CD spectra with temperature were monitored at 222 nm in 0.01 mol/L phosphate buffer (pH 7.4). In Vitro Release Experiments. For the resultant hydrogels loaded with various amounts of BSA, their in vitro release experiments were carried out. For this purpose, the BSA-encapsulated hydrogel samples were immersed in 20 mL of PBS (pH 7.4) at 37 °C, and their release profiles were studied. At predetermined time points, 2 mL of the solution was taken out and a 2 mL fresh PBS was added back to maintain the same total solution volume. The percentage of cumulative amounts of release BSA was calculated from standard calibration curve. All release studies were carried out in triplicate.

Results and Discussion The amylopectin-based hydrogelator (La-amylopectin) used in this study was prepared by the covalent fixation of lauryl chains onto the amylopectin backbone. To confirm this, 1H NMR and XRD analyses were carried out for the native amylopectin and its amphiphilic derivative. From the 1H NMR spectra shown in Figure 1, it was found that the spectrum of the amylopectin derivative showed not only the characteristic peaks for the proton species in the glycopyranan ring of the native amylopectin at 3.0-5.5 ppm,39 but also new peaks for the proton species of the lauryl chain at 0.8-1.5 ppm. From the XRD patterns shown in Figure 2, it was found that the strong reflections of the native amylopectin at 14.7, 17.8, and 22.6 °, which resulted from high crystallization,40 were not observed in the pattern of its derivative. This could be attributed to the disruption of the

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Figure 1. 1H NMR spectra of native amylopectin and its amphiphilic derivative (La-amylopectin) in D2O.

Figure 2. XRD patterns of the native amylopectin and its amphiphilic derivative (La-amylopectin).

Figure 3. Optical photos for (a) instant gelation of aqueous 1.0% Laamylopectin system and (b) no gelation of aqueous 2.0% amylopectin system at room temperature (25 °C).

crystalline structure by the incorporated lauryl chains. With 1H NMR spectroscopy, the molar substitution degree of the lauryl chains, defined as the average number of the lauryl chains per anhydroglucose unit (AGU), was calculated to be 1.7% by using the integrated intensities of the signals from the methyl protons in the lauryl chains [H(CH3)] and the sum of the integrated intensities of the signals from the anomeric protons (H-1) according to the method reported by Richardson et al.41 For such an amphiphilic amylopectin derivative, we found that it could be easily dissolved in water and display instant gelation ability for its aqueous 1.0% solution at room temperature, as shown in Figure 3. In contrast, no gelation phenomenon was observed at room temperature for aqueous solution of the native amylopectin even at a higher concentration of 2.0% (Figure 3). Figure 4 shows the angular frequency dependence of storage modulus (G′) and loss modulus (G′′) for aqueous systems of

Figure 4. Elastic modulus (G′, filled symbol) and viscous modulus (G′′, open symbol) as a function of frequency for (a) aqueous native amylopectin systems with different concentrations and (b) aqueous La-amylopectin systems with different concentrations. The data have been vertically shifted by a factor of 10a with given a to avoid overlapping.

the native amylopectin and La-amylopectin. The data were vertically shifted by a factor of 10a to avoid overlapping. For aqueous systems of native amylopectin, the G′ and G′′ were too small to be measured accurately even at a concentration of 1.5%. Only when the concentration of native amylopectin was greater than 2.0%, the system showed the viscoelastic behavior with dominating elastic property. In contrast, a low concentration (0.5%) of La-amylopectin could ensure the accurate measurements of the dynamic moduli. Moreover, the G′ value was observed to be considerably greater than the G′′ value for aqueous La-amylopectin system, even at a lower concentration of 1.0%. At the same polysaccharide concentration, aqueous La-amylopectin system has greater G′ and G′′ values when compared with aqueous amylopectin system. When La-amylopentin concentration was greater than 0.5%, a higher degree of elasticity was observed for aqueous La-amylopectin system, with G′ only weakly dependent on frequency and more greater than G′′ over the entire frequency range. These results show that aqueous La-amylopectin system has a strong liquid-solid transition ability and can behave as a strong viscoelastic hydrogel at certain concentration. Further investigation was dealt with the determination of the gel point for aqueous systems of the native amylopectin and La-amylopectin. For this purpose, the method known as the frequency-independence of loss tangent (tan δ) was adopted, by which the gel point could be determined from a multifrequency plot of tan δ versus the polysaccharide concentration. This method has been successfully used by Li and Aoki42 to determine the gel point of poly(vinyl chloride)/bis(2-ethylhexyl)phthalate systems. Figure 5 shows the tan δ values as a function of the native amylopectin concentration (Camyl) or La-amylopectin concentration (CLa-amyl) for two systems at various angular frequencies. All curves in each figure pass through the common point at the certain polysaccharide concentration, which is defined as the gel point (Cg). In this way, the Cg value could be determined to be 3.0% for aqueous amylopectin system and 0.6% for aqueous La-amylopectin system, respectively. It is clear

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Figure 5. Loss tangent (tan δ) as a function of polysaccharide concentration for aqueous systems of the native amylopectin and Laamylopectin at various angular frequencies (25 °C).Cg is the gel point.

Figure 7. Steady shear viscosity of aqueous native amylopectin (a) and La-amylopectin (b) systems with different concentrations (25 °C). Table 1. K and m Values as Well as the Corresponding Determination Coefficients (R) for Aqueous Amylopectin and La-Amylopectin Systems with Different Concentrations (25 °C) aqueous systems

conc. (%)

K

m

R

native amylopectin

3.0 5.0 7.0 9.0 3.0 5.0 7.0 9.0

6.080 ( 0.458 11.021 ( 0.777 16.838 ( 1.308 25.011 ( 2.785 19.717 ( 1.059 57.269 ( 1.446 151.586 ( 4.168 164.494 ( 4.317

0.281 ( 0.026 0.317 ( 0.025 0.356 ( 0.027 0.362 ( 0.038 0.151 ( 0.022 0.145 ( 0.010 0.020 ( 0.011 0.098 ( 0.011

0.985 0.985 0.980 0.961 0.994 0.999 0.999 0.999

La-amylopectin

Figure 6. Double-logarithmic plots of the relaxation modulus [G(t)] vs time (t) for aqueous amylopectin and La-amylopectin systems (25 °C).

that La-amylopectin has a strong gelation ability in aqueous system when compared with the native amylopectin. Durrani and Donald43 carried out the physical characterization of amylopectin hydrogels, and found that only aqueous amylopectin solutions of suficiently high concentration could form physically crosslinked thermoreversible hydrogels upon cooling to below room temperature. To understand the features of aqueous native amylopectin and La-amylopectin systems at the gel points, some stress relaxation experiments were performed. Figure 6 gives the double-logarithmic plots of the relaxation modulus (G(t)) versus time (t) for two systems. It was found that such plots gave good linear relationships with the determination coefficients of more than 0.990. This implies that the change of G(t) with t could be described well by the following power law relaxation mode44

G(t) ) Sgt-n

(5)

where Sg is the gel strength or stiffness, and n is the relaxation exponent. As a result, the Sg and n values were respectively determined to be 1.27 Pa · sn and 0.15 for aqueous native

amylopectin system (Camyl ) 3.0%) as well as 4.21 Pa · sn and 0.04 for aqueous La-amylopectin system (CLa-amyl ) 0.6%). In general, a greater value of Sg and a lower value of n imply the formation of a more highly elastic hydrogel.43,44 Figure 7 gives steady shear viscosity (η) as a function of shear rate for aqueous native amylopectin and La-amylopectin systems with different concentrations (25 °C). As seen, there is a shear-thining behavior for each system. All data can be represented satisfactorily by the following power-law equation45

η ) Kγm-1

(6)

where K is the consistency index and m is the flow behavior index. The K and m values obtained for each system as well as the corresponding determination coefficients (R) are listed in Table 1. For aqueous native amylopectin systems, the K and m values were observed to increase with the increase of the polysaccharide concentration. For aqueous La-amylopectin systems, the K value was found to increase obviously but the m value to decrease when the La-amylopectin concentration increased. A greater K value and a smaller m value of aqueous La-amylopectin system show the formation of a denser hydrogel network with a stronger shear-thining property, which is typical for associating and gel-forming polymer systems.46,47 This may be attributed to the intermolecular hydrophobic interactions in aqueous La-amylopectin system due to the incorporated lauryl



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Figure 8. Changes of the intensity ratio (I1/I3) with the polysaccharide concentrations for aqueous systems of (a) La-amylopectin system and (b) native amylopectin (25 °C).

chains. In this case, upon shearing the physically crosslinked network is disrupted into associative clusters and further to smaller fragments, which leads to the observed shear thinning behavior. For the detection of the hydrophobic clusters, fluorescence spectroscopy analyses were carried out for aqueous native amylopectin and La-amylopectin systems in the presence of pyrene as the fluorescent probe, as shown in Figure 8. It is known that the variation in the ratio (I1/I3) of intensity of the first (372 nm) to the third (383 nm) vibronic peaks, the socalled polarity parameter, is sensitive to the polarity of the microenvironment where pyrene is located.48,49 In the case of aqueous La-amylopectin system, the I1/I3 values remained nearly constant at lower concentrations. With a further increase in the concentration, the intensity ratio started to decrease, implying the formation of the hydrophobic clusters. The critical associating concentration (csc) could be determined to be 0.003 mg/ mL by the crossover point of two straight lines. However, few changes of I1/I3 with the concentration were observed for aqueous system of the native amylopectin. It seems that the gelation of the native amylopectin results only from the chain entanglements at higher concentrations while the gelation of La-amylopectin results from both the hydrophobic interactions and the chain entanglements. To obtain further insight into the structural charactristics of the La-amylopectin-based hydrogel and the native amylopectinbased hydrogel, we related the microscopic structure parameters of two kinds of hydrogels to their macroscopic elastic properties by means of strain sweep measurements and a fractal model developed by Shih et al.37 Figure 9 shows the storage modulus (G′)-strain profiles for two physical hydrogels, from which the maximum G′ values in the linear region and the critical strain values could be determined. As seen, the G′ remained almost constant when the strain increased and then suddenly decreased, which indicated the bond breakage within the gel network and a transition from linear to nonlinear behavior. Figure 10a shows the double-logarithmic plot of the critical strain (γ0) versus the polysaccharide concentration for each hydrogel. Because γ0 values tended to increase with the increase of the polysaccharide concentrations, these hydrogel samples could be confirmed to be weak-link hydrogels. Figure 10b shows the doublelogarithmic plots of the maximum elastic modulus (G0) versus the polysaccharide concentration for the same samples as those in Figure 10a. From the slopes of the plots, the fractal dimension Df was evaluated, using eq 3, to be 1.98 for the native amylopectin-based hydrogel and 2.35 for the La-amylopectinbased hydrogel. Compared with the native amylopectin-based

Figure 9. Storage modulus-strain profiles for (a) aqueous native amylopectin system and (b) aqueous La-amylopectin system (1.0 rad/ s, 25 °C).

Figure 10. The double-logarithmic plots of (a) the critical strain (γ0) vs the polysaccharide concentration and (b) the maximum elastic modulus (G0) vs the polysaccharide concentration for the native amylopectin- and La-amylopectin-based hydrogels.

hydrogel, the La-amylopectin-based hydrogel has a higher Df value. This suggests that a more complex network structure was formed in the La-amylopectin-based hydrogel, which was also confirmed by the SEM observation (Figure 11). As an effective polymeric hydrogelator, La-amylopectin was used for the in situ encapsulation and release of bovine serum albumin (BSA) as a model protein drug. When 1.0% Laamylopectin was added into aqueous BSA solutions with different concentrations, we found that the BSA-loaded hydrogels could be formed easily under the test temperature condition. Figure 12 shows the circular dichroism (CD) spectra of native BSA and the BSA encapsulated in the hydrogel. Compared with the CD spectrum of native BSA, the CD spectrum of the encapsulated BSA shows insignificant difference. This indicates

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Figure 11. SEM photos of (a) the native amylopectin-based hydrogel and (b) La-amylopectin-based hydrogel.

Figure 12. Circular dichroism spectra under various wavelengths for the native BSA and the BSA entrapped in the La-amylopectin-based hydrogel (PBS; pH, 7.4; 25 °C).

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Figure 14. Cumulative release of the encapsulated BSA from the hydrogel formed from 1.0% La-amylopectin in PBS (pH 7.4) at 37 °C.

there is a sustained release behavior in all cases. Before the hydrogels started to disintegrate, the BSA release was completed approximately within 4 h for the hydrogel loaded with 0.045% BSA, 8 h for the hydrogel loaded with 0.183% BSA and 10 h for the hydrogel loaded with 0.457% BSA, respectively. The reasons for the sustained BSA release might be as follows: (1) The diffusion of high molar mass BSA (Mw ) 67000 g · mol-1)51 was hindered by the hydrogel network; (2) there was an interaction between the hydrogel matrix and BSA due to the formation of hydrogen bonds. To understand the release machanism of the entrapped BSA from the La-amylopectinbased hydrogel, we fitted the accumulative release data using the following semiempirical equation52

Mt /M∞ ) ktn

Figure 13. Circular dichroism spectra under various temperatures for native BSA and the BSA entrapped in La-amylopectin-based hydrogel (wavelength, 222 nm; PBS; pH, 7.4).

that the entrapped BSA could keep its original structure and the encapsulation does not affect the protein secondary structure. To understand further the conformational stability of the protein, the CD spectra under various temperatures were also measured at 222 nm for the native BSA and the entrapped BSA, as shown in Figure 13. In the tested experimental conditions, the thermal denaturation of the native BSA in solution began at about 50 °C, while the CD spectrum of the entrapped BSA did not show any significant change. It is clear that the entrapped BSA is more stable than the native BSA in solution. The enhanced stability of the BSA after the encapsulation may due to the protective effect of the hydrogel matrix to the protein by the immobilization in the hydrogel network and the protein-matrix interaction, which impeded the thermal degradation and denaturation of the protein. Similar phenomenon was also observed by Teoli et al.50 when they entrapped BSA into wet sol-gel derived silica gels. For three La-amylopectin-based hydrogels loaded with various amounts of BSA during their formation, the in vitro release experiments after the immersion into physiological conditions were carried out. Figure 14 gives the cumulative BSA release against time for each BSA-loaded hydrogel sample. As seen,

(7)

where the k is the kinetic constant and the n is an exponent characterizing the diffusional mechanism, Mt and M∞ are the cumulative amount of the drug released at t and equilibium, respectively. Fickian diffusion (n ) 0.5) and case II transport (n ) 1) are often obtained when drugs are released from diffusion-controlled and swelling-controlled systems, respectively. A system controlled by both diffusion and swelling usually generates 0.5 < n < 1. In our work, the n value was determined to be 0.538 for the hydrogel loaded with 0.045% BSA (R ) 0.980), 0.644 for the hydrogel loaded with 0.183% BSA (R ) 0.980), and 0.852 for the hydrogel loaded with 0.457% BSA (R ) 0.994), respectively. Therefore, the release behavior of the entrapped BSA from the La-amylopectin-based hydrogel could be confirmed to be both diffusion- and swellingcontrolled kinetics.

Conclusions The grafting of lauryl side chains onto a hydrophilic amylopectin backbone was found to provide the amylopectin with strong gelation ability in aqueous system. The resultant amylopectin derivative hydrogelator could form rapidly a physical hydrogel under mild conditions by intermolecular hydrophobic interactions and macromolecular chain entanglements. For the obtained hydrogel matrix, its viscoelastic properties could be easily controlled by the concentration of this hydrogelator, and the gel point could be determined by the frequency independence of loss tangent in the vicinity of the sol-gel transition. By stress relaxation measurements, a power law time dependence of the relaxation modulus was detected at the gel point. In contrast to the physical hydrogel formed from aqueous amylopectin solutions of suficiently high concentration, the physical hydrogel based on this amphiphilic amylopectin derivative shows en-


hanced viscoelastic properties, gel strength, and shear thinning behavior. In particular, a more complex network structure with a higher fractal dimension was formed in such a hydrogel. As new matrix for the safe incorporation of bioactive molecules, the in situ crosslinkable hydrogel formed from this polysaccharide-based hydrogelator in aqueous solution holds a potential application for the sustained release of protein drugs. Acknowledgment. This work is supported by the Natural Science Foundation of China (Grant No. 20874116; 20676155) and the Natural Science Foundation of Guangdong Province in China (Grant No. 8151027501000004; 06023103; 039184). The authors appreciate the discussion with Professor Bing Xu from Brandeis University, U.S.A.

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