Journal of Colloid and Interface Science 283 (2005) 446–451

www.elsevier.com/locate/jcis

Preparation and adsorption properties of monodisperse chitosan-bound

Fe3O4 magnetic nanoparticles for removal of Cu(II) ions
Yang-Chuang Chang, Dong-Hwang Chen ∗
Department of Chemical Engineering, National Cheng Kung University, Tainan, Taiwan 701, Republic of China
Received 13 July 2004; accepted 9 September 2004

Abstract
Monodisperse chitosan-bound Fe3 O4 nanoparticles were developed as a novel magnetic nano-adsorbent for the removal of heavy metal
ions. Chitosan was first carboxymethylated and then covalently bound on the surface of Fe3 O4 nanoparticles via carbodiimide activation.
Transmission electron microscopy micrographs showed that the chitosan-bound Fe3 O4 nanoparticles were monodisperse and had a mean
diameter of 13.5 nm. X-ray diffraction patterns indicated that the magnetic nanoparticles were pure Fe3 O4 with a spinel structure, and the
binding of chitosan did not result in a phase change. The binding of chitosan was also demonstrated by the measurement of zeta potential,
and the weight percentage of chitosan bound to Fe3 O4 nanoparticles was estimated to be about 4.92 wt%. The chitosan-bound Fe3 O4
nanoparticles were shown to be quite efficient for the removal of Cu(II) ions at pH > 2. In particular, the adsorption rate was so fast that
the equilibrium was achieved within 1 min due to the absence of internal diffusion resistance. The adsorption data obeyed the Langmuir
equation with a maximum adsorption capacity of 21.5 mg g−1 and a Langmuir adsorption equilibrium constant of 0.0165 L mg−1 . The pH
and temperature effects revealed that the adsorption capacity increased significantly with increasing pH at pH 2–5, and the adsorption process
was exothermic in nature with an enthalpy change of −6.14 kJ mol−1 at 300–330 K.
 2004 Elsevier Inc. All rights reserved.

Keywords: Chitosan; Magnetic; Nanoparticles; Adsorbent; Cu(II)

1. Introduction as ionic exchange groups. It was shown to be quite fast

Most of commercialized or developing adsorbents are
submicrometer to micrometer sized and have large internal
porosities to ensure adequate surface area for adsorption.
However, the diffusion limitation within the particles leads
to decreases in the adsorption rate and available capacity.
Recently, the combination of organic and inorganic compo-
nents in a single particle at the nano-sized level has attracted
considerable attention because of the wide potential appli-
cations in many fields such as separation processes, opto-
electronics, catalysts and/or sensors, biotechnology, medical
diagnosis, and therapy [1–16]. Therefore, we developed
a novel magnetic nano-adsorbent which using iron oxide
(Fe3 O4 ) nanoparticles as cores and polyacrylic acid (PAA)
* Corresponding author. Fax: +886-6-2344496.
E-mail address: chendh@mail.ncku.edu.tw (D.-H. Chen).
0021-9797/$ – see front matter  2004 Elsevier Inc. All rights reserved.
doi:10.1016/j.jcis.2004.09.010
and efficient for the adsorption of enzyme and basic dyes
due to high specific surface area and the absence of internal
diffusion resistance [17–20]. Unfortunately, the PAA-based
magnetic nano-adsorbent is less effective for the adsorption
of heavy metal ions.
Chitosan is a natural polysaccharide with many useful
features such as hydrophilicity, biocompatibility, biodegrad-
ability, antibacterial properties, and remarkable affinity for
many biomacromolecules. It and its derivatives have great
potential applications in the areas of biotechnology, bio-
medicine, food ingredients, and cosmetics [21–27]. Note-
worthily, chitosan is also capable of adsorbing a number
of metal ions because its amino groups can serve as chela-
tion sites [28–30]. Thus, the binding of chitosan onto Fe3 O4
nanoparticles will probably yield another novel magnetic
nano-adsorbent for the efficient removal of heavy metal
ions.
 Y.-C. Chang, D.-H. Chen / Journal of Colloid and Interface Science 283 (2005) 446–451
The preparations of chitosan-conjugated Fe3 O4 particles
by a coprecipitation method, crosslinking method, and co-
valent binding method using coupling agents have been
reported [31]. The resultant magnetic composites were ei-
ther aggregated or unstable due to polymer crosslinking
or physisorption, the particle sizes were above 100 nm,
even larger than 1000 nm. So, following our previous work
on PAA-bound magnetic nanoparticles, the preparation of
monodisperse chitosan-bound Fe3 O4 nanoparticles was at-
tempted in this study. Since chitosan has no suitable func-
tional groups to bind directly onto Fe3 O4 nanoparticles, it
was first carboxmethylated and then covalently bound onto
Fe3 O4 nanoparticles via carbodiimide activation. The sizes,
structures, and surface charges of the product were charac-
terized by transmission electron microscopy (TEM), X-ray
diffraction (XRD), and zeta potential meter. The capability
and behavior for the adsorption of heavy metal ions were in-
vestigated using Cu(II) ions as a model compound.
2. Materials and methods
2.1. Materials
Chitosan (low viscous), chloroacetic acid sodium salt,
o-phthaldialdehyde, β-mercaptoethanol, and ferrous chlo-
ride tetrahydrate were purchased from Fluka (Buchs). Car-
bodiimides (cyanamide, CH2 N2 ) were supplied from Sigma
Chemical Co. (St. Louis, MO). Ferric chloride, 6-hydrate
was purchased from J.T. Baker (Phillipsburg). Glycine,
natriumtetraborate-10-hydrate, and Cu(II) nitrate-2,5-hy-
drate were obtained from Riedel-deHaën (Seelze). Ammo-
nium hydroxide (29.6%) was supplied by TEDIA (Fairfield).
The water used throughout this work was the reagent-grade
water produced by the Milli-Q SP ultrapure water purifica-
tion system of Nihon Millipore Ltd., Tokyo. All other chem-
icals were the guaranteed or analytic grade reagents com-
mercially available and used without further purification.
2.2. Preparation of chitosan-bound Fe3 O4 nanoparticles
Fe3 O4 nanoparticles were prepared by coprecipitating
Fe2+ and Fe3+ ions by ammonia solution and treating un-
der hydrothermal conditions [17–20]. Ferric and ferrous
chlorides (molar ratio 2:1) were dissolved in water at a
concentration of 0.3 M iron ions. Chemical precipitation
was achieved at 25 ◦ C under vigorous stirring by adding
NH4 OH solution (29.6%). During the reaction process, the
pH was maintained at about 10. The precipitates were heated
at 80 ◦ C for 30 min, washed several times with water and
ethanol, and then finally dried in a vacuum oven at 70 ◦ C.
Chitosan was carboxymethylated according to the method
of Chen and Park [32]. First, 3 g chitosan and 15 g sodium
hydroxide were added into 100 ml of isopropanol/water
(80/20) mixture at 60 ◦ C to swell and alkalize for 1 h. Then,
20 ml of monochloroacetic acid solution (0.75 g ml−1 in
447
isopropanol) was added into the reaction mixture in drops
in 30 min. After reaction for 4 h at the same temperature,
200 ml of ethyl alcohol (70%) was added to stop the reac-
tion. Finally, the solid was filtered, rinsed with 70 and 99%
ethyl alcohol to desalt and dewater, and dried in an oven
at 50 ◦ C.
The binding of carboxymethyl chitosan was conducted
following our previous work on the preparation of PAA-
bound magnetic nanoparticles [17]. First, 100 mg of Fe3 O4
nanoparticles was added to 2 ml of buffer A (0.003 M phos-
phate, pH 6, 0.1 M NaCl). Then, the reaction mixture was
sonicated for 10 min after adding 0.5 ml of carbodiimide
solution (0.025 g ml−1 in buffer A). Finally, 2.5 ml of car-
boxymethyl chitosan solution (50 mg ml−1 in buffer A) was
added and the reaction mixture was sonicated for 60 min.
The chitosan-bound Fe3 O4 nanoparticles were recovered
from the reaction mixture by placing the bottle on a per-
manent magnet with a surface magnetization of 6000 G.
The magnetic particles settled within 1–2 min and then were
washed with water and ethanol. NaCl was used for the
flocculation of magnetic nanoparticles. Its addition could
accelerate the magnetic separation, particularly in alkaline
solutions [33].
2.3. Characterizations of chitosan-bound
Fe3 O4 nanoparticles
The amino groups on the chitosan-bound Fe3 O4 magnetic
nanoparticles were determined by spectrophotometric as-
say based on the reaction of o-phthaldialdehyde (OPA) and
β-mercaptoethanol (BME) with primary amines [17,34].
First, two reagents were freshly prepared as follows. For re-
action of primary amines of the solid, 300 µl ethanol solution
containing 0.25 M OPA and 300 µl of 0.1 M sodium borate
(pH 9.5) containing 4% β-mercaptoethanol were added to
water to yield 50 ml of reagent 1 (R1). For the determina-
tion of excess OPA, 500 µl of 0.06 M glycine and 500 µl
of 0.1 M sodium borate (pH 9.5) containing 0.5% BME
were added to water to yield 30 ml of reagent 2 (R2). Sec-
ondly, 20 mg of magnetic nanoparticles was incubated with
5 ml reagent R1 for 5 min at 25 ◦ C. The liquid solution
was then separated from the magnetic nanoparticles via a
permanent magnet. To 4 ml of 0.1 M sodium borate and
200 µl of reagent R2 in a test tube, 100 µl of supernatant
was added. After mixing by vortex for several minutes, the
absorbance of the sample at 340 nm was determined on a
Hitachi U-3000 spectrophotometer. Finally, a reference so-
lution was prepared according to the above procedures but in
the absence of magnetic nanoparticles. The concentration of
amino groups on the surface of magnetic nanoparticles was
determined from the difference between the absorbances of
reference solution and sample, using a molar absorption co-
efficient ε = 6623 M−1 cm−1 obtained from the calibration
curve.
The size and morphology of magnetic nanoparticles
were observed by TEM using a JEOL Model JEM-1200EX
448 Y.-C. Chang, D.-H. Chen / Journal of Colloid and Interface Science 283 (2005) 446–451
at 80 kV. The sample for TEM analysis was obtained
by placing a drop of the magnetic nanoparticle-dispersed
aqueous solution onto a Formvar-covered copper grid and
evaporated in air at room temperature. Before the sample
was withdrawn, the dispersed solution was sonicated for
1 min to obtain the better particle dispersion on the cop-
per grid. XRD measurement was performed on a Rigaku
D/max III. V X-ray diffractometer using CuKα radiation
(λ = 0.1542 nm). The zeta potentials of the naked and
chitosan-bound Fe3 O4 nanoparticles were measured on a
Malvern ZEN2600 Zetasizer Nano Z.
2.4. Adsorption studies
The adsorption of Cu(II) ions by the chitosan-bound
Fe3 O4 magnetic nanoparticles (as a magnetic nano-adsor-
bent) was investigated in aqueous solutions at pH 2–5 and
300–330 K. In general, 105.17 mg magnetic nano-adsorbent
(i.e., 5.17 mg chitosan on 100 mg Fe3 O4 ) was added to
5 ml of copper nitrate solution. After mixing by vortex for
1 min, the magnetic nano-adsorbent was removed magnet-
ically from the solution. The solution pH was adjusted by
NaOH or HCl. The concentrations of Cu(II) ions were mea-
sured using a GBC Avanta atomic absorption spectrometer.
The concentration of magnetic nano-adsorbent was fixed at
21.034 mg ml−1 . Unless otherwise specified, the absorption
experiments were performed in aqueous solution at pH 5
and 300 K.
3. Results and discussion
3.1. Particle size and structure of chitosan-bound
Fe3 O4 nanoparticles
The typical TEM micrographs for the naked and chitosan-
bound Fe3 O4 nanoparticles are shown in Fig. 1. It was clear
that both the naked and the chitosan-bound Fe3 O4 nanopar-
ticles were essentially monodisperse and had a similar mean
Fig. 1. TEM micrographs for the naked (a) and chitosan-bound (b) Fe3 O4
nanoparticles.
Fig. 2. XRD patterns for the naked (a) and chitosan-bound (b) Fe3 O4 nano-
particles.
diameter of 13.5 nm. This reveals that the binding process
did not significantly result in the agglomeration and the
change in size of the particles. This could be attributed to
the reaction occurring only on the particle surface, and thus
our attempt to prepare monodisperse chitosan-bound Fe3 O4
nanoparticles in this work has been achieved.
Fig. 2 shows the XRD patterns for the naked and
chitosan-bound Fe3 O4 nanoparticles. Six characteristic peaks
for Fe3 O4 (2θ = 30.1, 35.5, 43.1, 53.4, 57.0, and 62.6◦ ),
marked by their indices ((220), (311), (400), (422), (511),
and (440)), were observed for both samples. These peaks
are consistent with the database in JCPDS file (PCPDFWIN
v.2.02, PDF No. 85-1436) and reveal that the resultant
nanoparticles were pure Fe3 O4 with a spinel structure. Also,
the binding process did not result in the phase change of
Fe3 O4 . It is explained that the binding process did not result
in the phase change of Fe3 O4 .
3.2. Amount of chitosan bound on Fe3 O4 nanoparticles
According to the increased weight after chitosan binding,
the amount of chitosan bound on Fe3 O4 nanoparticles could
be estimated to be 5.25 mg per 100 mg Fe3 O4 . To further
confirm this binding amount, the amino groups of chitosan
on Fe3 O4 nanoparticles were quantified by the spectrophoto-
metric assay based on the reaction of amino groups of solids
with an excess of OPA and subsequent quantitative deter-
mination of unreacted OPA by reaction with glycine. It was
found that about 0.0233 mmol of amino groups per 100 mg
Fe3 O4 nanoparticles was available. Accordingly, the quan-
tity of carboxymethyl chitosan on Fe3 O4 nanoparticles could
be estimated to be 5.08 mg per 100 mg Fe3 O4 . This was in
agreement with that obtained from weight difference. The
average value 5.17 mg (i.e., 4.92 wt%) was adopted in the
following investigations.
3.3. Zeta-potential analysis
The zeta potentials of the naked and chitosan-bound
Fe3 O4 nanoparticles (0.1 mg ml−1 ) were measured in 10−3 M
NaCl aqueous solutions at pH 2.5–9.5 (adjusted by NaOH
or HCl). As shown in Fig. 3, the isoelectric point (pI ) of the
naked Fe3 O4 nanoparticles was about 6.7, consistent with
 Y.-C. Chang, D.-H. Chen / Journal of Colloid and Interface Science 283 (2005) 446–451
Fig. 3. Zeta potentials of the naked (P) and chitosan-bound (!) Fe3 O4
nanoparticles at different pH.
Fig. 4. Dependence of adsorption capacity on contact time.
the literature value of 6.5 [35]. After chitosan bonding, the
pI was shifted to 5.95. This also confirms the binding of
chitosan and reveals that the chitosan-bound Fe3 O4 nanopar-
ticles were positively charged at pH < 5.95.
3.4. Adsorption properties
The feasibility of chitosan-bound Fe3 O4 nanoparticles
as a magnetic nano-adsorbent for the removal of heavy
metal ions from aqueous solutions was demonstrated using
Cu(II) ions as a model compound. First, the time required
to achieve adsorption equilibrium was determined. The typ-
ical experiments were conducted at pH 5, 300 K, and an
initial Cu(II) ion concentration of 1100 mg L−1 . As shown
in Fig. 4, the adsorption capacities (q) for the contact time
of 1 to 60 min had no significant differences. This reveals
that the adsorption equilibrium was reached within 1 min.
Such a fast adsorption rate could be attributed to the absence
of internal diffusion resistance, like the PAA-based magnetic
nano-adsorbent developed in our previous work.
The effect of pH on the adsorption of Cu(II) ions by
the magnetic nano-adsorbent at 300 K and the initial Cu(II)
ion concentrations of 200 and 1150 mg L−1 is illustrated in
Fig. 5. It was found that the adsorption capacity increased
with increasing the solution pH at an initial Cu(II) ion con-
centration of 1150 mg L−1 . This might be due to the less
insignificant competitive adsorption of hydrogen ions. At a
lower initial Cu(II) ion concentration (200 mg L−1 ), the ad-
sorption capacity increased with increasing the solution pH
and then remained unchanged at pH 3–5. This might be due
to the fact that almost all Cu(II) ions were adsorbed. At
449
Fig. 5. Effect of pH on the adsorption of Cu(II) ions by magnetic nano-
adsorbent. Initial Cu(II) ion concentration: 200 (!) and 1150 mg L−1 (P).
Fig. 6. Equilibrium isotherm for the adsorption of Cu(II) ions on magnetic
nano-adsorbent at pH 5 and 300 K. The inset illustrates the linear depen-
dence of Ce /q on Ce .
pH > 5, white Cu(OH)2 precipitate formed and so no ad-
sorption experiments were performed. Noteworthly, almost
no Cu(II) ions were adsorbed at pH 2. This condition might
be utilized for the desorption of Cu(II) ions from the mag-
netic nano-adsorbent.
The equilibrium isotherm for the adsorption of Cu(II)
ions on magnetic nano-adsorbent at pH 5 and 300 K is shown
in Fig 6. The adsorption data were analyzed according to the
linear form of the Langmuir isotherm as follow [36],
Ce 1 Ce
= + , (1)
q Kqm qm
where q is the adsorption capacity (mg g−1 ) based on the
dry weight of nano-adsorbent, Ce is the equilibrium concen-
tration (mg L−1 ) in solution, qm is the maximum adsorption
capacity (mg g−1 ), and K is the Langmuir adsorption equi-
librium constant (L mg−1 ). As shown in the inset in Fig. 6,
the plot of Ce /q vs Ce at various temperatures yielded
straight lines, revealing that the adsorption of Cu(II) ions
on magnetic nano-adsorbent obeys the Langmuir adsorption
isotherm. From the slope and intercept, the values of qm and
K might be estimated as 21.5 mg g−1 and 0.0165 L mg−1 ,
respectively. It was notable that only 4.92 wt% of chitosan
was responsible for the adsorption of Cu(II) ion. The max-
imum adsorption capacity based on the weight of chitosan
was 437 mg g−1 , which was significantly higher than the
value 284 mg g−1 observed for the chitosan-coated silica
beads by Shi et al. [37] and those values below 200 mg g−1
usually observed for the chitosan beads [30,38]. This might
be reasonably referred to the high specific surface area be-
450 Y.-C. Chang, D.-H. Chen / Journal of Colloid and Interface Science 283 (2005) 446–451
Fig. 7. Effect of temperature on the adsorption of Cu(II) ions by magnetic
nano-adsorbent. The inset is the plot of ln(q/Ce ) against 1/T .
cause the bound chitosan might be spread on the surface
of Fe3 O4 nanoparticles, leading to almost all chelation sites
available.
The effect of temperature on the adsorption of Cu(II)
ions by the magnetic nano-adsorbent was investigated at
pH 5, 300–330 K, and an initial Cu(II) ion concentration
of 1100 mg L−1 . As shown in Fig. 7, the adsorption ca-
pacity of Cu(II) ions decreased with increasing temperature.
The might be due to the fact that the electrostatic interac-
tion between Cu(II) ions and chitosan was lower at higher
temperatures. The plot of ln(q/Ce ) vs 1/T is shown in the
inset in Fig. 7. From the slope (−H /R), the change of
enthalpy (H ) at 300–330 K could be determined to be
−6.14 kJ mol−1 , indicating that the adsorption process was
exothermic in nature.
Accordingly, the chitosan-based magnetic nano-adsorbent
developed in this work was monodisperse and quite efficient
for the fast adsorption of Cu(II) ions due to high specific
surface area and the absence of internal diffusion resistance.
Compared to the PAA-based magnetic nano-adsorbent early
developed, the chitosan-based magnetic nano-adsorbent was
more efficient for the adsorption of heavy metal ions. Since
chitosan has wide applications in the areas of biotechnology
and biomedicine, the novel magnetic nano-adsorbent will
also be useful in the related fields as the magnetic carriers of
enzymes, proteins, DNA, drugs, etc.
4. Conclusions
A novel magnetic nano-adsorbent was fabricated by the
covalent binding of carboxymethylated chitosan on Fe3 O4
nanoparticles via carbodiimide activation. The analyses of
TEM and XRD indicated that the chitosan-bound Fe3 O4
nanoparticles were monodisperse with a mean diameter of
13.5 nm, and the binding of chitosan did not change the
spinel structure of Fe3 O4 . From the increased weight after
chitosan binding and the OPA method, the amount of chi-
tosan bound on Fe3 O4 nanoparticles was estimated to be
about 4.92 wt%. The binding of chitosan was also found
to result in the shift of isoelectric point from 6.7 to 5.95.
The chitosan-bound Fe3 O4 nanoparticles were quite effi-
cient as a magnetic nano-adsorbent for the fast adsorption
of Cu(II) ions from aqueous solutions at pH 2–5 due to
high specific surface area and the absence of internal diffu-
sion resistance. The time required to achieve the adsorption
equilibrium was only 1 min. Almost no Cu(II) ions were ad-
sorbed at pH 2, but the adsorption capacity was significantly
enhanced with the increase in pH. The adsorption behavior
followed the Langmuir adsorption isotherm with a maxi-
mum adsorption capacity of 21.5 mg g−1 and a Langmuir
adsorption equilibrium constant of 0.0165 L mg−1 at 300 K.
Also, the adsorption process was exothermic in nature with
an enthalpy change of −6.14 kJ mol−1 at 300–330 K. This
work will be helpful for the development of adsorbents and
magnetic carriers.
Acknowledgment
This work was performed under the auspices of the Na-
tional Science Council of the Republic of China, under
Contract NSC 93-2214-E006-023, to which the authors ex-
press their thanks.
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