Entrapment of white-rot fungus Trametes versicolor in

Ca-alginate beads: preparation and biosorption kinetic

analysis for cadmium removal from an aqueous solution
M. Yakup Arca
a,
*
, Yasemin Kac ar
a
,

Omer Genc
b
a
Department of Biology, Krkkale University, 71450 Yahs ihan, Krkkale, Turkey
b
Department of Chemistry, Hacettepe University, 06532 Beytepe, Ankara, Turkey
Received 22 January 2001; received in revised form 4 April 2001; accepted 26 April 2001
Abstract
The biosorption of cadmium ions onto entrapped Trametes versicolor mycelia has been studied in a batch system. The maximum
experimental biosorption capacities for entrapped live and dead fungal mycelia of T. versicolor were found as 102:3 3:2 mg Cd(II)
g
1
and 120:6 3:8 mg Cd(II) g
1
, respectively. Biosorption equilibrium was established in about 1 h and biosorption was well
described by the Langmuir and Freundlich biosorption isotherms. The change in the biosorption capacity with time was found to t
the pseudo-second-order equation. Since the biosorption capacities were relatively high for both entrapped live and dead forms,
those fungal forms could be considered as suitable biosorbents for the removal of cadmium in wastewater-treatment systems. The
biosorbents were reused in three consecutive adsorption/desorption cycles without a signicant loss in the biosorption
capacity. 2001 Elsevier Science Ltd. All rights reserved.
Keywords: Cadmium; Heavy metal; Ca-alginate; Entrapment; Biosorption; Trametes versicolor
1. Introduction
Pollution by cadmium usually comes from several
industrial processes such as electroplating, plastics
manufacturing, nickelcadmium batteries, fertilizers,
pigments, mining and metallurgical processes (Kefala
et al., 1999; Kaewsarn and Yu, 2001; Park et al., 1999).
Several chemical processes have been utilized for re-
moval of cadmium from cadmium-contaminated in-
dustrial wastewater, but it is dicult to apply chemical
processes to purication of cadmium contaminated soils
and water because they need enormous amounts of
chemicals, and lead to secondary pollution (Kapoor
et al., 1999; Gadd, 1993). Common treatments to re-
move cadmium ion from contaminated water are mostly
based on precipitation, evaporation and adsorption on
ion exchange resins (Kapoor et al., 1999; Gadd, 1993;
Kasan, 1993). The present technologies are expensive
when cadmium is present in the wastewater at a low
concentration or when a very low concentration of
cadmium in treated water is required. In order to de-
velop economic, practical and ecient techniques, fun-
gal and other microbial biomass have been studied to
remove and recover metallic elements such as cadmium,
mercury, lead, copper etc. (Volesky and Holan, 1995).
Biosorption utilizes the ability of biological materials to
accumulate heavy metals from wastewater by either
bioaccumulation, or biosorption. The several biological
materials that have been investigated for cadmium re-
moval include fungi, bacteria, yeast and algae.
Application of fungal biomass to remove heavy
metals from industrial wastewater and/or to recover
economically valuable metals is attractive for industry.
Many fungal species such as Rhizopus arrhizus (Aksu
et al., 1999), Phanerochaete chrysosporium (Saglam et
al., 1999; Say et al., 2001). Mucor miehei (Guibal et al.,
1992) and Aspergillus niger (Kapoor et al., 1999) have
been extensively studied for heavy metal biosorption
and the process mechanisms seem to be dependent upon
species. Trametes versicolor is a well-known white-rot
fungus and a strong degrader of various xenobiotics. It
could also be used to remove cadmium ions from
Bioresource Technology 80 (2001) 121129
*
Corresponding author. Tel.: +90-318-357-2477; fax: +90-318-357-
2329.
E-mail addresses: arica@turkuaz.kku.edu.tr, info@kgm.gov.tr (M.
Yakup Arca).
0960-8524/01/$ - see front matter 2001 Elsevier Science Ltd. All rights reserved.
PII: S 0 9 6 0 - 8 5 2 4 ( 0 1 ) 0 0 0 8 4 - 0
wastewaters by adsorbing the cadmium on its mycelium
(Zhou and Ki, 1991).
In industrial or technical operations, immobilized
microbial cell systems could also provide additional
advantages over freely suspended cells. These include
ease of regeneration and reuse of the biomass, easier
solidliquid separation and minimal clogging in con-
tinuous-ow systems (Ting and Sun, 2000; Arca et al.,
1993). Natural polymers such as alginate, chitosan,
chitin, and cellulose derivatives have been mostly used
as the matrix for the immobilization of microbial cells
via the entrapment technique. These polymers are also
known to bind metal ions strongly. Entrapment of mi-
crobial cells in these polymer supports could also en-
hance microbial cell performance and adsorptive
capacity of the biosorbent system for the heavy metal
ions. Immobilized fungal cells were found to be far more
stable during continuous operation in the bioreactor
than the fungal cells in the free form (Crist et al., 1994;
Sag et al., 1995; Jianlong et al., 2000).
In this work, T. versicolor was entrapped using Na-
alginate as the natural polymeric matrix. After uniform
growth on the surface of the matrix, the entrapped dead
and live fungal cells were used for biosorption of Cd(II)
ions from aqueous solutions in a batch system. The ef-
fect of pH on biosorption capacities of both entrapped
(dead and live) forms was characterized by measuring
the adsorption of Cd(II) ions at dierent pH values. The
maximum adsorption capacities of both entrapped
preparations, based on dry weight, were determined by
varying the concentration of the heavy metal ions in the
aqueous solutions. The biosorption kinetics of cadmium
removal and the relevance of biosorption isotherms for
characterization of uptake were examined. The eec-
tiveness of desorbing agent (HCl) in stripping adsorbed
cadmium ions from the alginate beads and the en-
trapped fungal preparations was also investigated.
2. Methods
2.1. Micro-organism and biosorbent preparation
White-rot basidiomycete, T. versicolor was main-
tained by subculturing on malt dextrose agar slants.
Growth medium consisted of (in g l
1
of distilled water);
D D-glucose, 10.0; KH
2
PO
4
, 20.0; MgSO
4
7H
2
O, 0.5;
NH
4
Cl, 0.1; CaCl
2
H
2
O, 0.1; thiamin, 0.001; and it was
supplemented with 1.0 ml of trace element solution. The
pH of the medium was adjusted to 4.5. Inocula were
obtained from a seven-day old agar slant culture. The
fungal mat (0.3 g) was removed and macerated asepti-
cally in 5.0 ml sterile medium using a homogenizer. It
was used to inoculate 50 ml of medium in a 250 ml ask,
and the asks were incubated on a shaker (150 rpm) for
5 days at 30C. After this period, the biomass was
harvested by ltration from the growth medium and
washed several times with distilled water. The fungal
biomass was obtained as discrete spherical clumps; it
was homogenized with a commercial blender to destroy
cell aggregates.
The entrapment of T. versicolor in Ca-alginate was
carried out as follows: Na-alginate (2.0 g; from Mac-
rosytia pyrifera, high viscosity, Sigma Chem., USA) was
dissolved in distilled water and then mixed with the
fungal mycelium (50 ml, containing fungal biomass 0.5
g). The mixture was introduced into a solution con-
taining 0:1 M CaCl
2
with a burette and the solution
was stirred to prevent aggregation of the fungus-en-
trapped Ca-alginate beads. The fungus-entrapped beads
(~4 mm) were cured in this solution for 1 h and then
washed twice with 200 ml sterile distilled water. The
beads with entrapped mycelia were then transferred to
the growth medium (50 ml) in 250 ml ask and were
incubated on an orbital shaker (150 rpm) at 30C for 3
days. The mycelia growth in/on the beads was followed
during the incubation period by using a microscope.
After a three-day incubation period, the Ca-alginate
beads with entrapped fungal mycelia were removed
from the medium by ltration and washed twice with
distilled water. This washed biomass is hereafter called
``entrapped live fungus''. At times entrapped live fungus
was heated in 5 mM CaCl
2
solution at 90C for 10 min
and it will be referred to as ``entrapped dead fungus''.
The entrapped preparations were then stored at 4C
until use in 5 mM CaCl
2
solution. The dry weight of the
microbial growth in/on the entrapped preparations was
determined by weighing (after drying in an oven at 50C
overnight) the alginate beads before and after cell
growth.
2.2. Biosorption procedures
The biosorption of Cd(II) ions, on the plain alginate
beads and on both entrapped live and dead T. versicolor
from water was investigated in batch biosorption-equi-
librium experiments. The eects of the medium pH and
the initial concentration of Cd(II) ions on the biosorp-
tion rate and capacity were studied.
The eect of pH on the biosorption rate of the algi-
nate beads and both entrapped fungal preparations with
Cd(II) ions was investigated in the pH range of 3.07.0
(which was adjusted with HCl or NaOH at the begin-
ning of the experiment and not controlled afterwards) at
25C. Cd(II) ion in each solution (200 mg l
1
) was
prepared in distilled water (25 ml) and alginate beads,
entrapped live or dead fungus preparation was trans-
ferred into this medium and agitated magnetically at 400
rpm.
The eect of the initial Cd(II) ions concentration on
the biosorption was studied at pH 6.0 as described
earlier except that the concentration of Cd(II) ions in the
122 M.Y. Arca et al. / Bioresource Technology 80 (2001) 121129
adsorption medium was varied between 30 and
600 mg l
1
.
2.3. Analytical procedure
Biosorption of Cd(II) ions from aqueous solutions
was studied in batch systems. Nitrate of cadmium ions
was used. After the desired incubation period (about 120
min) the aqueous phases were separated from the bio-
sorbents and the concentrations of the metal ions in
these phase were measured by using an atomic absorp-
tion spectrophotometer ((AAS), Shimadzu AA 6800,
Japan). Deuterium background correction was used and
the spectral slit width was 0.5 nm. The working current/
wavelength for Cd(II) ions was 8.0 mA/228.8 nm.
The instrument response was periodically checked
with known Cd(II) solution standards. For each set of
data, standard statistical methods were used to deter-
mine the mean values and S.D.s. Condence intervals of
95% were calculated for each set of samples in order to
determine the margin of error.
The amount of metal ions adsorbed per unit on the
plain and fungus immobilized alginate preparations (mg
metal ions g dry beads
1
) was obtained by using the
following expression:
q
eqex
= [(C
0
C)V [=M; (1)
where q
eqex
is the amount of metal ions adsorbed onto
the unit amount of the biosorbents (mg g
1
). C
0
and C
are the concentrations of the metal ions in the initial
solution (mg l
1
) and after biosorption, respectively. V
is the volume of the aqueous phase (l
1
) and M is the
amount of the biosorbent (g).
A known quantity of wet Ca-alginate or both fungus-
entrapped preparations was used in the biosorption test.
After biosorption process, the biosorbents were dried in
an oven at 50C overnight and the dry weight of the
preparations was used in the above equation. Each ex-
periment was repeated three times and the results given
are the average values.
2.4. Biosorption/desorption
In order to determine the reusability of the alginate
beads and both entrapped fungal preparations, consec-
utive biosorptiondesorption cycles were repeated three
times by using the same biosorbents. Desorption of
Cd(II) ions was performed by 10 mM HCl solution. The
alginate and both entrapped dead and live forms loaded
with Cd(II) ions were placed in the desorption medium
and stirred at 400 rpm for 60 min at 25C. The nal
Cd(II) ion concentration in the aqueous phase was de-
termined by using an AAS. The desorption ratio was
calculated from the amount of metal ions adsorbed on
the entrapped preparations and the nal Cd(II) ion
concentration in the adsorption medium.
Desorption ratio was calculated from the following
equation:
Desorption ratio =
amount of Cd(II) ions desorbed
amount of Cd(II) ions biosorbed
:
(2)
Percent desorption values were obtained by multiplying
the above equation by 100.
2.5. Scanning electron microscopy (SEM) studies
Sample of Ca-alginate and fungus-entrapped beads
were coated under vacuum with a thin layer of gold and
examined by SEM (JEOL, Model JMS 5600).
3. Kinetic models of adsorption
3.1. Equilibrium equations
Adsorption processes on the Ca-alginate and on the
fungus-entrapped systems (live or dead form) was
modeled by applying the Langmuir and the Freundlich
adsorption isotherms
ML
k
1
k
2
ML (3)
Metal ions and ligands on the biosorbents are de-
noted by M and L, respectively. k
1
and k
2
are the for-
ward and reverse interaction rates, respectively, which
include the metal ion movement from the bulk phase to
the biosorbent surface layer.
Eq. (3) can be expressed in the form of a rate equa-
tion with a second-order forward and a rst-order re-
verse kinetics, namely
dq=dt = k
1
C
eq
(q
m
q
eq
) k
2
q
eq
: (4)
Eq. (4) leads to
q
eq
= (q
m
C
eq
)=(k
d
C
eq
); (5)
where C
eq
and q
eq
also show the residual metal concen-
tration and the amount of metal adsorbed on the ad-
sorbent at equilibrium, respectively, k
d
= k
2
=k
1
is the
Langmuir constant of the system.
The semi-reciprocal plot of C
eq
=q
eq
vs C
eq
was em-
ployed to generate the intercept of k
d
=q
m
and the slope
of 1=q
m
.
The empirical Freundlich equation based on the
amount of a substance adsorbed (q
eq
) is related to the
concentration C
eq
by the equation
q
eq
= K
F
(C
eq
)
1=n
; (6)
where K
F
and n are the Freundlich constants charac-
teristic of the system. K
F
and n are indicators of ad-
sorption capacity and adsorption intensity, respectively.
M.Y. Arca et al. / Bioresource Technology 80 (2001) 121129 123
The slope and intercept of the linear Freundlich equa-
tion are equal to 1=n and ln K
F
, respectively.
3.2. Pseudo-rst- and second-order equations
In order to examine the controlling mechanism of
biosorption process such as mass transfer and chemical
reaction, kinetic models were used to test the experi-
mental data. The large number and dierent chemical
groups on the cell wall of the fungal mycelia (e.g.,
ACOOH, ANH
2
, @NH, ASH, AOH) imply that there
are many types of fungal myceliametal ion interactions.
The kinetic models (the pseudo-rst-order and pseudo-
second-order equations) can be used in this case as-
suming that measured concentrations are equal to cell
surface concentrations. The rst-order rate equation of
Lagergren is one of the most widely used for the sorp-
tion of solute from a liquid solution (Lagergren, 1898;
Ho and McKay, 1999; Cheung et al., 2001). It may be
represented as follows:
dq
t
=dt = k
1
(q
eq
q
t
); (7)
where k
1
is the rate constant of pseudo-rst-order bio-
sorption (min
1
) and q
eq
and q
t
denote the amounts of
biosorption at equilibrium and at time t (mg g
1
), re-
spectively. After integration by applying boundary
conditions, q
t
= 0 at t = 0 and q
t
= q
t
at t = t, gives
log(q
eq
=q
eq
q
t
) = (k
1
t)=2:303: (8)
Eq. (8) can be rearranged to obtain a linear form
log(q
eq
q
t
) = log q
eq
(k
1
t)=2:303: (9)
A plot of log(q
eq
q
t
) against t should give a straight
line to conrm the applicability of the kinetic model. In
a true rst-order process log q
eq
should be equal to the
intercept of a plot of log(q
eq
q
t
) against t.
In addition, a pseudo-second-order equation based
on adsorption equilibrium capacity may be expressed in
the form:
dq
t
=dt = k
2
(q
eq
q
t
)
2
; (10)
where k
2
(g mg
1
min
1
) is the rate constant of pseudo-
second-order adsorption. Integrating Eq. (10) and ap-
plying the boundary conditions, leads to
(1=(q
eq
q
t
)) = (1=q
eq
) k
2
t (11)
or equivalently for a linear form
(t=q
t
) = (1=k
2
q
2
eq
) (1=q
eq
)t: (12)
A plot of t=q
t
vs t (Eq. 12) should give a linear rela-
tionship for the applicability of the second-order kinet-
ics. The rate constant (k
2
) and adsorption at equilibrium
(q
eq
) can be obtained from the intercept and slope, re-
spectively.
4. Results and Discussion
4.1. Properties of biosorbent systems
The entrapment of T. versicolor into Ca-alginate
beads was carried out by a liquid curing method in the
presence of Ca(II) ions and was used in the removal of
Cd(II) from aqueous medium. Heavy metal removal by
ion-exchange resins is sensitive to the presence of Ca
2
,
Mg
2
and K

ions, whereas the fungal biomass does not

adsorb ions such as Ca
2
, Mg
2
and K

(Tobin et al.,
1994). Thus, the use of entrapped fungus in Ca-alginate
beads may be advantageous over the ion-exchange
resins when Ca
2
, Mg
2
and K

ions are present in an

adsorption medium or industrial wastewater at high
concentrations.
Alginate is a natural polymer and can easily be con-
verted into hydrogels via cross-linking with divalent
calcium ions. It was preferred over other materials be-
cause of its various advantages such as biodegradability,
hydrophilicity, presence of carboxylic groups, and nat-
ural origin. Other additional advantages are its low
density and mechanical stability that makes it highly
suitable for many biotechnological applications. The
alginate beads were very stable over the experimental
pH range of 3.08.0. The operational stability of the
support under specied experimental conditions is also a
very important parameter in the cell entrapment. One of
the most important disadvantages of cell entrapment is
the increase in the mass transfer resistance due to the
polymeric matrix. From this point of view, entrapped
cell matrix containing alginate could also be advanta-
geous when compared with other support materials such
as, agar, carrageenan, polyvinyl alcohol and 2-hy-
droxyethylmethacrylate, because the presence of carb-
oxylic groups in the alginate structure enhances the
heavy metal ion adsorption capacity of the system with a
combination of microbial cells.
Representative SEM micrographs for the plain Ca-
alginate and fungus entrapped bead surface are pre-
sented in Figs. 1(a) and (b), respectively. The SEM
micrograph of fungus-entrapped alginate bead was
found to be completely dierent from the empty one
and showed a uniform fungal growth on the bead sur-
face indicating that entrapped fungal mycelium was not
localized. This uniform distribution is an important
criterion for the proper biosorption of heavy metal ions
on the whole surface area of the fungus entrapped
beads. Thus, entrapment of fungal cells in/on the algi-
nate beads could also provide additional advantages
over the freely suspended fungal cell, in a batch-culture
fungal mycelia form in individually distributed spherical
clumps ( 24 mm). This tight packing of the fungal
cells could also lead to diusional restriction and less
adsorptive sites for heavy metals than the alginate
fungal cell systems.
124 M.Y. Arca et al. / Bioresource Technology 80 (2001) 121129
4.2. Eect of pH on the biosorption
It is well-known that metal ion adsorption on both
non-specic and specic sorbents is pH dependent
(Kapoor et al., 1999; Matheickal et al., 1999; Ting and
Sun, 2000). The medium pH aects the solubility of
metal ions and the ionization state of the functional
groups (i.e., carboxylic, phosphate, and amino groups)
on the fungal cell wall (Fourest and Volesky, 1997). Fig.
2 shows the eect of pH on biosorption. Biosorption of
Cd(II) ions rst increased with pH, and maximum
Cd(II) ions biosorption occurred between pH 5.0 and
6.0 and the interaction of the Cd(II) ions with the algi-
nate and entrapped fungal cell wall component could be
primarily with the carboxyl groups in both alginate and
the cell wall component of the mycelia. The metal bio-
sorption hence depends on the protonation or depro-
tonation of these carboxyl groups, and it is likely that
the groups responsible for metal are carboxyl groups,
which have pK
a
between 3 and 4 (Fourest and Volesky,
1997). The biosorption of Cd(II) increased with pH upto
6.0 and then declined with further increase in pH
(Fig. 2). As the pH increased, more carboxyl groups on
the cell wall component deprotonated and carried neg-
ative charges. This means that negative charge attracts
positively charged cadmium ions, and more cadmium
binding occurs. The decrease of the biosorption for pH
higher than 6.0 could be due to the formation of a
cadmium hydroxide complex which would prevent the
metal biosorption. It should be noted that the entrapped
dead fungal biomass, exhibited higher cadmium remo-
vals than the live form at all the tested pH values.
Similar observations were reported for other biomass
types (Fourest and Roux, 1992; Brady et al., 1994;
Kefala et al., 1999).
4.3. Biosorption rates
Fig. 3 shows the changes in the amount of Cd(II) ions
biosorbed with time. High biosorption rates were ob-
served at the beginning, and then plateau values (i.e.,
adsorption equilibrium) are gradually reached within 60
min. For example, the biosorption equilibrium time of
chromium (IV) on the dead and immobilized biomass of
R. arrhizus was 2 h (Prakasham et al., 1999). The lead
biosorption rate on P. chrysosporium is fast and reached
Fig. 1. A representative SEM micrograph for the plain Ca-alginate (a) and fungus-entrapped alginate beads (b).
M.Y. Arca et al. / Bioresource Technology 80 (2001) 121129 125
saturation value within 2 h (Yetis et al., 2000). The lead
biosorption equilibriumtime on A. niger biomass was 5 h
(Kapoor et al., 1999). The biosorption of cadmium onto
pre-treated biomass of the marine alga Durvillaea pota-
torum has been studied and the biosorption process was
very fast with 90% uptake taking place within 30 min
(Matheickal et al., 1999). Note that there are several
parameters which determine the biosorption rate, such
as stirring rate of the aqueous phase, structural prop-
erties both of the support and the biosorbent (e.g.,
protein and carbohydrate composition and surface
charge density, topography and surface area), amount
of sorbent, properties of ion under study (e.g., ionic
radius), initial concentration of ionic species and, of
course, existence of other metal ions which may compete
with the ionic species of interest for the active biosorp-
tion sites. The biosorption rates reported were dierent
because of these causes.
4.4. Eect of initial metal ion concentration
Cadmium ion biosorption capacities of the Ca-algi-
nate and both entrapped dead and live form are pre-
sented as a function of the initial concentration of Cd(II)
ions within the aqueous biosorption medium in Fig. 4.
This gure was prepared by using the plateau values of
the biosorption rate-curves (given in Fig. 3). The
amount of Cd(II) ions adsorbed per unit mass of the
biosorbent (i.e., biosorption capacity) increased with
the initial concentration of metal ions. In order to ob-
tain the maximum biosorption capacity of the Ca-algi-
nate and both entrapped live and dead form for the
Fig. 4. Biosorption capacities of plain Ca-alginate and entrapped T.
versicolor (dead and live form) for Cd(II) ions. biosorption conditions:
amount of biosorbent: 25 mg; volume of the biosorption medium: 25
ml; pH: 5.5; temperature: 25C.
Fig. 2. Eect of pH on the biosorption capacities of the Ca-alginate,
and entrapped T. versicolor (dead and live form) for Cd(II) ions.
Biosorption conditions: initial concentration of Cd(II) ions: 200 mg
l
1
; amount of biosorbent 25 mg; volume of biosorption medium: 25
ml; temperature: 25C.
Fig. 3. Biosorption rates of Cd(II) ions by plain Ca-alginate and en-
trapped fungus T. versicolor (dead and live form). Biosorption condi-
tions: initial concentration of Cd(II) ions: 400 mg l
1
; pH: 5.5;
temperature: 25C.
126 M.Y. Arca et al. / Bioresource Technology 80 (2001) 121129
Cd(II) ions, the initial concentration of Cd(II) ions was
increased up to 600 mg l
1
. As seen in Fig. 4, the amount
of Cd(II) ions adsorbed on the plain alginate beads was
39:1 1:45 mg per g dry alginate beads. Maximum
biosorption capacity for entrapped live and dead fungal
mycelia of T. versicolor was found as 102:3 3:2 mg
Cd(II) g
1
and 120:6 3:8 mg Cd(II) g
1
, respectively.
The biosorption capacity of the entrapped dead T. ver-
sicolor for cadmium was about 20% higher than that of
the entrapped live counterpart. Thus, it was reasonable
to assume that increased cadmium biosorption capacity
was due to the changes in biosorptive characteristics of
the fungus as a result of heat treatment. From these
observations, it appeared that the surface properties of
the fungal cells could be improved upon application of
heat. The heat treatment could erode microbial cell
surface integrity causing the walls to become leaky with
a marked increase in the passive diusion of metal ion to
the interior part of the cell wall (Churchill et al., 1995).
A wide range of biosorption capacities for heavy
metal ions by dierent biosorbents have been reported.
The biosorption capacity of heat-treated fungus P. Sa-
jor-caju was 29.6 mg Cd(II) g
1
dry biomass (Cihangir
and Saglam, 1999). The biosorption capacity of dead
Fusarium occiferum mycelium was 19.2 mg Cd(II) g
1
dry biomass (Delgado et al., 1998). The pre-treated alga
D. Potatorum has been used for cadmium removal and
the biosorption capacity of the biosorbent was 114.4 mg
g
1
dry biomass (Matheickal et al., 1999). In the case of
fungal biomass of the white-rot fungus P. chrysosporium
used for heavy metal removal from articial wastewater,
the cadmium removal capacity of the dry fungal bio-
mass was 23.4 mg Cd(II) g
1
(Say et al., 2001). The
adsorption capacity of 120:6 3:8 mg Cd(II) g
1
bio-
sorbent, obtained with entrapped dead T. versicolor, was
comparable with the values reported in the previous
studies.
4.5. Equilibrium isotherm
Fig. 5 shows the Langmuir plots for Cd(II) biosorp-
tion by Ca-alginate and both entrapped fungus live and
dead forms. The Langmuir constants (q
m
and k
d
) along
with correlation coecients (R
2
) have been calculated
from the plots (Fig. 5) for biosorption of Cd(II) on the
biosorbents and the results are presented in Table 1. The
maximum capacity q
m
determined from the Langmuir
isotherm denes the total capacity of the biosorbents for
Cd(II) ions.
Fig. 6 shows the Freundlich plots for Cd(II) ad-
sorption by Ca-alginate and both entrapped live and
dead form. The magnitude of K
F
and n values showed
easy uptake of Cd(II) from aqueous medium with a high
adsorption capacity of entrapped live and dead fungus.
With all experimentally tested biosorbents, n values were
found high enough for separation (Table 1). The ex-
perimental equilibrium data tted both the models well
(Table 1).
4.6. Biosorption kinetics modeling
In order to analyze the biosorption kinetics of Cd(II)
ions, the pseudo-rst-order and the pseudo-second-
order kinetic models were applied to the data. The
pseudo-second-order equation tted well with the
experimental data. The comparison of experimental
biosorption capacities and the theoretical values
estimated from the above two equations are presented in
Table 2. The theoretical q
eq
values estimated from the
rst-order kinetic model gave signicantly dierent
Fig. 5. Langmuir plot of Cd(II) ions on the Ca-alginate and entrapped
T. versicolor (dead and live forms).
Table 1
Isotherm model constants and correlation coecients for biosorption of Cd(II) ions from an aqueous solution
Biosorbent Langmuir Freundlich
q
m
(mg g
1
) k
d
M (10
7
) R
2
K
F
N R
2
Ca-alginate 40.1 22.1 0.996 5.5 3.09 0.999
Live T. versicolor 141.3 7.2 0.996 3.5 1.68 0.986
Dead T. versicolor 164.8 6.4 0.996 6.6 2.96 0.965
M.Y. Arca et al. / Bioresource Technology 80 (2001) 121129 127
values compared to experimental values, and the cor-
relation coecients were also found to be slightly lower
(Fig. 7). These results showed that these biosorption
systems were not described by the rst-order kinetic
model.
The correlation coecients for the linear plots of t=q
t
against t for the second-order equation are greater than
0.996 for all the biosorbents for contact times of 120 min
(Fig. 8). The theoretical q
eq
values for all the tested
biosorbent systems were very close to the experimental
q
eq
values in the case of second-order kinetics. The sec-
ond-order kinetics best described the data.
4.7. Desorption and reuse
The desorption of the adsorbed Cd(II) ions from the
biosorbents was studied in a batch system. The Cd(II)
ions adsorbed onto biosorbents were eluted with 10 mM
HCl. More than 95% of the adsorbed Cd(II) ions was
desorbed from the biosorbents. In order to examine the
reusability of the biosorbents, adsorptiondesorption
cycle of Cd(II) ions was repeated three times by using
the same preparations. The adsorption capacities for all
the biosorbents did not noticeably change (only a
maximum 3% change was observed with the tested
biosorbent) during the repeated adsorptiondesorption
operations. These results showed that alginate beads
and both fungus-entrapped biosorbents can be repeat-
edly used in heavy metal adsorption studies without
detectable losses in their initial adsorption capacities.
5. Conclusions
The plain alginate beads and both entrapped live and
dead forms have been successfully used as the biosorb-
ing agent for removal of Cd(II) ions from an aqueous
medium. The mechanism and the kinetics of Cd(II) ions
biosorption on the biosorbents depended on the exper-
imental conditions particularly medium pH and Cd(II)
ions concentration. The equilibrium was well described
Table 2
The rst-order and second-order kinetic constants for biosorption of Cd(II) ions on the alginate and both immobilized live and dead white-rot fungi
T. versicolor
Biosorbent Experimental First-order kinetic Second-order kinetic
q
eqex
(mg g
1
)
k
1
10
2
(min
1
)
q
eq
(mg g
1
)
R
2
k
2
10
3
(g mg
1
min
1
)
q
eq
(mg g
1
)
R
2
Ca-alginate 39.1 1.24 40.1 0.998 7.73 44.6 0.998
Live T. versicolor 102.3 0.97 95.6 0.992 2.74 103.4 0.996
Dead T. versicolor 120.6 1.22 102.8 0.980 1.70 116.1 0.996
Fig. 6. Freundlich plot of Cd(II) ions on the Ca-alginate and en-
trapped T. versicolor (dead and live forms).
Fig. 7. (log q
eq
q
t
) vs t plot for Cd(II) ions on the Ca-alginate and
entrapped T. versicolor (dead and live form).
128 M.Y. Arca et al. / Bioresource Technology 80 (2001) 121129
by the Langmuir and Freundlich adsorption isotherms.
For all the fungus-entrapment systems studied, the
biosorption of Cd(II) ions on the biosorbents followed
the pseudo-second-order biosorption kinetics. The fun-
gus, T. versicolor, entrapped in live or dead form could
be regenerated using 10 mM HCl with up to 97% re-
covery. The biosorbents were reused in three biosorp-
tion and desorption cycles with negligible decrease in the
biosorption capacities.
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M.Y. Arca et al. / Bioresource Technology 80 (2001) 121129 129