of

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different sizes. The kinetics parameters Kdye and Vdye max for the decolorization process for all the seven dyes were estimated through

Microbial decolorization processes offer a complete

cleanup of pollutants in a natural way as it reduces the color

decolorizing common dyes as these strains produce

peroxidases that oxidize the various structure of the dyes

peroxide and oxalates produced by the fungus catalyzes the

oxidative degradation of the pollutants [16]. These extra

Process Biochemistry 40 (200cellular peroxidases, are non-specific towards the substrate so

that it can attack some recalcitrant chemicals of diverse

structures, including organic-pollutants. The lignolytic* Corresponding author. Tel.: +91 44 2220 3507; fax: +91 44 2235 2642.

E-mail address: tmgesan_57@yahoo.com (T. Murugesan).

1359-5113/$ see front matter # 2005 Elsevier Ltd. All rights reserved.doi:10.1016/j.procbio.2005.03.033noticeable at a dye concentration of more than 1 mg/l and

an average concentration of 300 mg/l have been reported in

effluents from textile manufacturing processes [1,2]. Many

authors have reported physico-chemical treatments for the

removal of color from industrial wastewaters [35]. Even

though these procedures prove to be efficient, the operational

costs are relatively high and leads to other disadvantages like

sludge formation, biomass accumulation, etc. [6].

chrysosporium have been studied in the field of decolorization

of industrial effluents. This is due to the versatile ability of the

fungus to degrade, partially or completely various dyes such

as heterocyclic, azo, anthraquinone, vat and polymeric dyes

[1114]. P. chrysosporium displayed color reduction abilities

for all such dyes including the dyes used in newsprint, writing

and printing paper industries [15]. Extracellular peroxidases

such as lignin peroxidases, manganese peroxidases, hydrogenrelease of the dyes into the environment are the textile and

dyestuff manufacturing industries. Normally colors areand are found to be more suitable for decolorization [810].

The white rot fungi, more specifically, strains of P.variety of dyes in these industries, pollution from the effluents

has become increasingly alarming. The two major sources of# 2005 Elsevier Ltd. All rights reserved.

Keywords: Phanerochaete chrysosporium; Decolorization; Dyes; Immobilization; Calcium alginate beads; Kinetics

1. Introduction

Syntheticdyesareusedextensively for textiledyeing,paper

printing, leather dyeing, color photography and as additives in

petroleum products. With the increasing usage of the wide

components to carbon dioxide, ammonia and water by

initiating cleavage of the bonds in the dyes rather than

creating possible toxic fragments of dyes [7]. Strains such as

Phanerochaete chrysosporium, Trametes versicolor, Pseu-

domonas luteola have been reported to be suitable forLineweaverBurk plots.Decolorization studies

Phanerochaete chrysos

K.V. Radha, I. Regupathi,

Department of Chemical Engineering, AC College

Received 19 July 2004; received in revised

Abstract

Treatment of effluents from dye-based industries poses a major p

In this study, Phanerochaete chrysosporium, a commonly used wh

structures, namely azo, anthraquinone, thiazine and vat dyes. The de

violet, Congo red, Acid orange, Acid red 114, Vat magenta, Methy

parameters, namely dyes concentration (20400 mg/l), pH (27), te

percentage decolorization were investigated. Studies were carried osynthetic dyes using

ium and their kinetics

runagiri, T. Murugesan *

hnology, Anna University, Chennai 600025, India

2 February 2005; accepted 26 March 2005

m and biotreatment with white rot fungi seems to be a viable option.

t fungus, was used to biodegrade several synthetic dyes of varying

ization potential of P. chrysosporium for seven dyes namely, Methyl

lue and Acid green was studied. The effect of various operational

ature (2045 8C) and inoculum size (0.254 ml) on the maximuming free cells and fungal cell entrapped calcium alginate beads of

www.elsevier.com/locate/procbio

5) 33373345

iochem2. Materials and methods

2.1. Microorganism

The white rot fungus P. chrysosporium MTCC 787 was

obtained from the Culture Collection of Institute of

Microbial Technology, Chandigarh, India and the stock

cultures were maintained by periodic subculture on malt

agar medium at 4 8C.

2.2. Inoculum

The fungus P. chrysosporiumwas inoculated on malt agar

and incubated at 35 8C until extensive spore growth occurred.The basal medium [20] used to study the fungal biomass and

decolorization test consists of: D-glucose, 5.0 g/l; KH2PO4,

2.0 g/l; NH4Cl, 0.050 g/l; MgSO47H2O, 0.5 g/l;CaCl22H2O, 0.1 g/l; thiamine HCl, 100 mg; trace elementsolution, 10 ml and the final pH of the medium was

maintained at pH 4.5. Trace element solution consisting of

MnSO4, 0.5 g/l; FeSO47H2O, 0.1 g/l; ZnSO47H2O, 0.1 g/lwas prepared separately and 10 ml was added to the medium.

2.3. Dyes and decolorization studies

Seven commercial dyes belonging to the groups of azo

(Acid orange, Acid red 114), triphenylmethane (Methylenzymes of P. chrysosporium (due to their oxidative

mechanism) are considered responsible for the aerobic

decolorization of the dyes that not only decolorizes but also

detoxifies the effluents completely [17].

Recently, the application of immobilized cells has been

receiving increasing attention in the field of wastewater

decolorization. Many researchers have studied the effect of

immobilized whole cells and enzymes on decolorization

characteristics, since immobilization provides distinct

stability over free cells [12,18]. In this present work,

experiments on decolorization of the dyes were carried out

in batch mode using P. chrysosporium (MTCC 787) free

cells, to study the decolorization of seven structurally

different dyes and hence to find the optimum conditions viz.,

initial concentrations of the dyes, initial pH, glucose and

nitrogen concentrations, inoculum concentrations and

temperatures for the design of continuous reactors for

decolorization. Attempt have also been made for whole cells

immobilization using calcium alginate entrapment, due to

the gentle gelation procedure compared to that of chemical

polymerization procedures [19]. Similar to that of free cells,

studies were also carried out using immobilized beads of

varying bead sizes. The data obtained have been used to

study the kinetic effects on decolorization of individual dyes

since the kinetics studies are rarely reported with respect to

dyes decolorized by P. chrysosporium.

K.V. Radha et al. / Process B3338violet), anthraquinone (Acid green), thiazine (Methyleneblue), Vat (Vat magenta) and diazo group (Congo red) were

used. Individual dyes were added to Erlenmeyer flasks

(250 ml) containing 100 ml of the medium, which was

inoculated with approximately 3.2 105 cells. The experi-ments were carried out in an orbit shaker at 60 rpm for 7 days

at 35 8C. The dye concentrations were measured usingsamples collected at regular intervals using a spectro-

photometer [21] [UV/VIS shimadzu spectrophotometer

(model U2000)]. Control experiments for each test were

carried out using uninoculated medium with dye addition.

2.4. Enzyme assays

Enzyme activities were determined spectrophotometri-

cally at 35 8C lignin peroxidase (LiP) activity wasdetermined by the oxidation of veratryl alcohol at 310 nm

as described by Tien and Kirk [22]. Manganese peroxidase

activity was assayed at 468 nm using dimethoxyphenol as

the substrate as suggested by Field et al. [23]. Laccase

activity was analyzed spectrophotometrically according to

Niku-Paavola et al. [24], with 2,20-azino-di(3-ethyl-benzo-thiazolin-sulphonate) (ABTS) as substrate. One unit was

defined as the amount of enzyme that oxidized l mmol ofsubstrate per minute and the activities are reported as U/l.

2.5. Biomass determination

To determine biomass growth, a fixed volume of culture

broth was centrifuged at 1000 rpm for 45 min. The pellet

was removed, washed, filtered through a predried, pre-

weighed filter paper. The filter paper was dried to a constant

weight and the dry weight of the biomass was determined as

gram per litre [25].

2.6. Biosorption studies

In order to study the role of mycelium in dye

decolorization, the mycelium and the supernatant were

separated when the fungus showedmaximum activity. To the

mycelium the dye was added and tested for biosorption.

Enzyme activities were monitored during biosorption.

2.7. Immobilization

P. chrysosporium were grown into the stationary phase in

malt agar slants. Spore suspension of 2 ml (approximately

3.2 105 cells) was added to 100 ml of 2% sodium alginate.The mixture was gently stirred at room temperature to

produce a uniform suspension and then dropped into 100 ml

of 20% calcium chloride solution. Five different nozzles

were used to form beads of uniform sizes (2, 3, 4, 5 and

6 mm). The beads so obtained were stored in calcium

chloride solution at 4 8C for 2 h to complete gel formation[26]. The insoluble and stable immobilized P. chrysospor-

ium alginate beads thus obtained were further used for the

istry 40 (2005) 33373345decolorization studies.

3. Results and discussion

3.1. Effect on initial concentrations of the dyes

P. chrysosporium was used to study the percentage

decolorization alongwith the maximum time requirements

for decolorization process. For initial experiments, keeping

the parameters such as initial pH and temperature as

constant, initial concentrations of the individual dyes were

varied from 0.02 to 0.4 g/l.

The maximum time taken for decolorization varies with

the nature of individual dyes and the longer time taken for

decolorization is a result of the production of extra cellular

orange, whereas Vat magenta, Methylene blue, Congo red

and Acid red 114 showed 8892% of decolorization.

Decolorization was far less for Acid green, which showed

only 75%. Fig. 2a and b show the maximal percentage

decolorization of the individual dyes and it is evident that P.

chrysosporium shows the potential to transform the dyes to

colorless substances.

Compared to all the dyes used in the present study

(Fig. 2a), Methyl violet had a high percentage of

decolorization due to the sequential demethylation with

the removal of penta, tetra and trimethyl groups [31]. The

tentativemetabolic pathways of methyl violet decolorization

by different species are explained by Sarnaik and Kanekar

[32]. As concluded by Chizuko et al. [33], the presence of

hydroxyl group in the para position of the aromatic ring

leads to a faster cleavage of the bond by the organisms. This

could be the reason for the fast decolorization as the Acid

orange has a hydroxyl group in the para position. The

percentage decolorization of methyl violet upto an initial

concentrations of 0.2 g/l are at maximum and nearly uniform

(Fig. 2a), whereas for concentrations greater than 0.2 g/l, a

sudden drop in percentage decolorization was observed.

This may contribute to the fact that the fungus showed high

sensitivity and low tolerance to the dye [34]. For the case of

all other dyes a gradual decrease in the percentage

decolorization with respect to the initial concentration is

observed.

K.V. Radha et al. / Process Biochemistry 40 (2005) 33373345 3339peroxidases, which are available only after 23 days of the

growth of the fungus [27]. The enzyme production slowly

increased after 8 h of growth of the fungus and reached a

maximum of 234 U/l for LiP and 172 U/l for MnP by 120 h.

Thereafter, a drop in activity was observed (Fig. 1). Laccase

was tested for its activity from the day 1 but there was no

noticeable quantity of laccase production till the day 5, after

which 14 U/l was observed on day 7, further there was no

increase. From these results, it was observed that LiP and

MnP were the key enzymes responsible for the decoloriza-

tion process. A similar trend was also observed by Sami and

Radhouane [28], using different medium for the production

of enzymes using the fungus P. chrysosporium. They have

also concluded that decolorization starts on the second day

and reaches a maximum during the fourth day but higher

activity of the enzyme was reported on the fifth day after

which it declined.

Acid orange and Vat magenta took less than 2 days for

decolorization which might be due to the fact that dyes acts

as suitable substrate for the peroxidases and oxidases

produced by the fungus [23,29]. Even though the same

amount of the inoculum was used for all the tested dyes, the

differences found in the decolorization characteristics for

the individual dyes are attributed to the dissimilarity in

specificities and structures of different dyes. Similar strains

of white rot fungi also decolorize the dyes in the same

manner like that of P. chrysosporium [30]. About 98% of

decolorization is achieved for Methyl violet and Acid

Fig. 1. Enzymes production during the growth of the fungus Phanerochaetechrysosporium.Fig. 2. Effect of initial concentration of the dyes on the percentage

decolorization (initial pH:4.5; T:35 8C; initial concentration (g/l): (a)(^) 0.05; (&) 0.1; (~) 0.2; () 0.3; ( ) 0.4) and (b) (^) 0.02; (&)

0.04; (~) 0.05; () 0.06; ( ) 0.08; (*) 0.1).

The growth of the P. chrysosporium and the correspond-

ing decolorization process were essentially controlled by the

pH of the medium. The percentage decolorization of the

dyes, using free cells of P. chrysosporium at various initial

iochemistry 40 (2005) 33373345In the present study, it is observed that P. Chrysosporium

showed lesser activity towards decolorization of Acid green

than other dyes tested (Fig. 2b). The organism was not able

to decolorize Acid green at a concentration greater than

0.08 g/l even though toleration in the medium was little

higher. The dye is expected to become toxic to the

microorganism at higher concentrations (>0.08 g/l), as aresult, an incomplete decolorization is observed [35]. Hence,

the present studies were made upto a concentration of

0.08 g/l only for Acid green, whereas for other dyes P.

chrysosporium was able to grow at higher concentrations

and an appreciable decolorization was observed.

Congo red and Acid red 114 showed a maximum

decolorization of upto 90% for an initial dye concentration

of 0.02 g/l. Even though there was an identical percentage

decolorization for an initial concentration of 0.02 g/l in

these two dyes, at higher concentrations, P. chrysosporium

proved to be more effective for the case of Congo red than

Acid red 114. A similar observation were also made for

Acid green and Methylene blue. As it is observed in all

these cases, beyond an optimum initial concentration of

dyes the rate of decolorization decreases and further

increase in concentration does not have any effect on

decolorization. This could be attributed to the fact that the

color removal depends on the destruction of the

chromophore. The peroxidases of the fungus needs to

attack one molecule of the dyes several times, a lower

concentration of the dye facilitate the destruction of the

molecules and the higher the concentration of the dyes the

slower the rate of color removal [36]. No general trend was

seen regarding the behaviour of dyes of different nature

(azo, anthraquinone, triphenylmethane, etc.) but the

structure of individual dye seems to an have influence

on the decolorization. The relevant factors that influence

the decolorization process are: (i) the structure of the dyes,

(ii) loss of the vital requirements by the organism and (iii)

might require an additional amount of veratryl alcohol

apart from the production by the microorganism itself

[37].

3.2. Biomass growth

To evaluate the effect of toxicity of the dyes on the

growth of P. Chrysosporium batch studies were conducted

with varying dye concentrations (0.020.1 g/l). A sample

graph is given for Congo red as the biomass growth was

found to have a similar pattern for all the dyes (Fig. 3). The

maximum biomass concentration decreased from 1.35 g/l

in the control to 1.25 and 0.75 g/l for the initial dye

concentration of 0.02 and 0.1 g/l, respectively. At an initial

dye concentration of 0.3 g/l there was complete inhibition

of growth. Further experiments were carried out under the

same conditions to produce biomass that is suitable for the

production of enzymes. The agitation was kept at 60 rpm

with the addition of Tween 80 in a nitrogen-limited

K.V. Radha et al. / Process B3340medium [38].3.3. Biosorption studies

The role of mycelium in dye decolorization was

investigated by separating the mycelium and the supernatant

at the time at which the fungus showed maximum activity.

To this 0.05 g/l of Methyl violet was added and the time

course of the enzyme activity and dye decolorization was

studied and the results are shown in Fig. 4. The

decolorization was limited to 39% by physical adsorption

of the mycelium, as the enzyme activity was very low after

separation from the supernatent.

The experimental results clearly indicate that the mycelia

individually cannot produce significant decolorization [39].

As the fungal peroxidases appears to be extracellular

enzymes the maximum activity decreased to 58 U/l from the

initial value of 234 U/l for LiP and 36 U/l from 172 U/l for

MnP. Therefore, both the mycelium (biosorption) and the

extra cellular fungal enzyme (biodegradation) are necessary

for the dye decolorization.

3.4. Effect of pH and temperature

Fig. 3. Growth of biomass during decolorization process.Fig. 4. Biosorption of the mycelium on decolorization.

pH conditions ranging from 2.0 to 7.0 were studied (Fig. 5).

Maximum decolorization for all the dyes studied in this

work was observed at a pH range of 4.05.0 and the

percentage decolorization decreased at both extremes of pH

(5.0). Although the dyes were decolorized at apH range of 4.05.0, Methylene blue, Acid green, Congo red

and Vat magenta were mostly decolorized at around a pH

5.0. On the other hand, the maximum decolorization for

Acid orange, Methyl violet and Acid red 114 occurs at pH

K.V. Radha et al. / Process Biochemistry 40 (2005) 33373345 3341

Fig. 5. Effect of initial pH of the dyes on the percentage decolorization

(initial concentration: Methyl violet, Acid orange, Vat magenta: 0. 05 g/l;

Acid red 114, Acid Green, Congo red, Methylene blue: 0. 02 g/l; T: 35 8C;initial pH: (^) Methyl violet; (&) Acid orange; (~) Vat magenta; () Acidgreen; ( ) Acid red 114; (*) Congo red; (j) Methylene blue).

Fig. 7. Effect of initial pH of the dye methyl violet on the percentage

decolorization using immobilized beads (initial concentration: 0.05 g/l; T:

35 8C; diameter of the bead: 2 mm; initial pH: (^) 2; (&) 3; (~) 4; ()4.5;( ) 5; (*) 6; (j) 7).4.04.5 and 4.05.0, respectively, above and below which

the decolorization process decreased remarkably. TheseFig. 6. Variation in pH during decolorization process (T: 35 8C; initialconcentration (0.05 g/l): (^) Methylene blue; (&) Methyl Violet; (~) Vatmagenta; () Congo red; ( ) LiP; (*) MnP) (T: 35 8C; initial concentration(0.02 g/l): (^) Acid orange; (&) Acid green; (~) Acid red 114; ( ) LiP; (j)MnP).observations indicate that the optimum pH for the fungus P.

chrysosporium depends on the nature of the substrate used as

observed and reported by Alberto et al. [40].

The variation in pH during the course of the decoloriza-

tion using free cells are shown in Fig. 6. It is observed that,

for the case of Methyl violet, Vat magenta, Congo red and

Methylene blue even though there was a slight variation of

pH during the process of decolorization, the final pH was

maintained at 4.05.0. But for the case of Acid orange, Acid

green and Acid red 114, the final pH shoots upto 4.55.0

even though the initial pH was 2.5 (Acid green and Acid

orange) and 3 (Acid red 114). The decrease in initial pH did

not have any effect on decolorization [41]. As observed by

Dirk et al. and Ana et al. [42,43] the fungus produces organic

acids such as malonate, oxalate during the initial growth

period, which later decomposed by the enzyme (manganese

peroxidase). The fungus was biologically active during this

period.

The experiments were also carried out to study the

effect of initial pH on the percentage decolorization using

calcium alginate beads, since the medium pH is expected

to affect the ionization state of the functional groups on the

fungal cell walls and the entrapped fungus (carboxylic,

phosphate and amino groups). For example, Fig. 7 shows

the trend observed for Methyl violet, which is similar toFig. 8. Effect of glucose on dye decolorization (initial concentration of

Methyl violet: 0.05 g/l; T: 35 8C; concentration of glucose (g/l): (^) 1; (&)2; (~) 2.5; () 5; ( ) 10; (*) 15).

K.V. Radha et al. / Process Biochemistry 40 (2005) 333733453342

Fig. 10. Effect of inoculum size on dye decolorization (initial concentrationthat of free cells. This may be possibly due to the same

functional group that gets reacted in both alginate and the

cell wall component of the mycelia [26].

In order to study the variation in temperature on

decolorization studies were carried out at temperatures

ranging from 20 to 45 8C. At higher (>35 8C) or lower(

of immobilized are reported by Kuo-Cheng et al. [52].

Nurdan et al. [53] have observed higher degrading activity in

P. chrysosporium on immobilization. The reason for lower

Kdye compared to that of free cells could be attributed to the

fact that the calcium alginate used for entrapment might act

as a barrier for immediate dissociation of the dyes. As

iochem

3 (mm) 0.6550 0.0083

4 (mm) 0.6026 0.0076

5 (mm) 0.5771 0.0075

6 (mm) 0.5496 0.0070

Acid orange

Free cells 0.6884 0.0078

2 (mm) 0.8926 0.0076

3 (mm) 0.8335 0.0072

4 (mm) 0.8073 0.0071

5 (mm) 0.8162 0.0068

6 (mm) 0.8014 0.0068

Vat magenta

Free cells 0.9097 0.0246

2 (mm) 0.8898 0.0224

3 (mm) 0.8898 0.0224

4 (mm) 0.9806 0.0248

5 (mm) 0.9036 0.0217

6 (mm) 0.7777 0.0219

Congo red

Free cells 0.3029 0.0042

2 (mm) 0.2525 0.0035

3 (mm) 0.2460 0.0034

4 (mm) 0.2279 0.0032

5 (mm) 0.2090 0.0031

6 (mm) 0.2026 0.0029

Acid green

Free cells 0.4011 0.0035

2 (mm) 0.2529 0.0029

3 (mm) 0.2418 0.0027

4 (mm) 0.2385 0.0027

5 (mm) 0.2359 0.0027

6 (mm) 0.2197 0.0025

Methylene blue

Free cells 0.1328 0.0021

2 (mm) 0.1115 0.0020

3 (mm) 0.1078 0.0020

4 (mm) 0.1065 0.0020

5 (mm) 0.1009 0.0018

6 (mm) 0.1001 0.0018

Acid red 114

Free cells 0.1101 0.0022

2 (mm) 0.1362 0.0021

3 (mm) 0.1441 0.0021

4 (mm) 0.1383 0.0019

5 (mm) 0.1385 0.0018

6 (mm) 0.1433 0.00183.7. Studies on immobilized beads

Decolorization experiments with an estimated optimum

concentration (0.05 g/l for Methyl violet, Acid Orange and

Vat magenta and 0.02 g/l for Acid green, Acid red 114,

Congo Red and Methylene blue) were carried out with

immobilized calcium alginate beads of different sizes (2

6 mm). The maximum percentage decolorizations for

different sizes of beads are reported in Table 1. The

percentage decolorization decreased with increasing bead

diameter for all the dyes tested. The reason could be

attributed to the increase in the surface area in the smaller

beads compared to beads with larger diameters [19,48].

Basic experiments with alginate beads (without immobili-

zation) showed an initial reduction of 20% of the color,

which is due to the absorption by the alginate beads and the

rest is being decolorized by P. chrysosporium and the time

taken for decolorization was nearly the same compared to

that of free cells.

3.8. Kinetic studies

The dye decolorization process is mainly an extra cellular

enzymatic process, hence studies have been made to find the

kinetics of the decolorization. For the purpose of establish-

ing the kinetic parameters for the decolorization process

LineweaverBurk plots were used.

1

V Km

Vmax

1

s 1Vmax

The estimated values of Kdye and Vdye max are tabulated in

Table 2 for free cells and immobilized cells. For the case of

free cells the results shows the Kdye values in the range of

0.10.6 g/l for all the dyes tested except for Methyl violet

and Vat magenta which showed a Kdye value of 0.9304 and

0.9097 g/l, respectively. As Kdye is a measure of the enzyme

substrate complex, a high Kdye indicates a weak binding

[49]. For the case of Methyl violet the reason could be due to

the instant dissociation of the dye with the free cells as

explained by Bumpus and Brock [31]. The value of Kdye and

Vdye max for the decolorization of Azo dye (reactive red 22)

by P. luteolawas found to be 0.156 g/l and 0.012 g/l/h) [50],

respectively, which is in good agreement with the present

range of results obtained using P. chrysosporium.

Similar experiments were carried out for immobilized

beads of diameter ranging from 2 to 6 mm to study the effect

on surface area. The entrapped P. chrysosporium in alginate

beads showed a low Kdye value for the dyes Methyl violet,

Congo red and Acid green and almost constant for Vat

magenta, Methylene blue and Acid red 114. For the case of

Acid orange the Kdye values are found to be quite high. The

variation in Kdye value might be due to the surface effects of

the immobilized cells and thereby affinity of the hydro-

phobic substrates either increases or decreases [51]. Similar

K.V. Radha et al. / Process Bresults of change in Kdye values from that of free cells to thatistry 40 (2005) 33373345 3343

Table 2

Estimated kinetic parameters using Phanerochaete chrysosporium free cells

and immobilized beads

Dyes Kdye (g/l) Vdye max (g/l h)

Methyl violet

Free cells 0.9327 0.0131

2 (mm) 0.8743 0.0112observed from the results (Table 2) Vdye max value is lesser

ml) was obtained. Similar experiments were carried out for

entrapped cells in alginate beads in batch mode using shake

[5] Sangkil N, Paul GT. Reduction of azo dyes with zero valent iron.Water

Res 2000;34(6):183745.

K.V. Radha et al. / Process Biochemistry 40 (2005) 333733453344[6] Mishra G, Tripathy M. A critical review of the treatment for decolor-

ization of textile effluent. Colourage 1993;358.

[7] Paszczynski A, Pasti-Grigsby MB, Goszczynski S, Crawford RL,

Crawford DL. Mineralization of sulfonated azo dyes and sulfanilic

acid by Phanerochaete chrysosporium and Streptomyces chromfuscus.

Appl Environ Microbiol Biotechnol 1992;58:3598604.

[8] MarkWP, Yitzhak H, Illan C. The decolorization of the polymeric dye

poly-blue (poly vinalamine sulphonate-anthraquinone) by ligninAcknowledgement

The authors wish to express their appreciation to Anna

University for the award of Research fellowship to K.V.

Radha for support of this investigation.

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estimated for decolorization process using free cells and

immobilized cells. The kinetic parameters Kdye and Vdye

max using different diameter of immobilized beads can well

be used for the design and scale up of continuous reactors.for immobilized cells compared to that of free cells for all

the dyes tested and decreases with increase in the diameter

of the beads. This might be due to the fact that rate of

decolorization will be much faster in freely suspended cells

and at higher initial concentrations, the substrate inhibited

the decolorizing activity.

4. Conclusion

P. chrysosporium MTCC 787 is found to be suitable for

the decolorization of the different classes of dyes. An

optimum glucose and nitrogen concentration of 5 and

0.05 g/l, respectively, in the basal medium are found to be

useful for the enhancement of maximum decolorization.

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K.V. Radha et al. / Process Biochemistry 40 (2005) 33373345 3345

Decolorization studies of synthetic dyes using Phanerochaete chrysosporium and their kineticsIntroductionMaterials and methodsMicroorganismInoculumDyes and decolorization studiesEnzyme assaysBiomass determinationBiosorption studiesImmobilization

Results and discussionEffect on initial concentrations of the dyesBiomass growthBiosorption studiesEffect of pH and temperatureEffect of glucose and nitrogenEffect on size of inoculumStudies on immobilized beadsKinetic studies

ConclusionAcknowledgementReferences