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360 Journal of Basic Microbiology 2010, 50, 360 367
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Research Paper
Production of laccase fromPleurotus florida
usingagro-wastes and efficient decolorization of Reactive blue 198
P. Sathishkumar1, K. Murugesan
2and T. Palvannan
1
1Bioprocess and Genetic Engineering Lab, Department of Biochemistry, Periyar University, Salem,Tamil Nadu, India
2School of Environmental Science and Engineering, Pohang University of Science and Technology,San, Hyojadong, Namgu, Pohang, Republic of Korea
Pleurotus florida NCIM 1243 produced laccase as the dominant lignolytic enzyme during the dye
decolorization. Banana peel was the best substrate for extracellular laccase production under
solid state fermentation when compared to mandarin peel and cantaloupe peel. The maximum
activity of laccase (5.4 U/g) was detected on the 10 day. The ratio of banana peel : mandarin
peel : cantaloupe peel (5:2:3) showed increased production of laccase (6.8 U/g). P. florida pro-
duced two extracellular laccase isoenzymes (L1 and L2). The half life of laccase at 60 C was 2 h
and at 4 h it retained 25% residual activity. P. florida laccase showed high thermostability and
an interesting difference was noticed in the behavior of laccase isoenzymes at different
temperature. The L1 isoenzyme of laccase showed remarked thermostability at 60 C in the
native PAGE when compared to L2 isoenzyme. The optimum pH, temperature and enzyme
concentration for maximum decolorization was found to be 4.5, 60 C and 1.2 U/ml, respec-
tively. Partially purified laccase enzyme showed excellent decolorization activity to Reactive
blue 198. The maximum decolorization (96%) was observed at lower dye concentrations
(50100 ppm) which decreased markedly when the dye concentration was increased beyond
150 ppm. The thermostable laccase ofP. florida could be effectively used to decolorize the
synthetic dyes in the textile effluent and other biotechnological applications.
Keywords: Laccase /Pleurotus florida /Thermostability / Solid state fermentation / Dye decolorization /
Reactive blue 198
Received: December 02, 2009; accepted: April 07, 2010
DOI 10.1002/jobm.200900407
Introduction*
Synthetic dyes are widely used in the textile, paper,
cosmetics, leather, dyeing, color photography, pharma-
ceutical and food industries [1]. In textile industries,during dyeing process, about 1030% or more of the
dyes used are released into water bodies [2], causing
serious environmental problem in many parts of the
world. Although some of the dyes are not themselves
toxic, after release into the aquatic environment their
degradation products are often carcinogenic [3, 4]. De-
colorization of dyes by physical or chemical methods
Correspondence: Dr. T. Palvannan M.Sc., Ph.D., Bioprocess andGenetic Engineering Lab, Department of Biochemistry, Periyar Univer-sity, Salem 636 011, Tamil Nadu, IndiaE-mail: [email protected]: +91 427 2345766, 2345520
Fax: +91 427 2345124
including adsorption and precipitation methods, che-
mical degradation or photodegradation has practical
disadvantages. Further, they are also expensive, com-
mercially unattractive, time-consuming and mostly
ineffective [5]. On the other hand, dye decolorizationusing oxidative enzymes has received great attention in
recent years due to its efficient application [6].
White rot fungi have been shown to be able to decol-
orize synthetic dyes due to their lignolytic enzymes
such as lignin peroxidase, manganese peroxidase and
laccase [710]. Generally, the white rot fungi produce
either one of the above or all the three types of the
above lignolytic enzymes. Most of the white rot fungal
strains produce laccase as the main enzyme during dye
decolorization process.
Laccase (p-diphenol: oxygen oxidoreductase, EC
1.10.3.2) is a widespread group of multi-copper enzy-
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mes which can catalyzes the oxidation of a variety of
organic compounds, with concomitant reduction of
molecular oxygen to water. The potential applications
of laccases have been shown in various biotechnological
process including pulping, detergents, textile dyes,
biosensors, food industries, detoxification of pollution
and enzymatic conversion of chemical intermediates [6,
1115]. One of the advantages associated with laccases
is that they do not require H2O2 for substrate oxidation
unlike peroxidases and moreover they have broad sub-
strate specificity [16]. Thus, laccases of fungi have at-
tracted considerable attentions for academic and indus-
trial applications. Up to date, more than 100 laccases
have been isolated from fungal cultures and their bio-
chemical properties as well as their catalytic characters
have been characterized.Submerged fermentation does not mimic the natural
living conditions of white rot fungi. Solid state fermen-
tation (SSF), defined as the fermentation of solids in the
absence of free water, has the advantage of supporting
the growth and metabolism of microorganisms under
moisture conditions [17]. Production of enzymes by SSF
on agro-wastes has gained much attention in biotech-
nology due to its higher productivities and low produc-
tion costs [18]. The use of such wastes, besides providing
alternative substrates, helps to solve environmental prob-
lems, which are caused by their disposal. In addition,
most of these wastes contain lignin or/and cellulose andhemicellulose, which act as mediators of the lignolytic
activities. Furthermore, most of them are rich in sugars,
which make the whole process much more economical.
All these facts make them very suitable as raw materials
for the production of secondary metabolites of indus-
trial significance by microorganisms [19]. Laccase pro-
duced from SSF system using agro-wastes has been
proved as an effective synthetic dye decolorizer [20, 21].
Pleurotus florida is a white rot fungi which has great
biotechnological importance and its application on
industrial effluent is well known [2224]. In this study,
we used banana (Musa cavendishii), mandarin (Citrus re-ticulata) and cantaloupe (Cucumis melo) peels as the sub-
strate to produce extracellular lignolytic enzyme from
P. florida by the SSF. The extracellular enzyme was
evaluated for its decolorization capability against syn-
thetic dye, Reactive blue 198.
Materials and methods
Microorganism and chemicals
The white rot fungus, Pleurotus florida NCIM 1243 em-
ployed in this study was purchased from National Col-
lection of Industrial Microorganism, National Chemical
Laboratory (NCL), Pune, India.
ABTS (2,2-azinobis-3-ethylbenzothiazoline-6-sulfonic
acid) was purchased from Sigma-Aldrich. Dye used in
this study [Reactive blue 198 (RB 198)] was procured
from commercial textile dyeing industry at Erode, Ta-
milnadu, India and all other chemicals were of analyti-
cal grade.
Dye decolorization ability on agar plate
Dye degradation ability ofP. floridawas screened in low
nitrogen basal medium [25] amended with RB 198 dye
at concentration of 100 ppm in the presence of 1.5%
agar plate. Mycelia was placed on the centre of dye agar
plate and incubated at 30 C under dark condition.
Plates were regularly monitored for dye decolorizationactivities through the change of color from blue to
colorless for every 24 h.
Agricultural wastes
Chopped banana (Musa cavendishii), mandarin (Citrus reti-
culata) and cantaloupe (Cucumis melo) peels (particle size
7.5 mm 7.5 mm) were procured from local market,
and used as substrate for laccase production by P. florida
under SSF condition. Substrates were pre-treated as
follows: 10 g of each substrate which was freshly
collected were first soaked for an hour in 30 ml of
83.17 mM KOH at room temperature to neutralise or-ganic acids [26]. Then, they were thoroughly washed
with distilled water and dried at 50 C temperature.
Enzyme production and preparation
Laccase production was carried out in SSF using three
different agro-wastes such as banana peel (BP), manda-
rin peel (MP) and cantaloupe peel (CP) as the substrate.
Ten grams of moistened peels were transferred sepa-
rately to conical flask (250 ml) and autoclaved at 121 C
for 15 min. Five mycelial discs were transferred to each
flask supplemented with 100 M CuSO4 to induce lac-
case production and incubated at room temperature.Cultures were harvested at every 2 d interval for the
quantification of extracellular laccase enzyme.
Extracellular enzyme from SSF was extracted by
soaking the culture with 100 mM sodium acetate buffer
pH 5.0 at 4 C overnight. The culture supernatant was
filtered and centrifuged at 10,000 g for 15 min to
remove the fine particles. The supernatant was concen-
trated by Millipore Amicon ultrafiltration stirred cell
through a 0.22 m (10 kDa) filter under vacuum, until a
20-fold concentration was achieved. The enzyme was
precipitated with ammonium sulfate to a final concen-
tration of 80% (w/v). After standing in the ammonium
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sulfate solution for 5 h at 0 C, the precipitate was col-
lected by centrifugation at 10,000 g for 30 min and
resuspended with 100 mM of sodium acetate buffer
(pH 5.0) and dialysed against the same buffer for 12 h.
The resulting solution was used for the decolorization
studies.
Enzyme assay and protein estimation
Laccase activity was measured using ABTS as the sub-
strate at 30 C [27]. The assay mixture contained 50 mM
sodium acetate buffer (pH 5.0), 1 mM ABTS and diluted
laccase source. One activity unit was defined as the
amount of enzyme that oxidised 1 mol ABTS per min-ute. The absorbance increase of assay mixture was
monitored at 420 nm (420 = 36.0 mM1 cm1) in a UV
Vis spectrophotometer (Perkin-Elmer Lambda 25, Ger-many). The enzyme activities were expressed as unit
(U/ml or U/g).
Protein was estimated by the method of Lowry et al.
[28] using bovine serum albumin (BSA) as a standard.
Decolorization of RB 198 by P. floridalaccase
Dye decolorization efficiency of P. florida laccase en-
zyme obtained from SSF, was accessed using RB 198.
Reaction mixture contained of RB 198 dye, partially
purified laccase and 100 mM of buffer in a total volume
of 1 ml. The reaction mixture was incubated for dye
decolorization and the decolorization was measuredby monitoring the decrease in absorbance maximum
(592 nm) of RB 198 dye in a UVVis spectrophotometer
and expressed in terms of percentage. In parallel, con-
trol samples were maintained with heat inactivated
partially purified laccase.
Effect of different parameters on dye decolorization
The effect of pH ranging from 3 to 9 on the enzymatic
decolorization was monitored with fixed concentration
of dye (100 ppm) and enzyme concentration (2 U/ml) at
30 C. The pH of reaction mixture was adjusted by
citrate-acetate-phosphate (37) and TrisHCl (89). Tofind out the effect of temperature on the enzymatic
decolorization, the reaction mixture was incubated
with fixed concentration of dye (100 ppm), enzyme
(2 U/ml) and pH (4.5). The temperature range was be-
tween 20 to 80 C with 20 C increment. In order to
check the effect of enzyme quantity on dye decoloriza-
tion with fixed concentration of dye (100 ppm) and pH
(4.5) at 60 C, reaction was started with different quan-
tity of enzyme from 0.2 to 2 U/ml with the increment of
0.2 U/ml. The effect of dye concentration was tested
using 50, 100, 150, 200 and 250 ppm concentrations
with fixed concentration of enzyme (1.2 U/ml) and pH
4.5 at 60 C. For all these experiments, the decoloriza-
tion was monitored after 10 min incubation as descri-
bed above.
Thermostability of laccase
The thermostability of laccase was studied by incubat-
ing the enzyme at various temperatures from 20 to
80 C with the increment of 20 C. The residual laccase
activity was measured at different time intervals using
ABTS as substrate under standard assay condition.
Native PAGE
In order to identify the number of laccase isoenzymes
produced by P. florida and their thermostability, the
enzyme was subjected to non-denaturing polyacryla-
mide gel electrophoresis (10%). For zymogram analysis,the gel was incubated in 100 mM sodium acetate buffer
with 10 mM guaiacol for the laccase activity detection.
Results and discussion
Screening of dye decolorization
Decolorization ability ofP. floridawas screened by agar
plate amended with RB 198 dye. The mycelial growth
covered the agar plate completely on 6 d with 70%
decolorization zone. Further incubation lead to com-
plete decolorization on the 10 d (Fig. 1b). The decolori-zation of RB 198 proceeded through a color change
from blue to light pinkish blue and later colorless. Af-
ter 10 d of incubation, the plate was evaluated for the
type of lignolytic enzymes involved in dye decoloriza-
tion. In plate incorporated with 1 mM MnSO4, a very
slight brownish flake was observed indicating the in-
significant production of manganese peroxidase (Fig. 1c).
Another plate was screened for the laccase production
by incubating the plate with 1 mM ABTS in 50 mM
sodium acetate buffer (pH 5.0). An intense oxidation of
ABTS was observed by the formation of bluish green
color, indicating that the decolorization process wasmainly due to the production of laccase from P. florida
(Fig. 1d). The same result was observed during the de-
colorization of Remazol Brilliant Blue R (RBBR) by white
rot fungi Ganoderma lucidum [10].
Laccase production in solid state fermentation
Laccase production from P. florida in SSF using agricul-
tural wastes such as banana peel, mandarin peel and
cantaloupe peel are presented in Fig. 2 and Table 1.
Mycelial growth on the substrates was observed after
36 h incubation and the fungal mycelium completely
colonized the substrate within 6 days. The production
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Figure 1. Screening for dye decolorization by P. floridaNCIM 1243and its lignolytic enzymes on 100 ppm of RB 198 dye amendedagar plate. (a) Control; (b) after decolorization (10 days incubated);(c) Formation of manganese flakes (brown color) in MnSO
4
amended plate indicates manganese peroxidase production. (d)Oxidation of 1 mM ABTS in 100 mM sodium acetate buffer (pH 5.0)indicates laccase production.
of laccase was monitored for every second day. The
maximum laccase activities ofP. florida was found to
be 5.4 U/g (banana peel), 3.1 U/g (mandarin peel) and
4.0 U/g (cantaloupe peel) on 10 d incubation. Previously,
Osma et al. [29] reported that banana skin was novel
substrate for the growth and laccase production fromTrametes pubescens under SSF. This was attributed to
high carbohydrate content that might be easily metabo-
lized by the microorganism [30]. The same result was
observed in our study using P. floridawhich suggested
that banana peel can be used as cheap source for lac-
case production. Further experiments were carried out
with different ratio of BP:MP:CP for the laccase pro-
duction. The results indicated that a ratio of 5:2:3
(BP:MP:CP) increased production of laccase (6.8 U/g)
from P. florida (Table 1). This may be due to the syner-
getic action of the components present in all three
agro-wastes.
Figure 2. Production of laccase in SSF by P. floridaNCIM 1243.
Table 1. Effect of mandarin peel and cantaloupe peel supp-lementation to the banana peel in laccase production fromP. floridaNCIM 1243.
Ratio of banana peel : mandarin peelcantaloupe peel (w/w)
Laccase activity(U/g)
Banana peel (control) 5.4Mandarin peel (control) 3.1Cantaloupe peel (control) 4.05:5:0 4.75:4:1 5.65:3:2 6.15:2:3 6.85:1:4 6.25:0:5 5.6
Effect of different parameters on dye decolorization
by P. floridalaccase
P. florida extracellular laccase prepared from mixed
substrateculture (BP:MP:CP 5 :2 :3) was employed for
dye decolorization studies. The influence of pH on dye
decolorization was determined at different pH ranging
from 3 to 9 (Fig. 3). The results showed that the laccasehighly decolorize the dye in the range of pH 4 to 6. The
optimum pH for maximum decolorization (96%) was
pH 4.5. The decolorization activity decreased at pH 7
and no activity was observed at alkaline pH. Our result
shows that P. florida laccase favoring acidic range for
higher decolorization of RB 198 dye. This observation
was similar to the previous reports obtained by Mu-
rugesan et al. [10] and Kandelbauer et al. [31].
The effect of temperature on RB 198 dye decoloriza-
tion is shown in Fig. 4a. Temperature is one of the fac-
tors involved in dye decolorization. The result clearly
show that the optimum temperature for maximum de-
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Figure 3. Effect of pH on decolorization of RB 198 by P. floridaNCIM 1243 laccase.
colorization was 60 C, beyond that decolorization de-
creased sharply. In the case of dye samples at 30 C,
29% of decolorization was decreased when compared to
60 C. Murugesan et al. [10] reported that the white rot
fungi G. lucidum laccase had an optimum temperature
for RBBR decolorization at 60 C, which was achieved
by means of 1 h incubation. This was similar to our
result. However, P. florida laccase decolorize the dyemore rapidly (within 10 min). When incubated below
60 C, the percentage of decolorization was increased
with increase in time (data not shown). Some of the
white rot fungal laccases have been described as ther-
mostable, even though the most laccases are not active
at higher than 50 C [3235]. For example P. ostreatus
laccase showed a half life of 30 min at 60 C [36] and
P. eryngii laccase retained 10% residual activity at 60 C
after 30 min incubation [37]. However P. florida laccase
showed a half life of 2 h at 60 C and at 4 h it retained
around 25% residual activity (Fig. 4b) which indicates
its better thermostability than other Pleurotus specieslaccases. This could be due to the different ecological
origination of strains and entirely different culture
conditions. Further, in the above studies liquid culture
with chemically defined medium was used for laccase
production whereas in our study SSF with banana peel
was used. The laccase activity decreased rapidly at 80 C
and the complete inactivation occurred at 1 h incuba-
tion.
The effect of enzyme concentration on dye decolori-
zation (50 ppm) was studied to find out the amount of
enzyme essential for maximum decolorization. The
optimum concentration of laccase required for maxi-
Figure 4. a) Effect of temperature on decolorization of RB 198 byP. floridaNCIM 1243 laccase; b) Thermostability of P. floridaNCIM1243 laccase at various temperature.
mum decolorization was 1.2 U/ml (Fig. 5). However the
decolorization increased with increase in enzyme quan-tity. Previously, Soares et al. [38] had also reported that
increasing concentration of laccase increased the rate
of dye decolorization. Similarly, Pant and Adholeya [39]
reported that the concentrated laccase from P. florida
EM1303 obtained by ultrafiltration efficiently decolor-
ized undiluted distillery effluent. This further proves
the P. florida laccase utility in treating various effluents.
The effect of dye concentration on decolorization was
tested with different initial dye concentration from 50
to 250 ppm with constant amount of enzyme (1.2 U/ml).
The maximum decolorization (94%) was observed at
lower dye concentrations (50100 ppm) which decreas-
a)
b)
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Figure 5. Effect of enzyme quantity on RB 198 dye decolorization.
ed markedly when the dye concentration was increased
beyond 150 ppm (Table 2). The results revealed that the
decolorization decreased with increasing dye concen-
tration. However on further increasing the time of
incubation, a maximum decolorization was observed
(data not shown). It indicates that the tested dye con-
centrations up to 250 ppm are not inhibitory to the
enzyme whereas only the percentage of decolorization
was decreased with increasing dye concentration. At
high dye concentration the efficiency of enzyme wasreduced but did not completely diminish. This opti-
mized conditions, RB 198 was decolorized by partially
purified P. florida laccase and the absorbance spectra
were recorded Fig. 6. RB 198 decolorization was carried
in a UVVis spectrophotometer with the above opti-
mized condition and the results indicate the complete
decolorization of dye evident from the spectral scan
done between 480 and 680 nm. Interestingly, our P.
florida laccase rapidly decolorize the dye without addi-
tion of any mediator.
Laccase isoenzymesMany of the white rot fungi produce more than one
laccase isoenzyme. P. florida NCIM 1243 crude laccase
Table 2. Decolorization of RB 198 by P. florida NCIM 1243laccase at different dye concentration.
Decolorization of RB 198 (%)Dye concentration (ppm)
10 min 120 min
50 96 99100 93 99150 79 97200 43 93250 22 84
Figure 6. UVvis absorbance spectra of RB 198 decolorization byP. floridaNCIM 1243 laccase under 60 C and pH 4.5.
was subjected to zymogram analysis for the identifica-
tion of laccase isoenzymes. The results revealed that
two laccase isoenzymes (L1 and L2) were produced by
this strain (Fig. 7a). Of these two isoenzymes, L1 was
more dominant than L2. Similar pattern of laccase
isoenzymes was observed by Das et al. [32] in a previous
study using P. florida. In order to identify thermostabil-
ity of these isoenzymes, crude laccase was incubated at
different temperature for 1 h and analysed the stability
for guaiacol oxidation on native-PAGE (Fig. 7b). L1isoenzyme was more stable at 60 C than L2 isoenzyme
indicating that L1 isoenzyme of laccase is mainly re-
sponsible for dye decolorization at 60 C. At 80 C both
isoenzymes lost their activity within 1 h.
Figure 7. (a) Zymogram analysis of P. floridaNCIM 1243 laccaseisoenzymes (L1 and L2) on native PAGE by guaiacol oxidation. (b)Thermostability of P. florida NCIM 1243 laccase isoenzymes after1 h incubation at different temperature. Lane 1, 2 and 3 incubated
at 80 C, 60 C and 40 C, respectively.
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Conclusion
To our knowledge, this is the first report on thermosta-
ble laccase fromP. florida and its efficient dye decoloriz-
ing activity. Production and preparation of laccase from
this fungus using agro-wastes is easy and economical.
As the laccase in the present study shows good thermo-
stability at high temperatures it could be effectively
used to decolorize the synthetic dyes in textile effluent.
Acknowledgements
The corresponding author (Principal Investigator
Project No.: BT/PR8973/GBD/27/57/2006 Dt 14/08/07) is
grateful to Government of India, Ministry of Science &
Technology, Department of Biotechnology for financialsupport.
References
[1] Marmion, D.M., 1991. Handbook of US colorants. Foods,Drugs, Cosmetics and Medical Devices. John Wiley &Sons, New York, USA.
[2] Srinivasan, S.V., Murthy, D.V.S., 2009. Statistical optimi-zation for decolorization of textile dyes using Trametes ver-sicolor. J. Hazard. Mater., 165, 909914.
[3] Shyam, S.D., Satyshari, D., Bhattacharyya, B.C., 1995. Dyedecolorization in a column bioreactor using wood-degrad-
ing fungusPhanerochaete chrysosporium. Indian Chem. Eng.,A37, 176 180.
[4] Banat, I.M., Nigam, P., Singh, D., Marchant, R., 1996.Microbial decolorization of textile dye-containing efflu-ents: a review. Bioresour. Technol., 58, 217227.
[5] Cooper, P., 1993. Removing colour from dye housewastewater e a critical review of technology available. J.Soc. Dyers Colour, 109, 97108.
[6] Couto, S.R., Toca Herrera, J.L., 2006. Industrial and bio-technological applications of laccases: A review. Biotech-nol. Adv., 24, 500 513.
[7] Covino, S., Svobodov, K., Kesinov, Z., Petruccioli, M.,Federici, F., DAnnibale, A., vanarov, M., Cajthaml, T.,2010.In vivo and in vitro polycyclic aromatic hydrocarbons
degradation by Lentinus (Panus) tigrinus CBS. Bioresour.Technol., 101, 3004 3012.
[8] Saha, S.K., Swaminathan, P., Raghavan, C., Uma L., Subra-manian, G., 2010. Ligninolytic and antioxidative enzymesof a marine cyanobacterium Oscillatoria willei BDU 130511during Poly R-478 decolourization. Bioresour. Technol.,101, 30763084.
[9] Pant, D., Singh, A., Satyawali, Y., Gupta, R.K., 2008. Effectof carbon and nitrogen source amendment on syntheticdyes decolourizing efficiency of white-rot fungus, Phane-rochaete chrysosporium. J. Environ. Biol., 29, 7984.
[10] Murugesan, K., Nam, I.H., Kim, Y.M., Chang, Y.S., 2007.Decolorization of reactive dyes by a thermostable laccaseproduced by Ganoderma lucidum in solid state culture. En-
zyme Microb. Technol., 40, 1662 1672.
[11] Khlifi, R., Belbahri, L., Woodward, S., Ellouz, M., Dhouib,A., Sayadi, S., Mechichi, T., 2010. Decolourization and de-toxification of textile industry wastewater by the laccase-mediator system. J. Hazard. Mater., 175, 802808.
[12] Minussi, R.C., Pastore, G.M., Durn, N., 2002. Potentialapplications of laccase in the food industry. Trends FoodSci. Technol., 13, 205216.
[13] Majeau, J.A., Brar, S.K., Tyagi R.D., 2009. Laccases forremoval of recalcitrant and emerging pollutants. Biore-sour. Technol., DOI:10.1016/j.biortech.2009.10.087.
[14] Vianello, F., Cambria, A., Ragusa, S., Cambria, M.T., Zen-naro, L., Rigo, A., 2004. A high sensitivity amperometricbiosensor using a monomolecular layer of laccase as bio-recognition element. Biosens. Bioelectron., 20, 315321.
[15] Sadhasivam, S., Savitha, S., Swaminathan, K., 2009. De-ployment of Trichoderma harzianum WL1 laccase in pulpbleaching and paper industry effluent treatment. J. Clean.Product., DOI:10.1016/j.jclepro.11.014.
[16] Saito, T., Hong, P., Kato, K., Okazaki, M., Inagaki, H.,Maeda, S., 2003. Purification and characterization of anextracellular laccase of a fungus (family Chaetomiaceae)isolated from soil. Enzyme Microb. Technol., 33, 520526.
[17] Pandey, A., 2003. Solid state fermentation. Biochem. Eng.J, 13, 81 84.
[18] Pandey, A., Soccol, C.R., Nigam, P., Soccol, V.T., 2000. Bio-technological potential of agro-industrial residues. I. Sug-arcane Bagasse Bioresour. Technol., 74, 69 80.
[19] Kalogeris, E., Iniotaki, F., Topakas, E., Christakopoulos, P.,Kekos, D., Macris, B.J., 2003. Performance of an intermit-tent agitation rotating drum type bioreactor for solid-state fermentation of wheat straw. Bioresour. Technol.,86, 207213.
[20] Gmez, J., Pazos, M., Couto, S.R., Sanromn, M.A., 2005.Chestnut shell and barley bran as potential substrates forlaccase production by Coriolopsis rigida under solid-stateconditions. J. Food Eng., 68, 315319.
[21] Shrivastava, R., Christian, V., Vyas, B.R.M., 2005. Enzyma-tic decolorization of sulfonphthalen dyes. Enzyme Mic-rob. Technol., 36, 333337.
[22] Shanmugam, S., Palvannan, T., Sathish Kumar, T., Micha-el, A., 2005. Biological decolourization of textile and pa-per effluents by Pleurotus florida and Agaricus bisporus.World J. Microbiol. Biotechnol., 21, 11491151.
[23] Pant, D., Adholeya, A., 2007. Identification, ligninolyticenzyme activity and decolorization potential of two fungiisolated from a distillery effluent contaminated site. Wa-
ter Air Soil Pollut., 183, 165 176.[24] Pant, D., Adholeya, A., 2007. Enhanced production of
ligninolytic enzymes and decolorization of molassesdistillery wastewater by fungi under solid state fermenta-tion. Biodegradation, 18, 647659.
[25] Arulmani, M., Murugesan, K., Arumugam, P., Dhandapa-ni, R., Kalaichelvan, P.T., 2005. Decolorization of basic dy-es methyl violet and emerald green by Pleurotus sajor-cajuand its effluent decolorization activity. Indian J. Appl.Microbiol., 1, 4752.
[26] Stredansky, M., Conti, E., 1999. Xanthan production bysolid state fermentation. Proc. Biochem., 34, 581 587.
[27] Niku-Paavola, M.L., Raaska, L., Itvaara, M., 1990. Detec-tion of white-rot fungi by a non-toxic stain. Mycol. Res.,
94, 27 31.
-
7/30/2019 jobm.200900407aa
8/8
Journal of Basic Microbiology 2010, 50, 360367 Production of laccase from Pleurotus florida using agro-wastes 367
2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.jbm-journal.com
[28] Lowry, O.H., Rosebrough, N.J., Farr, A.F., Randall, R.J.,1951. Protein measurement with the folin phenol re-agent. J. Biol. Chem., 193, 265275.
[29] Osma, J.F., Toca Herrera, J.L., Couto, S.R., 2007. Bananaskin: A novel waste for laccase production by Trametes pu-bescens under solid-state conditions. Application to syn-thetic dye decolouration. Dyes Pigm., 75, 3237.
[30] Essien, J.P., Akpan, E.J., Essien, E.P., 2005. Studies onmould growth and biomass production using waste bana-na peel. Bioresour. Technol., 96, 14511456.
[31] Kandelbauer, A., Erlacher, A., Cavaco-Paulo, A., Guebitz,G.M., 2004. Laccase catalyzed decolorization of the syn-thetic azo-dye diamond black PV 200 and of some struc-turally related derivatives. Biocatal. Biotransfor., 22, 331339.
[32] Das, N., Chakraborty, T.K., Mukherjee, M., 2001. Purifica-tion and characterization of a growth-regulating laccase
fromPleurotus florida. J. Basic Microbiol., 41, 261267.[33] Cambria, M., Cambria, A., Ragusa, S., Rizzarelli, E., 2000.
Production, purification and properties of an extracelularlaccase from Rigidoporus lignosus. Protein Expres. Puri., 18,141147.
[34] Hublick, G., Schinner, F., 2000. Characterization and im-mobilization of the laccase from Pleurotus ostreatus and its
use for the continuous elimination of phenolic pollutants.Enzyme Microb. Technol., 27, 330336.
[35] De Souza, C.G.M., Peralta, R.M., 2003. Purification and
characterization of the main laccase produced by thewhite-rot fungusPleurotus pulmonarius on wheat bran solidstate medium. J. Basic Microbiol., 43, 278286.
[36] Palmieri, G., Giardina, P., Marzullo, L., Desiderio, B., Nitti,B., Cannio, R., Sannia, G., 1993. Stability and activity ofphenoloxidase from lignolytic fungus Pleurotus ostreatus.
Appl Microbiol. Biotechnol., 39, 632636.
[37] Munoz, C., Guillen, F., Martinez, A.T., Martinez, M.J.,1997. Laccase isoenzymes ofPleurotus eryngii: Characteri-zation, catalytic properties and participation in activationof molecular oxygen and Mn
2+oxidation. Appl. Environ.
Microbiol., 63, 21662174.
[38] Soares, G.M.B., Costa-Ferreira, M., Amorim, M.T.P., 2001.Use of laccase together with redox mediators to decolori-
ze Remazol Brilliant Blue R. J. Biotechnol., 89, 123129.[39] Pant, D., Adholeya, A., 2009. Concentration of fungal lig-
ninolytic enzymes by ultrafiltration and their use in dis-tillery effluent decolorization. World J. Microbiol. Bio-technol., 25, 17931800.
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