




































In ternationa l
Scholars
Journa ls

 

African Journal of Environmental Economics and Management ISSN 2375-0707 Vol. 2 (4), pp. 195-197, April, 
2014. Available online at www.internationalscholarsjournals.org © International Scholars Journals 

 

Author(s) retain the copyright of this article. 

Short Communication 
 

A study of synthetic lignin decolorization ability of P. 
chrysosporium, T. hirsuta and marine fungi  

 

Basavapunnaiah Naidu1, Ranga Reddy1, Rao Koratala Satyanarayana2 and Venkayya 
Prakasam2  

 
1
Department of Environmental Sciences, Faculty of Agricultural Sciences, Delhi Technological University, New Delhi, India. 

2
Department of Environmental Studies, Faculty of Biological Sciences, Taleigão, Goa, India, Goa University. 

 

Accepted 23 February, 2014 
 

Lignin is an organic compound abundantly found in wood. Degradation of lignin poses major problems for paper mill 
industries all over the world with regard to the dark- brown color of lignin in wastewater, which is also a major problem 
facing the environment today. The decolourization capability was tested with species like Phanerocheate chrysosporium, 
Trametes hirsuta and an isolated native Marine fungi – M1 using synthetic lignin. The result showed that, isolated strain 
decreased the lignin color by up to 20% after cultivation for 7 days. Where the decolourisation ability of the fungi 
Phanerocheate chrysosporium and Trametes hirsuta was observed to 80 and 70% respectively. 

 
Key Words: Lignin, decolourisation, Phanerocheate chrysosporium, Trametes hirsuta. 

 
INTRODUCTION 
 

Bioremediation is defined as the application of biological 
processes to the treatment of pollution. Most research 
within the field of bioremediation has focused on bacteria, 
with fungal bioremediation (Mycoremediation) attracting 
interest just within the past two decades. White rot fungi 
can withstand toxic levels of most organopollutants (Aust 
et al., 2003). White rot fungi is a physiological grouping of 
fungi that can degrade lignin (and lignin – like 
substances). Four main genera of white rot fungi have 
shown potential for bioremediation: Phanerocheate sp, 
Trametes sp, Bjerkandera sp and Pleurotussp (Hestbjerg 
et al., 2003). The main mechanism of biodegradation 
employed by this group of fungi, however, is the lignin 
degradation system of enzymes. These extracellular 
lignin – modifying enzymes have very low substrate 
specificity so they are able to mineralize a wide range of 
high recalcitrant organo pollutants that are structurally 
similar to lignin (Cajthaml et al., 2002; Mansur et al., 
2003; Pointing, 2001; Veignie, 2004).  

The fact that these fungal enzymes work extracellularly 
allows them to access many of the non-polar, non-soluble 
toxic compounds that intracellular processes cannot 
(Reddy and Mathew, 2001; Levin et al., 2003). The three 
main lignin – modifying enzymes are lignin peroxidases, 
 
 
 
Corresponding Author Email: basavapunnaiah123@gmail.com 

Mn-dependent peroxidase and laccase. All three of these 

enzyme groups are stimulated by nutrient limitation 

(Mansur et al., 2003; Aust et al., 2003). They are most 

effective at degrading lignin and lignin-like substances 

when certain nutrient levels, primarily nitrogen are low.  
Conversely, activities of these enzymes are completely 

suppressed in media containing high levels of nitrogen. This 

characteristic is advantageous for the fungi inhabiting 
highly contaminated sites with very low productivity due 
to toxic levels of organo pollutants (Reddy and Mathew, 
2001).  

In recent years, many researches indicated that white 
rot fungus is a promising microbe in wastewater 
treatment. The basidiomycete Phanerochaete 
chrysosporium in white rot fungi is the most extensively 
studied and discussed (Tien, M. and Kirk, T. K, 1983). 
The extracellular ligninolytic enzymes of white rot fungi 
have the ability to degrade a wide spectrum of recalcitrant 
organo-pollutants such as chlorinated phenols and 
various types of synthetic dyes because of their 
nonspecific characteristics (Heinfling, A et al., 1997; 
Novotny, C., et al., 2001). And some significant mineralization 

(20%&48%) was observed during degradation of azo dyes 

(Spadaro, J. T et al., 1992).  
The study of biodegradation of lignin by wood-rotting 
fungi is limited not only by lack of taxonomy in referring 
to genetic variety, but also in their potential for 
industrial  use.  The  studies  have  mainly  focused  on 



Basavapunnaiah et al.           195 
 
 
 
 
 
 
 
 
 

 
(Before Decolorization) (After Decolorization)  
(a)  
Decolorization by Phanerochaete chrysosporium in 
5 gm/l concentration 

 
 
 
 
 

 
(Before Decolorization) (After Decolorization) 
(b)  
Decolorization by Phanerochaete chrysosporium 
in 10gm/l concentration 

 
 
 
 
 

 
(Before Decolorization) (After Decolorization) 
(c)  
Decolorization by Phanerochaete chrysosporium 
in 15gm/l concentration 

 
Figure 1.  Decolorization by Phanerochaete chrysosporium 

 
 

 
Phanerochaetechrysosporium (Boominathan and Reddy, 
1992). Until now, selectively few species of 
basidiomycetes have been used in lignin degradation 
studies, such as Bjerkandera, Pleurotus, Polyporus, 
Pycnoporus and Trametes (Addleman and Archibald, 
1993; Paice et al., 1993; Eggert et al., 1996). Species of 
marine fungi that may have potential capacity for use in 
the pulp and paper industry can possibly be used 
effectively, since these are available in great number 
along the coastal zones. Studies are therefore required in 
order to determine not only variations in fungal species 
and lignin degradation capabilities, but also to establish 
species or strains that may be suitable for 
biotechnological applications. The purpose of this work 
was to study the ability of white rot fungi Phanerochaete 
chrysosporium, Trametes hirsute and a marine fungus to 
decolorize synthetic lignin. 
 
 
METHODOLOGY 
 
Chemicals 
 
All the chemicals used were procured from Himedia Laboratories 
Pvt.Ltd, Mumbai, India. 

 
 
 

 
Microorganisms 
 
Commercial strains of Phanerocheate chrysosporium (MTCC 787), 
Trametes hirsuta (MTCC 136) were procured from IMtech, 
Chandigarh, India. A native marine fungal species (Here by named 
as M1) was isolated from the coastal waters of Visakhapatnam. 

 
Culture Conditions 
 
All the cultures were cultured on PDA. The cultures were 

maintained at 27
0
C and were sub-cultured periodically. 

 
Decolorization Studies 
 
Comparative decolorization ability of the three fungi was studied by 
adding commercial lignin. The experimental procedure was as 
follows. The 7 days, 10 days and 7 days old cultures of  
Phanerochaete chrysosporium and Trametes hirsuta and M1 
respectively, were inoculated in the medium containing synthetic 
lignin under sterile conditions. The concentration of synthetic lignin 

in the culture medium was 2 mg·L
-1

, 4 mg·L
-1

, 6 mg·L
-1

. The 
decolorization potential of the fungi was observed after 7 days. 
 
 
Results 
 
The main colored substances of general kraft pulp 
bleaching effluent were lignin and its related compounds. 
Decolorizing activity by the test fungal species toward a 
commercial lignin on plate assay is obvious, as shown in 
Figures. In this solid state decolorization, the mycelial 
growth of the microorganisms and the decolorization 
patterns were observed to be as follows: 
 
Phanerochaete chrysosporium: Ligninocellulosic 
materials were able to induce ligninolytic enzyme 
production in many fungi. The increase in lignin 
concentration is correlated with the decolorization 
capacity of extracellular medium. The growth of the 
fungus started on the third day in the concentrations 5 

g·L
-1

, 10g·L
-1

, where as no growth was observed in 

15g·L
-1

 till seventh day, which may be attributed to higher 
concentration of the lignin. The growth was observed to 
be less in comparison with the other two fungi. The 
decolorization of synthetic lignin by P. chrysosporium was 
observed to be faster than the decolorization by T. hirsuta 
and the Marine fungi. The decolorization by P. 
chrysosporium was observed to be more than the other 

two fungi on the 21
st

 day of incubation in the 

concentrations 5 g·L
-1

, 10g·L
-1

, however the ability 
decreased at higher concentration. (Figure 1).  

Many workers have reported that Lignin peroxidase has 
been shown to be involved in dye decolorization, mainly 
in P. chrysosporium cultures (Bumpus and Brock, 1988; 
Cripps et.al., 1990; Ollikka et.al.,1993; Paszczynski and 
Crawford 1991 and Young and Yu, 1997). 
 
Trametes hirsuta: The growth of the fungus started on 

the Eighth day in the concentrations 5g·L
-1

, 10g·L
-1

, 



  
 
 
 
 
 
 
 
 
 

 
(Before Decolorization) (After Decolorization)  
(a) 
Decolorization by Trametes hirsuta in 5gm/l concentration 

 
 
 
 
 
 
 

(Before Decolorization) (After Decolorization) 
(b) 
Decolorization by Trametes hirsuta in 10gm/l concentration 

 
 
 
 
 
 

 
(Before Decolorization) (After Decolorization) 
(c) 
Decolorization by Trametes hirsuta in 15gm/l concentration 

 
Figure 2.  Decolorization by Trametes hirsute 

 
 
 

where as no growth was observed in 15g·L
-1

 till Eleventh 
day, which may be attributed to higher concentration of 
the lignin. The growth was observed to be luxurious when 
compared with other two fungi. The decolorization of 
synthetic lignin by T. irsute was observed to be slower 
than the decolorization by P chrysosporium and the 
Marine fungi. The decolorization by T. irsute was 

observed to be lesser than P.chrysosporium on the 21
st

 
day of incubation in all the three concentrations. Even 
though the growth was observed to be luxurious the 
ability to decolorize has reduced with the increase in 

concentrations of 10g·L
-1

 and 15g·L
-1

 (Figure 2).  
Sathiya Moorthi et.al., 2007 has studied the 

decolorization capability of T.hirsuta and proved the 
laccase activity from this fungus through a Dark brown 
color that indicated laccase activity of culture filtrate (well-
1 Glass distilled water; well-2 20 µl of crude enzyme and 
well-3 30 µl crude enzyme). 
 
Marine Fungi: The growth of the fungus started on the 

Second day in the concentrations 5g·L
-1

, 10g·L
-1

, and in 

15g·L
-1

. The growth was observed to be luxurious when 
compared with P. chrysosporium and less when 
compared to T.hirsuta. The decolorization of synthetic 
lignin by marine fungi was observed to be slower than the 

 
 
 
 
 
 
 
 
 
 
 
(Before Decolorization)   (After Decolorization) (a) 
Decolorization by Marine fungi in 5gm/l concentration 
 
 
 
 
 
 
 
 
(Before Decolorization) (After Decolorization) (b) 
Decolorization by Marine fungi in 10gm/l concentration 
 
 
 
 
 
 
 
(Before Decolorization) (After Decolorization) (c) Decolorization 
by Marine fungi in 15gm/l concentration 
 
Figure 3. Decolorization by Marine fungi 
 
 

 
decolorization by P. chrysosporium. The decolorization by 
marine fungi was observed to be lesser than the other 

two fungi on the 21
st

 day of incubation in all the three 
concentrations. (Figure 3). 
 
 
Discussion 
 
Pointing (1999) showed that qualitative assays are 
powerful tools used in screening fungi for lignocellulose 
degrading enzyme production. Such tests give a positive 
or negative indication of enzyme production. They are 
particularly useful in screening large numbers of fungal 
isolates for several classes of enzyme, where definitive 
quantitative data are not required. All fungal isolates were 
screened for the presence of laccase, lignin-peroxidase 
and manganese dependent peroxidase activities by using 
an agar plate assay as a qualitative method for the 
determination of lignocellulolytic enzyme production 
(Atalla et.al., 2010).  

With the aim of finding decolorization activity at higher 
concentrations and because extracellular ligninolytic 
enzymes have been shown to be induced by growth on 
natural lignin substrates, the three fungal species were 
grown in media containing different concentrations of 
synthetic lignin substrates. In the present study the 
synthetic lignin decolorization ability of P. chrysosporium, 

196         Afr. J. Environ. Econ. Manage. 



Basavapunnaiah et al.           197 
 
 
 
T. hirsuta and marine fungi were studied. Compared with  
T. hirsuta and marine fungi, P. chrysosporium have 
shown the highest ability to decolorize. However, the 
extent of color removal decreased with increase in 
concentration. Decolorization depends upon on the 
laccase production, media and dyes. Similar observation 
regarding dye degradation by the white rot fungus P. 
chrysosporium has been observed by Spardaro et al. 
(1992). These results also show that in conclusion, 
several industrial dyes were decolorized by extra cellular 
enzymes from different strains of fungi. This appears to 
be a good application for immobilization and use as a 
bioreactor for effluent treatment from the dye and printing 
industries. 
 
 
Conclusion 
 
Lignin, which is widespread in nature, especially in all-
higher plants, is a hydrocarbon aromatic compound. Its 
complex structure makes lignin very difficult to degrade 
and therefore it can be persistent in the environment. 
Lignin degradation is important in the pulp and paper 
industry worldwide, which uses chemical substances to 
breakdown lignins in pulp processing. The process 
releases hazardous lignin-compound effluents into the 
environment that are toxic and carcinogenic (Harazono et 
al., 1996; Elizabeth Rodrý´guez, 1999). P. chrysosporium 
showed the highest ability to decolorize synthetic lignin, 
the study indicated that the lignin decolorization by 
marine fungus was competent with the standard species 
and can provide a cost-effectivess in using native 
organisms. But further studies are required to prove the 
ability of the fungi to degrade such very high 
concentrations of lignin, so that it can be adopted and 
applied for lignin degradation in paper and pulp mill 
effluents. 
 
 
References 
 
Addleman K, Archibald F (1993). Kraft pulp and delignification by 

dikaryons and monokaryons of Trametes versicolor. Appl. Environ. 
Microbiol 59: 266-273.  

Atalla M, Mabrouk Z, Kheiralla H, Eman R, Hamed A, Youssry A, Abeer 
AA (2010). Screening of some marine-derived fungal isolates for 
lignin degrading enzymes (LDEs) production. Agric. Biol. J. North 
Amer., 1(4): 591-599.  

Aust SD, Swaner PR, Stahl JD (2003).Detoxification and metabolism of 
chemicals by white-rot fungi. Pesticide decontamiantionand 
detoxification J.J.P.C Zhu S.D.Asut A.T.Lemley Gan, Washington, 
D.C.: Oxford University Press. 3 – 14.  

Boominathan K, Reddy CA (1992). Fungal biodegradation of lignin: 
biotechnological applications. In: Handbook. Appl. Microbiol. 4 (eds. 
D.K. Arora, R.P. Elander and K.G. Mukerji). Marcel Dekker, New 
York: 763-822.  

Bumpus JA, Brock BJ (1988). Biodegradation of crystal violet by white 
rot fungus Phanerochaete chrysosporium. Appl Environ Microbiol 
54:1143–1150.  

Cajthaml TM, Moder P, Kacer V, Sasek PP (2002). Study of fungal 
degradation products of polycyclic aromatic hydrocarbons using gas 

 
 
 

 
chromatography with ion trap mass spectrometry detection. J. 
Chromatogr. A 974: 213 – 22.  

Cripps C, Bumpus JA, Aust SD (1990). Biodegradation of azo and 
heterocyclic dyes by Phanerochaete chrysosporium. Appl Environ 
Microbiol 56:1114–1118.  

Eggert E, Temp U, Eriksson KEL (1996). The ligninolytic sistem of the 
white rot fungus Pycnoporus cinnabarinus: purification and 
characterization of the laccase. Appl. Environ. Microbiol. 62: 1151-
1158.  

Elizabeth R, Michael AP, Rafael V-D (1999). Industrial Dye 
Decolorization by Laccases from Ligninolytic Fungi. Current 
Microbiol. 38: 27–32.  

Heinfling A, Bergbauer M, Szewzyk U (1997). Biodegradation of azo 
and phthalocyanine dyes by Trametes versicolor and Bjerkandera 
adusta, Appl. Microbiol. Biotechnol., 48(2): 261 – 266.  

Levin LA, Viale AF (2003). Degradation of organic pollutants by the 
white rot basidiomycetes Trametes trogii. Int. Biodeterior. 
Biodegradation 52: 1 – 5.  

Mansur MME, Arias JL, Cop-Patino MF, Gonzalez AE (2003). The 
white-rot fungus Pleurotus ostreatus secretes laccase isoenzymes 
with different substrate specificities. Mycologia 95(6): 1013 –1020.  

Novotny C, Rawal B, Bhatt M, Milind P, Vaclav S, Hans PM (2001). 
Capacity of Irpex lacteus and Pleurotus ostreatus for decolorization 
of chemically different dyes, J. Biotechnol., 89(2-3): 113 – 122.  

Ollikka P, Alhonmaki K, Leppanen V-M, Glumoff T, Raijola T, Suominen 
Y (1993). Decolorization of azo, triphenyl methane, heterocyclic, and 
polymeric dyes by lignin peroxidase isoenzymes from Phanerochaete 
chrysosporium. Appl Environ Microbiol., 59: 4010–4016.  

Paice MG, Reid IO, Bourbonnais R, Archibald FS, Jurasek L (1993). 
Manganese peroxidase produced by Trametes versicolor during pulp 
bleaching, demethylates and delignifies kraft pulp. Appl. Environ. 
Microbiol. 59: 260-265.  

Paszczynski A, Crawford RL (1991). Degradation of azo compounds by 
ligninase from Phanerochete chrysosporium: involvement of veratryl 
alcohol. Biochem Biophys Res Commun 178:1056– 1063.  

Pointing SB (1999). Qualitative methods for the determination of 
lignocellulolytic enzyme production by tropical fungi. Fungal Diversity, 
2: 17-33.  

Pointing SB (2001). Feasibility of bioremediation by white-rot fungi. 
Appl. Microbiol. Biotechnol. 57: 20 -33.  

Reddy CA, Mathew Z (2001). Bioremediation potential of white rot fungi. 
Fungi in bioremediation. G.M.Gadd Cambridge, U.K.:Cambridge 
University Press.  

Sathiya MP, Deecaraman M, Periyar SS, Murugesan K, Kalaichelvan 
PT (2007). Biosorption of textile dyes and effluents by Pleurotus 
florida and Trametes hirsuta with evaluation of their laccase activity. 
Iranian J. Biotechnol., 5(2): 114 – 118.  

Spadaro JT, Gold MH, Renganathan V (1992). Degradation of azo dyes 
by the lignin-degrading fungus Phanerochaete chrysosporium, Appl. 
Environ. Microbiol., 58: 2397 – 2401.  

Tien M, Kirk TK (1983). Lignin-degrading enzyme from the hymeno-
mycete Phanerochaete chrysosporium, Sci. 221: 661 – 663.  

Veignie EC, Rafin P, Woisel FC (2004). Preliminary evidence of the role 
of hydrogen peroxide in the degradation of benzo[a]pyrene by a non-
white rot fungus Fusarium solani. Environ. Pollut. 129: 1- 4.  

Young L, Yu J (1997). Ligninase-catalyzed decolorization of synthetic 
dyes.Water. Res., 31:1187–1193. 


