MANGAI.G,

II M.Sc., Biotechnology,

Guided By: Mr.SOURAV BHATTACHARYA, M.Sc.,

Faculty of Microbiology,

Genohelix,

Jain University,

Bangalore 560019

MYCOREMIDIATION OF CONGO RED - AN AZO DYE

INTRODUCTIONEnvironmental degradation due to

rapid growing population and economic development.

Chemical wastes and byproducts or discharges of poorly-treated or untreated sewage from the dye and textile industries are the major source of water pollution.

Bioremediation agents in the treatment of wastewater containing textile dyes, which are the major source of aromatic amines.

More than 10,000 dyes used in textile industry and 280,000 tons of textile dyes are discharged every year worldwide.

Azo dyes contribute to about 70% of all used dyes, and have complex structure and synthetic nature.

The colour of dyes is due to azo bond and Nitrogen to nitrogen double bond (–N N–) and associated chromophores.

Azo compounds are xenobiotic compounds that are very recalcitrant against biodegradative processes.

Disposal of dyes into surface water not only affects the aesthetic but cause also biotoxicity.

Congo red is one of the predominantly used azo dye in the textile industries. It is a secondary diazo dye and toxic in nature.

Many physicochemical techniques are high cost, low efficiency and inapplicability to a wide variety of dyes.

A definitive solution of the colour problem of textile effluents would provide a marked competitive advantage for this industrial sector.

OBJECTIVES♦Screening of the laboratory fungal

forms for degradation activity.

♦To check the degradation activity under static and shaking condition.

♦To check the degradation activity when dye is the sole source of Nitrogen.

♦To check the degradation activity when media is free of CuSO4.

♦To check the degradation activity when media is supplemented with inorganic fertilizer.

♦To check the degradation activity when media is supplemented with heavy metals.

METHODOLOGY

Screening of the laboratory fungal forms for degradation activity.

• Plating of semi synthetic media with congo red (0.050g/L).

• Plates are inoculated with the fungal species such as Aspergillus niger, Aspergillus oryzae, Aspergillus flavus, Penicillium spp, Cladosporium spp, Pleurotus spp.

• Zone of decolourization around the fungal growth was observed after the incubation at 27° C.

To check the degradation activity under static and shaking condition.

• Preparation of Semi synthetic broth with congo red (0.050g/L).

• Inoculation of fungal species.

• Incubation at static and shaking condition (120 rpm).

• Decolourization observed through spectrophotometer at 570 nm.

To check the degradation activity when dye is the sole source of Nitrogen.

• Preparation of Semi synthetic broth with congo red (0.050g/L) as the sole Nitrogen source.

• Inoculation of fungal species.

• Incubation at static and shaking condition (120 rpm).

• Decolourization observed through spectrophotometer at 570 nm.

To check the degradation activity when media is free of CuSO4.

• Preparation of Semi synthetic broth without CuSO4 and with congo red (0.050g/L).

• Inoculation of fungal species.

• Incubation at static and shaking condition (120 rpm).

• Decolourization observed through spectrophotometer at 570 nm.

To check the degradation activity when media is supplemented with inorganic fertilizer (NPK fertilizer).

• Preparation of Semi synthetic broth with 1% NPK fertilizer and congo red (0.050g/L).

• Inoculation of fungal species.

• Incubation at static and shaking condition (120 rpm).

• Decolourization observed through spectrophotometer at 570 nm.

To check the degradation activity when media is supplemented with heavy metals.

• Preparation of Semi synthetic broth with HgCl2 (0.1%) & congo red (0.050g/L).

• Inoculation of fungal species.

• Incubation at static and shaking condition (120 rpm).

• Decolourization observed through spectrophotometer at 570 nm.

RESULTS♦ Screening of the laboratory fungal forms for

degradation activity has shown the zone of decolourization.

♦ The percentage of decolourization varied between 8.6% - 32.5% (static) & 10.9% - 98.86% (shaker).

♦ When dye is used as the sole Nitrogen source, the percentage of decolourization varied between 4.68% - 24.75% (static) & 8.41% - 39% (shaker).

♦ When SS broth is free of CuSO4, the percentage of decolourization varied between 5.98% - 29.9% (static) & 4.5% - 89.6% (shaker).

♦ When media is supplemented with NPK fertilizer the percentage of decolourization varied between 47.82% - 81.2% (static) & 49% - 99.8% (shaker).

♦ When media is supplemented with the heavy metal HgCl2 the percentage of decolourization varied between 89.1% - 95.5% (static) & 88% - 90.6% (shaker).

Decolourization activity is calculated by the formula Initial absorbance – Observed absorbance × 100 Initial absorbance

DECOLOURIZATION BY Aspergillus flavus

U – Uninoculated sampleA - 24 hrsB - 48 hrsC - 72 hrsD – 96 hrs

RESULTS To check the degradation activity

under static and shaking condition.

Uninoculated sample’s OD @ 570 nm = 0.440

S.N

MICRO ORGANISMS

O D @ 570 nm

24 hrs 48 hrs 72 hrs 96 hrs

ST SH

ST SH ST SH

ST SH

1 Aspergillus niger

0.423 0.409 0.420 0.399 0.413 0.390 0.402 0.384

2 Aspergillus oryzae

0.401 0.397 0.386 0.386 0.367 0.385 0.355 0.225

3 Aspergillus flavus

0.399 0.336 0.390 0.273 0.354 0.005 0.297 0.005

4 Penicillium spp 0.410 0.410 0.405 0.389 0.400 0.405 0.398 0.376

5 Cladosporium spp

0.394 0.411 0.391 0.405 0.388 0.400 0.382 0.392

6 Pleurotus spp 0.396 0.402 0.392 0.398 0.389 0.392 0.381 0.364

RESULTS To check the degradation activity when

dye is the sole source of Nitrogen.Uninoculated sample’s OD @ 570 nm =

0.832S.N

MICRO ORGANISMS

O D @ 570 nm

24 hrs 48 hrs 72 hrs 96 hrs

ST SH

ST SH ST SH

ST SH

1 Aspergillus niger

0.794 0.784 0.790 0.739 0.776 0.724 0.775 0.709

2 Aspergillus oryzae

0.783 0.730 0.757 0.696 0.738 0.690 0.693 0.660

3 Aspergillus flavus

0.764 0.587 0.743 0.555 0.686 0.525 0.626 0.507

4 Penicillium spp 0.811 0.767 0.783 0.767 0.757 0.762 0.737 0.762

5 Cladosporium spp

0.802 0.771 0.799 0.769 0.795 0.766 0.793 0.761

6 Pleurotus spp 0.803 0.773 0.792 0.760 0.788 0.759 0.787 0.733

RESULTSTo check the degradation activity when

media is free of CuSO4.Uninoculated sample’s OD @ 570 nm =

0.618S.N

MICRO ORGANISMS

O D @ 570 nm

24 hrs 48 hrs 72 hrs 96 hrs

ST SH

ST SH ST SH

ST SH

1 Aspergillus niger

0.614 0.596 0.589 0.596 0.587 0.596 0.581 0.590

2 Aspergillus oryzae

0.585 0.585 0.569 0.550 0.533 0.171 0.520 0.141

3 Aspergillus flavus

0.583 0.582 0.559 0.440 0.478 0.184 0.433 0.184

4 Penicillium spp 0.583 0.583 0.583 0.570 0.512 0.167 0.486 0.064

5 Cladosporium spp

0.583 0.582 0.581 0.582 0.574 0.503 0.569 0.440

6 Pleurotus spp 0.570 0.570 0.570 0.570 0.569 0.450 0.555 0.410

RESULTS To check the degradation activity when

media is supplemented with inorganic fertilizer(NPK fertilizer).

Uninoculated sample’s OD @ 570 nm = 0.506 S.

NMICRO ORGANISMS

O D @ 570 nm

24 hrs 48 hrs 72 hrs 96 hrs

ST SH

ST SH ST SH

ST SH

1 Aspergillus niger

0.318 0.340 0.019 0.330 0.011 0.250 0.001 0.214

2 Aspergillus oryzae

0.267 0.295 0.034 0.240 0.005 0.205 0.005 0.205

3 Aspergillus flavus

0.225 0.302 0.009 0.241 0.016 0.142 0.001 0.095

4 Penicillium spp 0.299 0.310 0.295 0.300 0.264 0.266 0.264 0.264

5 Cladosporium spp

0.299 0.300 0.271 0.282 0.268 0.269 0.258 0.222

6 Pleurotus spp 0.295 0.307 0.272 0.285 0.244 0.285 0.243 0.265

RESULTSTo check the degradation activity when

media is supplemented with heavy metal (HgCl2).

Uninoculated sample’s OD @ 570 nm = 1.785

S.N

MICRO ORGANISMS

O D @ 570 nm

24 hrs 48 hrs 72 hrs 96 hrs

ST SH

ST SH ST SH

ST SH

1 Aspergillus niger

0.238 0.238 0.219 0.217 0.185 0.198 0.176 0.199

2 Aspergillus oryzae

0.204 0.224 0.170 0.217 0.089 0.196 0.080 0.176

3 Aspergillus flavus

0.188 0.212 0.158 0.199 0.109 0.181 0.100 0.168

4 Penicillium spp 0.218 0.222 0.216 0.222 0.158 0.217 0.152 0.214

5 Cladosporium spp

0.217 0.214 0.191 0.201 0.167 0.187 0.163 0.167

6 Pleurotus spp 0.216 0.210 0.212 0.210 0.196 0.194 0.194 0.193

THANK ‘U’