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Development in
Wastewater Treatment
Research and
Processes
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Development in
Wastewater Treatment
Research and
Processes
Innovative Microbe-Based
Applications for Removal of
Chemicals and Metals in
Wastewater Treatment Plants
Edited by
Maulin P. Shah
Senior Environmental Microbiologist, Environmental Microbiology
Lab, Bharuch, Gujarat, India
Susana Rodriguez-Couto
Department of Separation Science, LUT School of Engineering
Science, LUT University, Mikkeli, Finland
Riti Thapar Kapoor

Assistant Professor (Grade-III), Amity Institute of Biotechnology,
Amity University, Uttar Pradesh, India
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Typeset by TNQ Technologies

Contents
Contributors.......................................................................................... xix
CHAPTER 1 A comparative study between physicochemical and

biological methods for effective removal of textile
dye from wastewater ............................................... 1
Subhasish Dutta and Joyani Bhattacharjee
1 Introduction ...................................................................... 1
2 Types of dyes and their toxicity............................................ 2
2.1 Azo dyes...................................................................3
2.2 Acid dye ...................................................................4
2.3 Basic dye ..................................................................4
2.4 Direct dye .................................................................4
2.5 Vat dye .....................................................................4
3 Physical methods ............................................................... 4
3.1 Adsorption.................................................................5
3.2 Coagulation and flocculation.........................................7
3.3 Emerging physical method for the treatment of textile
dye effluents ..............................................................8
4 Membrane technology......................................................... 8
4.1 Microfiltration............................................................8
4.2 Reverse osmosis .........................................................9
4.3 Nanofiltration.............................................................9
4.4 Ultrafiltration .............................................................9
5 Chemical methods.............................................................. 9
5.1 Fenton method ......................................................... 10
5.2 Ozonation................................................................ 10
5.3 Cucurbitiril .............................................................. 11
5.4 Sodium hypochlorite ................................................. 11
5.5 Recent biochemical trend ........................................... 12
6 Advanced oxidation process................................................13
7 Biological methods............................................................14
7.1 Degradation with bacteria........................................... 15
7.2 Degradation with algal culture .................................... 16
7.3 Degradation with yeast .............................................. 16
7.4 Degradation with fungi .............................................. 16
7.5 Degradation with white-rot fungi................................. 16
7.6 Enzyme system of white-rot fungi ............................... 17
8 Conclusion.......................................................................19
References .......................................................................19
v
vi Contents
CHAPTER 2 An approach toward developing clean green

techniques to deal with heavy metal toxicity
using the microbiome.............................................23
Subhasish Dutta and Pitam Chakrabarti
1 Introduction .....................................................................23
2 Different heavy metals and their impacts...............................24
2.1 Arsenic ................................................................... 24
2.2 Cadmium ................................................................ 25
2.3 Lead....................................................................... 25
2.4 Nickel..................................................................... 26
2.5 Mercury .................................................................. 26
2.6 Copper.................................................................... 26
2.7 Chromium ............................................................... 26
3 Bioremediation: the savior of the environment .......................28
3.1 Biosorption.............................................................. 28
3.2 Bioaugmentation: a new green technology .................... 30
3.3 Phytoremediation...................................................... 31
3.4 Mycoremediation, erasing environmental pollutants........ 32
3.5 Bioventing and biosparging ........................................ 33
3.6 Cyanoremediation..................................................... 34
3.7 Biostimulation.......................................................... 35
3.8 Bioleaching ............................................................. 36
4 Factors affecting bioremediation ..........................................37
4.1 Availability of nutrients ............................................. 37
4.2 Temperature............................................................. 37
4.3 pH.......................................................................... 37
5 Conclusion.......................................................................38
References .......................................................................38
CHAPTER 3 Microbial degradation of pesticides: microbial
potential for degradation of pesticides....................41
Sangeeta Kumari, Deepak Kumar and
S.M. Paul Khurana
1 Introduction: pesticides effect on environment health ..............41
1.1 Pesticides ................................................................ 43
1.2 Classification of pesticide........................................... 43
1.3 Pesticide fate in the environment ................................. 43
2 Biomagnification: tenacious effect of pesticides......................47
2.1 Bioaccumulation and biomagnification ......................... 47
3 Microbial potential: effective degradation of pesticides............48
Contents vii
4 Types of bioremediation technologies ...................................50

4.1 Bioremediation factors............................................... 52
5 Biochemical mechanism of pesticides bioremediation..............52
5.1 Inference ................................................................. 54
References .......................................................................60
CHAPTER 4 Biodegradation and photocatalysis of
pharmaceuticals in wastewater ..............................69
Salman Farissi, Sneha Ramesh,
Muthukumar Muthuchamy and Anbazhagi Muthukumar
1 Introduction .....................................................................69
1.1 Pharmaceuticals........................................................ 70
2 Biodegradation .................................................................71
3 Biodegradation approaches for the treatment of
pharmaceutical wastes........................................................72
3.1 Bacterial degradation................................................. 72
3.2 Fungal degradation.................................................... 73
3.3 Enzymatic degradation............................................... 74
4 Factors affecting the biodegradation of pharmaceuticals...........77
4.1 Photocatalysis .......................................................... 78
4.2 Process enhancement conditions.................................. 84
4.3 Recent advancements and approaches........................... 85
4.4 Photoelectrocatalytic oxidation.................................... 88
5 Conclusion.......................................................................92
References .......................................................................93
CHAPTER 5 Recent trends in the microbial degradation and
bioremediation of emerging pollutants in
wastewater treatment system .................................99
Jayshree Annamalai, Sabeela Beevi Ummalyma and
Ashok Pandey
1 Introduction .....................................................................99
2 Emerging pollutants as micropollutants............................... 100
2.1 Pharmaceuticals...................................................... 101
2.2 Pesticides .............................................................. 102
2.3 Plasticizers ............................................................ 106
2.4 Brominated flame retardants ..................................... 107
2.5 Perfluorinated compounds ........................................ 108
3 Fate of emerging micropollutants in aqueous
environment ................................................................... 109
viii Contents
4 Microbial degradation of micropollutants ............................ 111

4.1 Degradation pathways and metabolic residues of
phthalate esters....................................................... 111
5 Microbial cells and their enzymes in wastewater treatment..... 112
5.1 Bacteria and fungi................................................... 114
5.2 Microalgae ............................................................ 116
6 Perspectives of microbial degradation and challenges ............ 117
7 Advancements in microbial cell-based wastewater treatment .. 117
7.1 Genetic engineering ................................................ 117
7.2 Biogenic nanoparticles............................................. 118
7.3 Integrated systems................................................... 118
8 Conclusion..................................................................... 119
Acknowledgment ............................................................ 119
References ..................................................................... 120
CHAPTER 6 Biological methods for textile dyes removal
from wastewaters................................................. 127
Ashish Kumar Sahoo, Anjali Dahiya and Bhisma K. Patel
1 Introduction ................................................................... 127
1.1 Types of dyes......................................................... 129
1.2 Textile dyes and their impact on the environment......... 131
1.3 Methods for the treatment of textile dyes .................... 134
1.4 Biological treatment ................................................ 134
1.5 Decolorization treatment of dispersed textile dyes ........ 144
1.6 Conclusion and outlook ........................................... 145
References ..................................................................... 146
CHAPTER 7 Importance and applications of biofilm in
microbe-assisted bioremediation .......................... 153
Janhavi Gadkari, Sourish Bhattacharya and
Anupama Shrivastav
1 Introduction ................................................................... 153
2 An overview of biofilm .................................................... 154
2.1 Composition of biofilms........................................... 155
2.2 Mechanism of biofilm formation ............................... 156
2.3 Factors affecting biofilm formation ............................ 157
2.4 Biofilm formed by different microbial species ............. 159
3 An overview of bioremediation.......................................... 161
4 Role of biofilm in bioremediation ...................................... 162
5 Strategies for use of biofilm in bioremediation ..................... 163
Contents ix
6 Types of pollutants remediated by biofilms.......................... 164

7 Application of biofilm in bioremediation............................. 165
7.1 Persistent organic pollutants (POPs)........................... 165
7.2 Inorganic pollutants: heavy metals and synthetic dyes ... 166
7.3 Oil-contaminated water............................................ 167
7.4 Pharmaceutical and personal care products (PPCPs) ..... 168
7.5 Pesticides .............................................................. 168
8 Challenges in biofilm mediated bioremediation .................... 169
References ..................................................................... 170
CHAPTER 8 Microorganism: an ecofriendly tool for waste
management and environmental safety ................. 175
Shubhangi Parmar, Sagar Daki, Sourish Bhattacharya
and Anupama Shrivastav
1 Introduction ................................................................... 175
2 Types of wastes and its sources ......................................... 176
3 Role of microorganisms in waste management ..................... 177
3.1 Sewage treatment.................................................... 178
3.2 Energy production................................................... 178
3.3 Treatment of soil .................................................... 178
3.4 Oil spills treatment.................................................. 179
4 Advantages of bioremediation over conventional methods...... 179
5 Different approaches for microbial waste management .......... 181
5.1 Bioleaching ........................................................... 182
5.2 Bioaugmentation..................................................... 182
5.3 Biostimulation........................................................ 183
5.4 Bioventing............................................................. 183
5.5 Biopiles ................................................................ 184
5.6 Biofiltration ........................................................... 184
5.7 Microbe assisted phytoremediation ............................ 185
6 Treatment of wastewater using microbes ............................. 186
6.1 Bacteria ................................................................ 186
6.2 Algae.................................................................... 186
6.3 Fungi.................................................................... 189
7 Challenges in microbial waste management by
bioremediation................................................................ 189
8 Role of indigenous microorganisms for environmental
protection ...................................................................... 190
9 Conclusion..................................................................... 191
References ..................................................................... 191
x Contents
CHAPTER 9 Microbial degradation of lignin: conversion,

application, and challenges ................................. 195
Aryama Raychaudhuri and Manaswini Behera
1 Introduction ................................................................... 195
2 Chemical structure of lignin and sources ............................. 196
3 Biological degradation of lignin......................................... 198
3.1 Lignin degradation by fungi...................................... 199
3.2 Lignin degradation by bacteria .................................. 201
4 Enzymes associated with lignin degradation ........................ 203
4.1 Lignin peroxidase ................................................... 203
4.2 Manganese peroxidase............................................. 204
4.3 Versatile peroxidase ................................................ 205
4.4 Laccase................................................................. 205
4.5 Bacterial ligninolytic enzymes .................................. 206
5 Regulation of ligninolytic enzymes .................................... 207
5.1 Induction of metal ions ............................................ 207
5.2 Induction of ethanol ................................................ 208
5.3 Induction nutrients .................................................. 208
5.4 Induction of phenolic compounds .............................. 208
6 Bioconversion of lignin to value-added bioproducts .............. 209
6.1 Microbial lipids ...................................................... 209
6.2 Polyhydroxyalkanoates ............................................ 212
6.3 Vanillin ................................................................. 213
6.4 Muconic acid ......................................................... 214
7 Current challenges and future perspectives .......................... 214
8 Conclusion..................................................................... 215
References ..................................................................... 216
CHAPTER 10 Ligninolytic enzymes: a promising tools for
bioremediation of waste water............................ 221
Hiren K. Patel, Rishee K. Kalaria, Divyesh K. Vasava
and Hiren N. Bhalani
1 Introduction.................................................................. 221
2 Ligninolytic enzymes..................................................... 223
2.1 Molecular structure and mechanisms of ligninolytic
enzymes...............................................................223
3 Laccases ...................................................................... 224
3.1 Laccase structure and catalytic mechanism.................224
4 Heme-peroxidases ......................................................... 225
5 Lignin peroxidase (LiP).................................................. 226
Contents xi
6 Manganese peroxidase (MnP).......................................... 227

7 Sources of ligninolytic enzymes....................................... 227
8 Application of ligninolytic enzymes ................................. 228
8.1 Delignification of lignocelluloses ............................229
8.2 Removal of recalcitrant polyaromatic hydrocarbons ...229
8.3 Conversion of coal to low molecular mass fraction ....230
8.4 Biopulping and biobleaching in paper industry..........230
8.5 Polymerization in polymer ventures ........................231
8.6 Biodegradation of colors .......................................231
8.7 Wastewater treatment............................................232
8.8 Soil treatment......................................................233
8.9 Lignolytic enzymes applications in various
industries............................................................234
8.10 Role of ligninolytic enzymes in lignin degradation.....235
9 Improvement strategies for ligninolytic enzyme production .. 236
10 Conclusions.................................................................. 237
References ................................................................... 237
CHAPTER 11 Bioaccumulation and detoxification of heavy
metals: an insight into the mechanism................ 243
Mohita Chugh, Lakhan Kumar, Deepti Bhardwaj and
Navneeta Bharadvaja
1 Introduction.................................................................. 243
2 Industrial effluent containing heavy metals ........................ 244
2.1 Metal finishing ......................................................244
2.2 Mining.................................................................245
2.3 Textiles industries ..................................................245
2.4 Nuclear plants .......................................................245
3 Heavy metals in industrial effluent ................................... 245
3.1 Arsenic ................................................................245
3.2 Lead....................................................................246
3.3 Mercury ...............................................................246
3.4 Cadmium .............................................................246
4 Analysis of heavy metals................................................ 246
5 Heavy metal toxicity...................................................... 247
5.1 Environment .........................................................247
5.2 Plants ..................................................................248
5.3 Microorganisms.....................................................248
5.4 Humans................................................................249
6 Bioremediation ............................................................. 250
7 Bioaccumulation ........................................................... 252
xii Contents
8 Detoxification............................................................... 253
9 Different types of bioremediators for heavy metals ............. 256
9.1 Algae...................................................................257
9.2 Fungi...................................................................258
9.3 Bacteria ...............................................................258
10 Integrated system .......................................................... 259
11 Conclusion and future perspectives................................... 260
References ................................................................... 261
CHAPTER 12 Membrane proteins mediated microbial-
electrochemical remediation technology............. 265
Jesús Pérez-Garcı´a, Javier Bacame-Valenzuela,
Diana Mayra Sánchez López,
José de Jesús Gómez-Guzmán,
Martha Leticia Jiménez González,
Luis Ortiz-Frade and Yolanda Reyes-Vidal
1 Introduction ................................................................. 265
2 Microbial electrochemistry ............................................. 267
2.1 Microbial-electrochemical systems for
bioremediation ..................................................... 268
3 Membrane protein complex in electrogenic bacteria for
bioremediation ............................................................. 270
3.1 Respiratory complexes of Shewanella oneidensis and
heavy metals biodegradation................................... 270
3.2 Redox mediators of Pseudomonas aeruginosa in
environmental bioremediation ................................. 272
3.3 Geobacter sulfurreducens cytochromes and
nanowires in heavy metals reduction ........................ 272
4 Biological enzymes for environmental bioremediation......... 273
4.1 Oxidoreductases ................................................... 274
4.2 Peroxidases.......................................................... 274
4.3 Oxygenases.......................................................... 275
4.4 Monooxygenases .................................................. 275
4.5 Methane oxygenase (MMO) ................................... 276
4.6 Laccases.............................................................. 276
5 Electrochemical characterization of redox enzymes ............ 277
5.1 Cyclic voltammetry............................................... 277
5.2 Electrochemical impedance spectroscopy (EIS).......... 278
5.3 Coupled spectroscopic and electrochemical
techniques ........................................................... 279
6 Perspectives ................................................................. 281
References ................................................................... 282
Contents xiii
CHAPTER 13 Bioremediation strategies to overcome heavy

metals and radionuclides from the
environment....................................................... 287
Sanchayita Basu, Pujaita Banerjee, Sudeshna Banerjee,
Bhaswati Ghosh, Arunima Bhattacharjee, Dipanjan Roy,
Pragati Singh and Ashutosh Kumar
1 Introduction ................................................................. 287
2 Microbial interactions with radionuclides and heavy metals . 289
3 Organisms involved in bioremediation.............................. 292
4 Bioremediation of heavy metals and radionuclides ............. 295
5 Limitations and future prospects...................................... 297
6 Conclusion .................................................................. 298
References ................................................................... 298
CHAPTER 14 Microbial remediation of tannery wastewater...... 303
Lakhan Kumar, Khushbu, Mohita Chugh and
Navneeta Bharadvaja
1 Introduction to tanneries ................................................ 303
2 Characteristics of tannery waste water.............................. 305
3 Environmental and health impacts ................................... 305
4 Wastewater treatment methods adopted in tanneries............ 308
4.1 Physicochemical remediation of tannery wastewater
constituents.......................................................... 310
4.2 Biological treatment or bioremediation of tannery
wastewater........................................................... 312
5 Microbial remediation ................................................... 312
5.1 Bacterial remediation of tannery effluent
constituents.......................................................... 313
5.2 Phycoremediation or algal remediation of tannery
wastewater constituents.......................................... 317
5.3 Fungal remediation or mycoremediation of tannery
wastewater constituents.......................................... 317
6 Challenges and limitations to biological wastewater
treatment methods employed in tannery industries.............. 320
7 Recent advancements .................................................... 321
7.1 Metagenomic approach for bioprospecting potential
microbes and enzymes for tannery wastewater
treatment............................................................. 321
7.2 Microbial biosensors for detection and monitoring of
contaminants present in tannery wastewater............... 322
8 Solid waste management practices................................... 323
References ................................................................... 323
xiv Contents
CHAPTER 15 Biological methods for degradation of textile

dyes from textile effluent .................................... 329
Muhammad Shoaib, Ambreen Ashar,
Zeeshan Ahmad Bhutta, Iqra Muzammil,
Moazam Ali and Ayesha Kanwal
1 Introduction ................................................................. 329
2 Types and characteristics of dyes..................................... 332
3 Methods of dye removal ................................................ 333
3.1 Physicochemical methods....................................... 333
3.2 Biological methods ............................................... 334
4 Conclusion .................................................................. 350
References................................................................... 351
CHAPTER 16 Biodegradation of azo dye using microbiological
consortium ........................................................ 355
Chitra Devi Venkatachalam, Mothil Sengottian,
Sathish Raam Ravichandran and
Sivakumar Venkatachalam
1 Introduction ................................................................. 355
2 Azo dyes in textile industry ............................................ 356
2.1 Classification of azo dyes....................................... 356
2.2 Impact of textile effluents containing azo dyes on
environment......................................................... 357
2.3 Different methods used in degradation of azo dyes ..... 359
3 Microbiological degradation of azo dyes........................... 359
3.1 Degradation mechanism with bacteria ...................... 360
3.2 Degradation mechanism with algae.......................... 362
3.3 Degradation mechanism with fungi.......................... 362
3.4 Advantages of using microbiological consortia .......... 363
4 Parameters involved during microbial azo-dye degradation .. 364
4.1 Effect of carbon source.......................................... 364
4.2 Effect of nitrogen source........................................ 364
4.3 Effect of dye concentration..................................... 366
4.4 Effect of inoculum size.......................................... 366
4.5 Effect of pH......................................................... 366
4.6 Effect of temperature............................................. 366
4.7 Effect of time....................................................... 367
4.8 Effect of agitation ................................................. 367
5 Conclusion .................................................................. 367
References................................................................... 368
Contents xv
CHAPTER 17 Removal of pesticides from water and waste

water by microbes ............................................. 371
Pinal Bhatt and Anupama Shrivastav
1 Introduction ................................................................. 371
2 Pesticide...................................................................... 372
2.1 Organochloride pesticide........................................ 374
2.2 Organophosphate pesticide ..................................... 375
2.3 Carbamate pesticide .............................................. 375
2.4 Other classes........................................................ 375
3 Impact of pesticide........................................................ 375
4 Metabolism and degradation of pesticide .......................... 378
4.1 Application of adsorbent ........................................ 378
5 Biodegradation ............................................................. 382
5.1 Types of pesticides-degrading microorganism ............ 384
5.2 The mechanism of microbial degradation of
pesticides ............................................................ 384
5.3 Commonly used pesticide degradation of
microorganisms .................................................... 388
5.4 Microbial degradation of pesticide technology ........... 388
6 Factors affect biodegradation .......................................... 389
6.1 Environmental factors............................................ 390
6.2 Effect of pesticide structure .................................... 391
6.3 The impact of microorganisms ................................ 392
7 Current scenario ........................................................... 392
8 Conclusion .................................................................. 394
References................................................................... 395
CHAPTER 18 An ecofriendly approach toward waste
management and environmental safety through
microorganisms ................................................. 401
Kunwali Das, Suraj Chetri, Priya Khadgawat,
Sidak Minocha, Aveepsa Sengupta,
Bipin Kumar Sharma and Ashutosh Kumar
1 Introduction ................................................................. 401
2 Microorganisms in the environment ................................. 403
2.1 Bacteria .............................................................. 403
2.2 Fungi.................................................................. 404
2.3 Viruses................................................................ 404
2.4 Protozoa.............................................................. 405
2.5 Algae.................................................................. 405
2.6 Archaea .............................................................. 406
xvi Contents
3 Microorganisms for waste management ............................ 406

3.1 Industrial waste .................................................... 406
3.2 Municipal waste ................................................... 408
3.3 Agricultural wastes ............................................... 409
3.4 Biomedical waste.................................................. 409
3.5 Radioactive waste ................................................. 410
4 Microorganisms in environmental safety........................... 410
5 Conclusion .................................................................. 413
References................................................................... 414
CHAPTER 19 Enzymatic decolorization and degradation of
azo dyes ............................................................ 419
Devikaben Bharatbhai Vishani and
Anupama Shrivastav
1 Introduction................................................................. 419
2 Dyes........................................................................... 420
3 Azo dye ...................................................................... 420
4 Classification of azo dyes............................................... 421
4.1 Acid dyes ............................................................ 421
4.2 Basic or cationic dyes............................................ 421
4.3 Direct dyes .......................................................... 421
4.4 Mordant dyes ....................................................... 421
4.5 Vat dyes .............................................................. 422
4.6 Azoic dyes........................................................... 422
4.7 Reactive dyes....................................................... 422
4.8 Disperse dyes....................................................... 422
4.9 Solvent dyes ........................................................ 423
5 Strucutre of dyes .......................................................... 423
6 Different method for the removal of dyes ......................... 423
6.1 Degradation methods of dyes.................................. 424
6.2 Biologycal method................................................ 426
7 Decolorization and degradation of azo dyes by
azoreductase ................................................................ 428
7.1 Decolorization and degradation azo dyes by laccase ... 428
8 Factor affecting dyes degradation by biological method ...... 429
8.1 pH...................................................................... 429
8.2 Temperature......................................................... 429
8.3 Oxygen ............................................................... 430
9 Mechanism of azo dyes ................................................. 430
10 Conclusion .................................................................. 430
Further reading............................................................. 431
Contents xvii
CHAPTER 20 Azo dyes: a notorious class of water pollutant,

and role of enzymes to decolorize and
degrade them..................................................... 433
Vivek Chauhan, Priya Gautam and
Shamsher S. Kanwar
1 Introduction ................................................................. 433
2 Enzyme-meditated decolorization and degradation of
azo dye ....................................................................... 434
3 Mechanism of degradation and decolorization by
peroxidases.................................................................. 436
3.1 Manganese peroxidase........................................... 437
3.2 Lignin peroxidase ................................................. 438
3.3 Horseradish peroxidase .......................................... 438
4 Mechanism of degradation and decolorization by laccase .... 439
5 Mechanism of degradation and decolorization by
azoreductases ............................................................... 441
6 Conclusion .................................................................. 444
Acknowledgments......................................................... 445
References................................................................... 445
CHAPTER 21 Biofilm mediated bioremediation and other
applications....................................................... 449
Rajalakshmi Sridharan and
Veena Gayathri Krishnaswamy
1 Introduction.................................................................. 449
2 Biofilms in bioremediation.............................................. 450
3 Bioreactors in biofilm formation ...................................... 451
4 Biofilm mediated remediation.......................................... 452
5 Marine biofilms............................................................. 453
6 Marine biofilm in elimination of plastic debris ................... 454
7 Factors affecting the remediation using biofilm................... 455
7.1 Nature of matrix ....................................................455
7.2 pH.......................................................................455
7.3 Temperature..........................................................456
8 Qs in pollutant degradation ............................................. 456
9 Biofilms as source for value added products....................... 456
10 Conclusion................................................................... 456
References................................................................... 457
Index...................................................................................................461
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Contributors
Kunwali Das
Department of Microbiology, Tripura University (A Central University), Agartala,
Tripura, India
Moazam Ali
Department of Clinical Medicine and Surgery, University of Agriculture,
Faisalabad, Punjab, Pakistan
Jayshree Annamalai
Centre for Environmental Studies, Department of Civil Engineering, Anna
University, CEG Campus, Chennai, Tamil Nadu, India
Ambreen Ashar
Department of Chemistry, Government College Women University, Faisalabad,
Punjab, Pakistan
Javier Bacame-Valenzuela
Center of Research and Technological Development in Electrochemistry,
Querétaro Technology Park, Pedro Escobedo, Querétaro, México; CONACYT-
Querétaro Technology Park, Pedro Escobedo, Querétaro, México
Pujaita Banerjee
Department of Life Sciences, Presidency University, Kolkata, West Bengal, India
Sudeshna Banerjee
Department of Microbiology, Amity University, Lucknow, Uttar Pradesh, India
Sanchayita Basu
Tripura, India
Manaswini Behera
School of Infrastructure, Indian Institute of Technology Bhubaneswar,
Bhubaneswar, Odisha, India
Hiren N. Bhalani
College of Agriculture, Junagadh Agricultural University, Junagadh, Gujarat,
India
Navneeta Bharadvaja
Plant Biotechnology Laboratory, Department of Biotechnology, Delhi
Technological University, New Delhi, Delhi, India
Devikaben Bharatbhai Vishani
Department of Microbiology, Parul Institute of Applied Sciences, Parul University,
Vadodara, Gujarat, India
xix
xx Contributors
Deepti Bhardwaj
Pinal Bhatt
Joyani Bhattacharjee
Department of Biotechnology, Haldia Institute of Technology, Haldia, West
Bengal, India
Arunima Bhattacharjee
Tripura, India
Sourish Bhattacharya
Process Design and Engineering Cell, CSIR Central Salt and Marine Chemicals
Research Institute, Bhavnagar, Gujarat, India
Zeeshan Ahmad Bhutta
The Royal (Dick) School of Veterinary Studies, University of Edinburgh, Easter
Bush Campus, Midlothian, Scotland, United Kingdom
Pitam Chakrabarti
Bengal, India
Vivek Chauhan
Department of Biotechnology, Himachal Pradesh University, Summer Hill,
Himachal Pradesh, India
Suraj Chetri
Department of Zoology, Cotton University, Guwahati, Assam, India
Mohita Chugh
Anjali Dahiya
Department of Chemistry, Indian Institute of Technology Guwahati, Guwahati,
Assam, India
Sagar Daki
José de Jesús Gómez-Guzmán
Contributors xxi
Subhasish Dutta
Bengal, India
Salman Farissi
Department of Environmental Science, School of Earth Science Systems, Central
University of Kerala, Kasaragod, Kerala, India
Janhavi Gadkari
Priya Gautam
Bhaswati Ghosh
Department of Microbiology, Sarsuna College, University of Calcutta, Kolkata,
West Bengal, India
Martha Leticia Jiménez González
Rishee K. Kalaria
ASPEE Shakilam Biotechnology Institute, Navsari Agricultural University, Surat,
Gujarat, India
Ayesha Kanwal
Institute of Biochemistry, Biotechnology and Bioinformatics, The Islamia
University of Bahawalpur, Bahawalpur, Punjab, Pakistan
Shamsher S. Kanwar
Priya Khadgawat
Department of Genetics, University of Delhi, New Delhi, Delhi, India
Khushbu
Department of Applied Chemistry, Delhi Technological University, New Delhi,
Delhi, India
Veena Gayathri Krishnaswamy
Department of Biotechnology Stella Maris College (Autonomous), Affiliated to
University of Madras, Chennai, Tamil Nadu, India
Deepak Kumar
Amity Institute of Biotechnology, Amity University Haryana, Gurugram, Haryana,
India
xxii Contributors
Lakhan Kumar
Ashutosh Kumar
Tripura, India
Sangeeta Kumari
India
Sidak Minocha
Department of Genetics, University of Delhi, New Delhi, Delhi, India
Muthukumar Muthuchamy
Anbazhagi Muthukumar
Iqra Muzammil
Department of Clinical Medicine and Surgery, University of Agriculture,
Faisalabad, Punjab, Pakistan
Luis Ortiz-Frade
Ashok Pandey
CSIR-Indian Institute of Toxicological Research, Lucknow, Uttar Praesh, India
Shubhangi Parmar
Hiren K. Patel
School of Sciences, P P Savani University, Surat, Gujarat, India
Bhisma K. Patel
Assam, India
S.M. Paul Khurana
India
Contributors xxiii
Jesús Pérez-Garcı́a
Sneha Ramesh
Sathish Raam Ravichandran
Department of Chemical Engineering, Kongu Engineering College, Erode, Tamil
Nadu, India
Aryama Raychaudhuri
School of Infrastructure, Indian Institute of Technology Bhubaneswar,
Bhubaneswar, Odisha, India
Yolanda Reyes-Vidal
Querétaro Technology Park, Pedro Escobedo, Querétaro, México; CONACYT-
Dipanjan Roy
Department of Agricultural Biotechnology, Ramkrishna Mission Vivekananda
Educational and Research Institute (RKMVERI), Kolkata, West Bengal, India
Ashish Kumar Sahoo
Assam, India
Mothil Sengottian
Department of Chemical Engineering, Kongu Engineering College, Erode, Tamil
Nadu, India
Aveepsa Sengupta
Tripura, India
Bipin Kumar Sharma
Tripura, India
Muhammad Shoaib
Institute of Microbiology, University of Agriculture, Faisalabad, Punjab, Pakistan;
Key Laboratory of New Animal Drug Project, Gansu Province, Key Laboratory of
Veterinary Pharmaceutical Development, Ministry of Agriculture, Lanzhou
Institute of Husbandry and Pharmaceutical Sciences of CAAS, Lanzhou, China
xxiv Contributors
Anupama Shrivastav
Pragati Singh
Tripura, India
Rajalakshmi Sridharan
Department of Biotechnology Stella Maris College (Autonomous), Affiliated to
University of Madras, Chennai, Tamil Nadu, India
Diana Mayra Sánchez López
Sabeela Beevi Ummalyma
Institute of Bioresources and Sustainable Development (IBSD), An Autonomous
Institute Under Department of Biotechnology, Government of India, Takyelpat,
Imphal, Manipur, India
Divyesh K. Vasava
College of Agriculture, Junagadh Agricultural University, Junagadh, Gujarat,
India
Chitra Devi Venkatachalam
Department of Food Technology, Kongu Engineering College, Erode, Tamil Nadu,
India
Sivakumar Venkatachalam
Department of Chemical Engineering, AC Tech Campus, Anna University,
Chennai, Tamil Nadu, India
CHAPTER
A comparative study
between physicochemical
and biological methods for
effective removal of textile
1
dye from wastewater
Subhasish Dutta, Joyani Bhattacharjee
Department of Biotechnology, Haldia Institute of Technology, Haldia, West Bengal, India
1. Introduction
One of the major sources of pollution in nature is industrial wastewater. Due to high
global development, various chemical agents like dyes, pigments, and other aro-
matic compounds are used in industries such as textile, printing, pharmaceuticals,
and plastics to generate products with better results (Khan and et al., 2020). Various
studies have confirmed that an average sized textile industry consumes around 1.6
million liters of water per day for the production of about 8000 kg of fabric. A
research from World Bank has said around 17%e20% of textile industry water
pollution comes from dyeing and finishing treatments that are applied to the fabric.
The processes included in the main steps of textile industry are resizing, dyeing,
printing and some finishing steps. The finishing steps are softening, cross-linking
and waterproofing and they require a huge amount of water supply (Nemr, 2012;
Bhatia et al., 2017). Large amount of effluents are generated from dyeing and the
finishing processes. Dyeing can be defined as the process of applying colors to
the fabrics which are resistant to the effect of light, water, and soap. Tannin and
lignin are considered examples of coloring agents. For various other processes to
be carried out in the textile industry, a particular mixture is made out of chemicals,
dye stuffs, and water. Once the process is completely done this mixture is released
into various water bodies (Anjaneyulu et al., 2005). The presence of suspended solid
particles, high chemical oxygen demand (COD), synthetic dyes, and heavy metals
such as lead, mercury, and cadmium have been considered to increase the toxicity
of water. These textile effluents alter the color and composition of the water bodies
making it very hazardous for the marine ecosystem (Nguyen and Ruey-Shin, 2013).
Dyes have high thermal and photo stability which helps them to persist for an
extended period of time in the water environment if left untreated. The dark color
imparted by these toxic chemicals reduces the sunlight penetration thus hampering
photosynthesis. They also inhibit the growth and activity of microorganisms.
Development in Wastewater Treatment Research and Processes. https://doi.org/10.1016/B978-0-323-85657-7.00003-1 1

Copyright © 2022 Elsevier Inc. All rights reserved.
2 CHAPTER 1 A comparative study between physicochemical and biological
The effluents generated from these textile industries are not only considered to be
toxic for the aquatic ecosystem but they are also enriched by the presence of various
carcinogenic and mutagenic substances which are harmful to human beings too. For
instance, the very famous azo dye, which is one of the main components of textile
industry, is very much responsible for bladder cancer in humans (Ramachandran
et al., 2013). They also cause severe damage to kidney, brain, reproductive system.
Therefore, in this chapter an effort has been made in discussing the preexisting phys-
iochemical methods to remove toxicity from textile effluents and their inefficiencies
and cost effectiveness, with special focus on biological methods which have been
found beneficial.
2. Types of dyes and their toxicity

One of the most important sources of water pollution globally is the toxic effluents
generated from textile industry (Porter, 1973). The main contaminants are gener-
ally produced from dyes and finishing processes (El Harfi and El Harif, 2017).
Dyes can be defined as complex, unsaturated organic compounds which are soluble
in either oil or water. When dyes are applied to substances, it imparts colors
through a process which changes the crystalline structure of colored substance
(Porter, 1973; Benkhaya et al., 2020). Solubility of different dyes in water also dif-
fers from one another. Dyes are widely used in various industries such as textile,
pharmaceuticals, plastics, cosmetics, and so forth. During the middle of 19th cen-
tury natural dyes were synthesized for the first time. They were synthesized mostly
from natural substances like animals, plants, insects and minerals. These dyes are
generally nonsubstantive and are usually applied on textiles with the help of mor-
dents which have affinity for both the fiber and the coloring substance. In compar-
ison to synthetic dyes, these natural dyes are much less toxic and are also easily
treated by biodegradation. In 1871, the first synthetic dye was produced by Woulfe
who treated indigo, a natural dye with nitric acid to produce picric acid (El Harfi
and El Harif, 2017). Nearly 10,000 different kinds of textile dyes and pigments are
used. 7 105 tons of synthetic dyes are produced globally, annually (Singh and
Arora, 2011; Ramachandran et al., 2013). Dyes can be classified into two cate-
gories, based on presence of chromophoric groups in their chemical structures
and usage or application methods (Nemr, 2012).
A chromophore is an atom or a group of atom who is responsible for production
of color because of its ability to absorb light in a near UV region. Compounds
bearing the chromophores are called chromogen. Examples of some chromophoric
groups are N¼O, -NO2, -N¼N-, -C¼O, (CH-CH)n, C¼S, and -C¼N (Benkhaya
et al., 2020).
Auxochromes can be defined as a group which does not itself act as a chromo-
phore, but whose presence alters the intensity as well as wavelength of absorption.
These groups can be acidic or basic in nature. Examples of some acidic groups are
-OH, -COOH, -SO3H. Examples of some basic groups are -NH2, -NHR, and -NR2
(Benkhaya et al., 2020; Singh and Arora, 2011).
2. Types of dyes and their toxicity 3
Based on the presence of chromophoric groups, dyes can be classified into more
than 20e30 groups. Out of which most important ones are nitro dyes, nitroso dyes,
azo dyes, trimethyl ethane dyes, phthalein dyes, indigo dyes, anthraquinone dyes,
and sulfur dye (Benkhaya et al., 2020; Ramachandran et al., 2013).
2.1 Azo dyes

Of all kinds of synthetic dyes, azo dye derivatives are the most widely used ones
around 70% in the textile industries (Nemr, 2012). It has been found out that nearly
1million azo dyes are produced annually. They have -N¼N-chromophoric group in
their structure. Azo dye can again be classified into disazo, trisazo, monazo, polyazo,
and tetrakisazo based on the number of azo dye linkage present (Benkhaya et al.,
2020). The immense popularity of azo dyes is due to some factors. They are the
variations and adaptations that are needed for the multipurpose use of these dyes
and the high molar extinction of azo dye compounds (El Harfi and El Harif,
2017). Azo dyes are formed in situ when two colorless or slightly colored com-
pounds called napthols react (Benkhaya et al., 2020). These azo dyes when conju-
gated with heterocyclic aromatic groups, complex structures are formed. These
structures help in the expression of various colors in the dye. One of the major
drawbacks of using azo dye in the textile industry is, azo dye on reaction with
any blue-violet range of colors gives a dull shade (Benkhaya et al., 2020). A total
of 896 dyes whose chemical structures are known and they are available in the textile
database. According to the European Annex XVII of reach, there are 22 regulated
aromatic amines and only 426 out of 898 dyes can generate a variety or two of
them (Benkhaya et al., 2020). Azo amino dyes can be considered as a suitable
example for organic compounds consisting of zwitterionic resonance system (Singh
and Ram, 2017). Nowadays synthetic dyes are preferably more used than natural
ones. Azo dyes, despite having such a huge impact in the textile industry, have a
lot of major harmful effects too (Saini, 2017). It was noticed that the nitro group
present in the azo dyes, after breaking down generates obnoxious products such
as 1, 4-phenylenediamine, and o-tolidine, which were proved to be mutagenic in
nature. 3-methoxy-4-aminoazobenzene, an aromatic amine was found to be hepato-
carcinogen in rats and a source of potent mutagen in bacteria (Saini, 2017; Singh and
Ram, 2017). There are few specific cases where the azo dyes after being biodegraded
by microorganisms are producing certain derivatives that are found to be highly
toxic. For example, there is a commercial textile dye, namely Acid Violet 7, which
is toxic enough to cause lipid peroxidation, chromosomal aberrations and inhibition
of enzyme acetylcholinesterase. This dye when biodegraded using Pseudomonas
putida, breaks down into respective metabolites such as 4-aminoacetanilide and
5-acetamido-2-amino-1-hydroxy-3, and 6-naphtalene disulfonic acid, which is far
more toxic than Acid Violet 7 (Nemr, 2012).
Based on application method, textile dyes can be classified into the following.
2.2 Acid dye

Due to interactions between the protons of the fiber and the negative charge of the
dyes due to the presence of sulfonic or carboxylic acid groups, acid dyes are consid-
ered to be highly water soluble anionic dyes. These dyes, compared to direct dyes
and vat dyes, and have poor lightfastness on textile dyes. According to chemical
composition, acid dyes can be classified into azo, anthraquinone, triarylmethane,
nitro, and nitroso compounds (Porter, 1973).
2.3 Basic dye

These are water soluble cationic dyes which give high intensity and bright colors that
are visible even at low concentration. These dyes bind with fibers like wool, poly-
ester which have exclusive anionic sites to bind with the cationic parts of these basic
dyes. Major chemical categories are diazahemicyanine, triarylmethane, cyanine,
hemicyanine, thiazine, oxazine, and acridine (Porter, 1973).
2.4 Direct dye

These are anionic dyes that are water soluble in nature. They contain sulfonic acid
group, which have affinity toward cellulose fibers mainly azo compounds that come
under this group. Main advantage of direct dye over fiber-reactive dyes is that direct
dyes are more resistive in fading out under light (Porter, 1973).
2.5 Vat dye

These are the type of dyes which require oxygen to develop their color. These dyes are
usually insoluble in water but are soluble in alkali reduction, i.e., sodium dithionite in
the presence of sodium hydroxide common compounds are anthraquinone or indi-
goids (Nemr, 2012). These synthetic dyes are highly toxic in nature. So when these
dye effluents from the industries are discharged into the water bodies, it not only
increases pollution but also is very much harmful to all the living beings. Thus, we
discuss few physico-chemical methods and the adversities associated with those pro-
cesses and the need to choose certain biological methods to eradicate the problem of
textile dye toxicity (Singh and Arora, 2011; Ramachandran et al., 2013).
The untreated effluents from the textile industries when dumped in water bodies
cause serious side effects on animals and plants (Natarajan et al., 2018). Thus
various conventional approaches have been taken into consideration to treat the
effluents (Anjaneyulu et al., 2005) (Table 1.1).
3. Physical methods
Physical methods can be defined as a straight forward method to remove toxic efflu-
ents from wastewater by the application of forces such as gravitation, electrical
3. Physical methods 5
Table 1.1 Classification of dyes.

Class of dye Chromophoric group Example
Azo dye Methyl orange
Nitro dye Acid yellow 24
Indigo dye Acid blue71
Anthraquinone dye Reactive blue 4
Pthalein dye Phenolphthalein
Triphenyl Methane dye Malachite green
Nitroso dye Fast green O
Sulfur dye Indophenol
Xanthene dye Rhodamine B
Modified from El Harfi, S., El Harfi, A., 2017. Classifications, properties and applications of textile dyes: a
review. Appl. J. Environ. Eng. Sci. 3 (3), 00000e00003 and Singh, P.K., Ram, L.S., 2017. Bio-removal
of azo dyes: a review. Int. J. Appl. Sci. Biotechnol. 5 (2), 108e126.
attraction, and van der Waal’s force. Of all the three methodologies (physical, chem-
ical, and biological), physical method is the most used one as it is much simpler, effi-
cient, and the least amount of chemicals and biological organisms are used (Vanitha
et al., 2018).
3.1 Adsorption
Of all the available physical methods, adsorption has been proven to be the most effi-
cient and widely used technology to treat waste water effluents (Anjaneyulu et al.,
2005). Adsorption can be defined as a physicochemical mass transfer process where
elements are concentrated on the surface of the adsorbent (solid or liquid). This pro-
cess happens in the interface of either two similar or different states for example,
liquid-liquid, gas-liquid, and solid-liquid (Natarajan et al., 2018). The process of
adsorption has been considered as one of the most feasible equilibrium separation
techniques to remove toxic effluents from water bodies due to its economic effi-
ciency and also due to the production of high quality products (Natarajan et al.,
2018). To perform adsorption, no pretreatment is required. Often, it is used as a post-
treatment for various other conventional methods as the process doesn’t produce
additional toxic matters as a result in the end (Vanitha et al., 2018). The surface
on which the process of adsorption takes place is called adsorbent and the adsorbates
are the molecules that get adhered to the surface of adsorbent. If the process of
adsorption is characterized by low heat generation and is reversible in nature, it is
called physisorption (physical adsorption). If the process is characterized by
high heat generation and is irreversible in nature with chemical force acting
between adsorbate and adsorbent, it is called chemisorptions (chemical adsorption)
(Natarajan et al., 2018).
Factors affecting adsorption: There are certain physicochemical factors associ-
ated with adsorption such as temperature, pH, solution concentration, nature of
adsorbate (molecular structure, size, and weight), surface charge, and time duration
of contact (Vanitha et al., 2018; Saini, 2017).
Due to the rising popularity of the adsorption process, a vast range of adsorbents
are used nowadays. Few of them are chitin, almond peels, fly ashes, baggase, acti-
vated carbon, sugarcane, etc.
3.1.1 Adsorption by clay particles

Clays are low cost raw materials used for making zeolites. Zeolites are used widely
in adsorption, ion exchange and irradiation process. Kaolinite and montmorillonite
are two types of clay minerals, they are widely used as dye adsorbents. Cationic dye,
methylene blue is absorbed by kaolinite at a rate of 16 mg/g min. Whereas, for mont-
morillonite the rate varies from 10 mg/g min in the first 5minto 0.55 mg/g min over
the next hour to 0.07 mg/g min for the final hour. Such difference in adsorption rate
is due to the variance in anionic exchange rate of the clays (Nguyen and Ruey-Shin,
2013).
3.1.2 Adsorption by activated carbon

This is one of the most common techniques of removal of effluents. Efficiency of
activated carbon depends on the nature of carbon particle used and also the kind
of dye effluent it is applied on. For instance, activated carbon is most useful in
adsorbing cationic, mordant and acid dyes (90%). Efficiency of activated carbon
reduces for removal of sulfur, dispersed, and reactive dyes (40%). But this issue
can be solved by using an excess amount of activated carbon (Saini, 2017; Robinson
et al., 2001). Activated carbon particles can be made from anything carbonaceous.
Scientists long used coal as a source, but because coal is a nonrenewable source
of energy, activated carbon is now synthesized from renewable, cost-efficient
3. Physical methods 7
substances like biomass, waste materials, and so forth (Vanitha et al., 2018). Acti-
vated carbon particle comes in different shapes and sizes. Powdered activated carbon
(PAC) is very fine in structure thus they can stay suspended for longer while and
making it difficult to be reused. Adding a little quantity of polyaluminum chloride
can enhance the decolorization rate making it easier to collect the sludge from
settling tank and reuse them. In case of granular activated carbon (GAC), it is easier
to replace the GAC packed bed with fresh amount of carbon particles when
exhausted (Nguyen and Ruey-Shin, 2013).
3.1.3 Adsorption by wood chips

These are best used for the adsorption of acid dyes. But they are not widely used as
these wood chips are usually nonrenewable and are burned down to generate power
after being used as adsorbents (Robinson et al., 2001).
3.1.4 Adsorption by silica gel

It is an effective way of removing basic dyes but it is not used much commercially
due to the huge amount of side reactions associated with it (Robinson et al., 2001).
3.2 Coagulation and flocculation

Coagulation is one of the conventional methods of treating wastewater toxicity. This
method is mostly effective for disperse dyes (Anjaneyulu et al., 2005). To reduce the
number of colored effluents from the water bodies, positively charged coagulants are
formed from hydrolysis of metal salts such as iron, aluminum etc. (Anjaneyulu et al.,
2005). These coagulants join together with the negative dye particles forming
neutral clusters which then become macro enough to get filtered. The coagulating
agents are generally polymers which are synthetic in nature having a high molecular
rate and linear structure. Generally used coagulants are aluminum sulfate, aluminum
chloride, ferric chloride, ferric sulfate, cationic organic polymers, etc. (Anjaneyulu
et al., 2005). During the preparation of the macroflocs, the flocculants are added
slowly. There are certain factors that alter the process of coagulation. There are
chemicals present, pH, temperature, mixing time, and retention speed of the system,
which affects the process of coagulation (Verma et al., 2012). Electro coagulation is
the electro chemical treatment of textile dyes, where electrolytic reactions are per-
formed followed by the process of coagulation and sedimentation. This treatment
has been proven to be extremely effective even at higher pH. This method has
been most effective for direct dye removal from wastewater (Verma et al., 2012).
From the research conducted by Yang et al. it was concluded that Al-coagulation
is much more effective for the removal of direct, disperse and reactive dyes. During
the process, OH is formed in the cathode which in turn increases the solution pH,
thus restoring the solution color and dissolving the coagulants. Whereas
Fe-coagulation method is used for color removal from effluents while forming
NaCIO, a strong oxidizing agent as a byproduct which helps in further decomposi-
tion of the dye structure (Yang and McGarrahan, 2005).
3.2.1 Irradiation
This is the simplest yet effective technique to remove textile dye effluents from
waste water. This process requires a large amount of UV radiation and constant dis-
solved oxygen supply. Titanium dioxide acts as a catalyst in the process. On treating
a secondary effluent with irradiation of gamma ray it was found that there was a
reduction of 64% COD, 34% TOC, and 88% of color (Anjaneyulu et al., 2005;
Robinson et al., 2001).
3.3 Emerging physical method for the treatment of textile dye

effluents
3.3.1 Biosorption
Various physical methods have been chosen for the removal of textile dye effluents
from waste water. But most of these methods are cost effective and are meant for low
scale in situ treatment. Biosorption can be defined as the process of accumulating
toxic effluents with the help of microorganisms. Depending on the kind of textile
dye, the rate of binding of the microorganism differs too (Robinson et al., 2001).
On the experiment conducted by Sulak et al. to study the biosorption of aqueous
textile effluents by wheat bran, it was inferred that wheat bran was effective in
removing effluents of certain textile dyes such as are Reactive Red 180, Reactive
black5, Reactiveorange16, Directred80, Acid red, Acid yellow 199 from the aqueous
solution. The percentage of dye removal was found to be directly proportional to the
amount of biosorbent and textile dyes present in the solution. Thermodynamically,
the process was noted to be spontaneous and exothermic. On plotting the biosorption
data in the Langmuir isotherm, adsorption was found to be monolayer (Sulak and
Cengiz Yatmaz, 2012).
4. Membrane technology
Membrane technology can be considered as one of the most effective and cost-
efficient methods which not only help in decolorization but also in reduction of
BOD and COD of wastewater. The main advantage of membrane technology is
that the drench can be further reused. This kind of technology is immensely useful
in places where there is shortage of water (Buckley, 1992). On the basis of classifi-
cation of dye and degree of separation of effluents, this process can be classified into
four types.
4.1 Microfiltration
This is one of the oldest yet most capable ways of removing suspended particles hav-
ing aperture approximately 0.1e1 mm. Through this process of microfiltration wide
range of contaminants such as suspended particles, yeast cells, large pathogens, and
so forth can be separated. Microfiltration is usually performed prior to reverse
osmosis and nanofiltration (Buckley, 1992; Anis et al., 2019).
5. Chemical methods 9
4.2 Reverse osmosis

This method is effective in both wastewater and desalination treatments. It can also
remove various organic contaminants and harmful pathogens. RO is considered to
be a pressure driven process where a semipermeable membrane reject particles
based on size, charge, and interaction between the solute and solvent. RO membranes
have nearly 90% retention rate when it comes to ionic compounds. The osmotic pres-
sure involved in the process is directly proportional to the amount of salts dissolved
therefore to the energy required (Xu and Lebrun, 1999; Malaeb and Ayoub, 2011).
4.3 Nanofiltration
The process of nanofiltration also known as charged filtration is performed after
adsorption to minimize the polarization concentration happening during the filtra-
tion process. It has the advantage of both reverse osmosis and ultrafiltration (UF),
i.e., application of moderate pressure like UF and separations of solutions like
reverse osmosis. Commercially available nanofiltration membrane have been used
to treat cotton textile dye effluents (Buckley, 1992; Xu and Lebrun, 1999).
4.4 Ultrafiltration
UF can be considered as a one-step removal of secondary effluents. UF can be
considered as an RO pretreatment. In many cases UF membranes have been found
using hollow fiber geometry (Xu and Lebrun, 1999).
From the experiment conducted by Paz’dzior et al. for the removal of azo dye by
the amalgamation of nanofiltration with biological methods, it was concluded that
almost 90% color removal was evident and pure colorless filtrate was obtained hav-
ing adequate quantity of salt and alkali concentration. Due to the formation of cleav-
age of Reactive Red 120, orthanilic acid was released. In an aerobic reactor, this acid
was further degraded whereas in the sequencing batch reactor, the aromatic amine
was not degenerated using the microbial culture (Pazdzior et al., 2009).
_
Zy11a et al. performed an experiment with real wastewater collected from two
textile plants. On applying nanofiltration on these textile effluents it was found
that there was significant deterioration in the COD and textile colors. The further
concentrated product in the nanofiltration was made to undergo anoxic biological
treatment. It was observed that there was approximately 50% reduction in the
COD. It was also inferred that the filtrate received from nanofiltration could be
_
used as process water for the rest of the process (Zy11a et al., 2006).
5. Chemical methods
Chemical methods of dye removal can be defined as a collection of certain conven-
tional methods performed by utilizing chemical theories to remove textile dye
effluents. This method requires specific set of equipments and high electrical energy
to work efficiently. Usually chemical technologies have been proven to have success
rate of 88%e99% (Vanitha et al., 2018).
5.1 Fenton method

H2O2eFe (II) is known as Fenton’s reagent. It is very useful for the removal of toxic
effluents from wastewater which normally resists the biological or physical degrada-
tion due to their toxicity to living biomass. CI Reactive Yellow 15 has high COD and
resists degradation. But on reaction with Fenton’s reagent it was found to have
decolorization of 98% and approximately 93% removal of COD. Fenton’s reagent
in association with oxidation process was reported to be effective in degradation
of approximately 20 different classes of dyes namely acid, reactive, direct, cationic,
disperse, and vat in their aqueous solutions. Whereas color removal was found to be
around 80%e100% for acid, reactive, cationic dyes and for water insoluble dyes like
vat and disperse dye it was found to be as low as 30%e60%. Electro-Fenton process
can be defined as an indirect electrochemical treatment used to degenerate and
decolorize toxic textile effluents in this process, hydroxyl radical which is formed
by Fenton’s reagent is produced electrochemically in situ. Hydroxyl radical, being
excellent oxidizing agent can oxidize organic effluents until complete mineraliza-
tion (Robinson et al., 2001; Lahkimi et al., 2007; Singh and Arora, 2011).
Panizza et al. in his experiment studied the electrochemical treatment of
real wastewater containing naphthalene and anthroquinone-sulfonic acid. The pro-
cedure was tried by the electro coagulation of Fenton’s reagent using graphite cath-
odes. To find out the optimum operating conditions, he also analyzed, the effect of
cathode potential, Fe2þ concentration and cathode surface pretreatment (Panizza
and Cerisola, 2001).
5.2 Ozonation
This is one of the most effective chemical processes for nearly complete removal of
textile dye effluents from waste water. Ozone is an excellent oxidizing agent
compared to chlorine, hydrogen peroxide, and other oxidizing agents due to its
high instability. The proportion of ozone to be used in the process is directly propor-
tional to the amount of color and residual COD to be removed. The end product of
ozonation can be easily discharged in the water bodies easily due to the lesser
amount of color and COD present. Ozone can be applied in gaseous state thus vol-
ume of the sludge is not increased (Vanitha et al., 2018). In an experiment conducted
by Lin et al., three strengths of waste water effluent were collected low, medium and
high. The COD and color content of the effluents were observed to intensify from
low to high in the three containers. For low strength, it was inferred that only ozon-
ation was sufficient for decolorization and to remove turbidity. But for medium and
high strength, ozonation was found to be effective for the removal of color but not
5. Chemical methods 11
for turbidity reduction. Thus, coagulation with aluminum sulfate or PAC was neces-
sary. Even with the amalgamation of chemical coagulation with ozonation, approx-
imately 70% COD could be reduced (Lin and Lin., 1993).
5.3 Cucurbitiril
These are cyclic oligomer molecules made up of glycoluril and formaldehyde.
Glycoluril is formed from glyoxal and urea. The monomers are held together by
methylene bridges. The origin of the name cucurbitiril is from the Latin word
Cucurbita maxima which is the scientific name for pumpkin, for its pumpkin like
structure. The uril part of cucurbitiril emphasizes on the presence of urea. It was
inferred that cucurbitiril has extremely good sorption capacity for several textile
dyes. It also forms host-guest relation with aromatic compounds thus removing
the toxicity for adsorption. Another mechanism is based on formation of insoluble
cucurbitiril dye-cation aggregate as adsorption occurs at a faster rate (Robinson
et al., 2001; Nagy et al., 2009) (Fig. 1.1).
5.4 Sodium hypochlorite

The process of bleaching can be defined as a method to remove the colors from
textile materials by chemically destroying the chromophores present thus increasing
FIGURE 1.1
Structure of cucurbitiril (https://pubchem.ncbi.nlm.nih.gov/compound/Cucurbit_8_uril).
the brightness of the product. The chloride molecule attacks the amino group of the
dye, thus accelerating an azo bonding cleavage. This method is not applied for azo
dyes. In an experiment conducted by Massoudinejad et al. it was concluded that
combining chemical oxidation method with sodium hypochlorite solution gives rele-
vant standard to biological method. It was also noticed that imported chorine was
about 3.65 times more efficient than Iranian native chlorine (Robinson et al.,
2001; Massoudinejad et al., 2015).
5.4.1 Ion exchange

This is not much widely used process due to the belief that ion exchangers fail to
accumulate huge amount of dye particles for removal. Advantage of this method
is the no loss of adsorbent molecules (Robinson et al., 2001). Wastewater containing
dyes are made to run over the ion-exchange resins until the favorable exchange sites
are obtained. One of the major disadvantages of this method is its cost. Organic sol-
vents are costly and disperse dyes cannot be removed using ion exchange method
(Anjaneyulu et al., 2005).
5.5 Recent biochemical trend

These physicochemical methods are highly applicable to the treatment of textile dye
effluents in waste water. But these treatments have certain drawbacks too. For
example, intoxicating sludge formation, inefficiency in large scale degradation,
too costly to be used for large scale. Thus few recent biochemical trends have
been discussed which were found to be effective.
5.5.1 Photocatalysis
Photo catalysis can be defined as a process by which a photo reaction is accelerated
in the presence of catalyst. This method degrades the dye molecule into water and
carbon dioxide by the action of UV rays in the presence of catalyst H2O2. Depending
on the product to be catalyzed and the length of the reaction, various byproducts are
formed. No sludge production is associated with the process. UV light on degrading
reaction with H2O2, forms two hydroxyl radicals which help in the further chemical
processes (Robinson et al., 2001).
Photo catalytic removals of organic pollutants have been found to be one of the
most effective ways. Ag/Cl can be considered as high performing photo catalysts
that can be used for degradation. Zhao et al. in his experiment successfully prepared
Ag/AgCl nanoparticles biochemically from the metabolin of living fungi. These
nanoparticles were found to be 3e5 nm in size, spherical in shape. On studying these
particles under microscope it was found that these particles show visible light driven
photo catalytic performance. This biochemical invention helped in the degradation
of a carcinogenic RhB (Zhao et al., 2015).
6. Advanced oxidation process 13
6. Advanced oxidation process

Oxidation can be defined as one of the efficient methods for the removal of effluents
chemically. Usually, in this process the oxidizing agents are activated by some
means. Hydrogen peroxide is the most widely used oxidizing agent in this case.
Advanced oxidation processes (AOP) can be defined as a set of emerging chem-
ical procedures which can remove soluble organic effluents from water and soil.
They use hydroxyl radicals which are powerful oxidants and are nonselective in
nature having the power to degrade complex toxic effluents. The complex molecules
are usually degraded completely into water and carbon dioxide. The biggest signif-
icance of AOP is the simplicity of its usage and production of lesser amount of
residuals. Fenton’s method, ozonation, photolysis can also be considered as types
of AOPs (Rauf and Ashraf, 2012).
As hydroxyl radicals are nonselective in nature thus EAOPs can be coupled with
biological processes in pretreatment as well as posttreatment way. Pretreatment is
considered when the effluent to be treated as BOD and COD ratio much lower
than 0.3 and contains toxic substance which can alter the microbial activities.
Thus EAOP is applied to decrease the toxicity level to such a point where biological
methods can be applied. Posttreatment is considered when the effluent is mostly
filled with biodegradable compounds except for some refractory pollutants. In those
cases, cost effective biological method is applied first followed by the application of
EAOP to remove the remaining minute toxicity (Ganzenko et al., 2014).
Azbar et al. performed a comparative experimental study on various oxidation
and chemical treatment methods for effluent and COD removal from polyester
and acetate fiber dyeing effluents. He inferred that AOPs have nearly 85% greater
success rate in comparison with any other chemical methods for the removal of
effluents derived while dyeing polyester and acetate. Among the different AOPs
for example O3, O3/UV, H2O2,/UV, the most efficient one is the combined result
of ozone, UV and peroxide having removed 99% of COD and 96% color (Azbar
et al., 2004).
In recent studies, coupling of AOPs has been considered. In simpler words,
coupling of AOP can be defined as a method of simultaneous application of more
than one AOP in a single step to increase the oxidative strength of the process.
Due to the wide number of oxidant production in a step, maximum number of times
the process has been considered useful. But there can be instances when due to the
excess production of reactive oxygen species (ROS), degradation rate might get
reduced (Dewil et al., 2017).
Recent trend has suggested that sulfate radical AOPs are gaining a lot of popu-
larity. This can be considered as an advanced alternative to OH- radical AOP. Sulfate
radicals have the redox potential from around 2.5e3.1 volt thus making it able to
oxidize a wider range of organics. They are pH independent thus organics of any
pH can be easily oxidized. It has been studied that sulfate radicals are selective in
nature thus can be used properly to oxidize specific functional groups responsible
for toxicity of wastewater (Lutze et al., 2015).
7. Biological methods
Various physicochemical methods such as oxidation, ozonation, membrane technol-
ogy, coagulation flocculation, and AOPs have been administered to reduce the COD
level and to remove the toxic effluents which are released from textile effluents in
various water bodies. But these processes have some limitations such as costing,
equipments and sludge formation in the end. Many of these processes even work
in situ for only small scale effluents. Thus biological methods have been used as
alternating techniques to remove large scale effluents using living microorganisms
in much cost-efficient and simpler way (Table 1.2).
Table 1.2 Factors affecting biological method of dye removal (Rauf and
Ashraf, 2012; Wesenberg et al., 2003).
Serial
no. Factor Affect
1 Oxygen Dye degradation can occur under aerobic and anaerobic
conditions. Under anaerobic condition, reductive enzyme
activity is higher. Under aerobic conditions, carbon sources
like glucose, starch, acetate affect the decolorization process.
2 Ph The enzymatic activity depends on the pH and on the acid-
base behavior of the substrate and amino acid side chain. The
optimum pH for color removal is often neural or slightly
alkaline. The rate of color removal decreases with acidic pH
values.
3 Temperature This is one of the most impactful parameters. With an increase
in temperature, the rate of color removal increases too. The
temperature required for maximum color removal is nearly
around 35 to 45 C, which is also optimum for cell growth.
4 Dye structure The more dye concentration, the more is the amount of
and toxicity. Dye structure varies in different dyes. Dyes with
concentration simple structure and low molecular weight give out higher rate
of color removal and the process is more difficult with dyes of
high molecular weight.
5 Redox Enzyme mediated degradation is a versatile process efficient
mediator process. The redox mediator is required to maintain the redox
equilibrium. The more redox potential, the faster the molecule
is reduced.
7. Biological methods 15
7.1 Degradation with bacteria

Aromatic compounds can be degraded biologically under both aerobic and anaer-
obic conditions. Under aerobic conditions, enzymes like monoazo oxygenase,
diazo oxygenase catalyzes the incorporation of oxygen into the aromatic amines
prior to ring fission. In maximum monoazo oxygenases, the electro donor is either
NADH or NADPH. In presence of azo reductases, few aerobic bacteria are able to
reduce azo compounds and produce aromatic amines. Certain aerobic azo
reductases, for example, K22 and KF46 strains of Pseudomonas species after
undergoing purification and characterization were found to be flavin-free. These
azoreductases could use both NADH and NADPH as their cofactors and cleave
not only the carboxylated substrate, but also the sulfonated structure (Dos et al.,
2007). Reductions of azo dye by bacterial species are usually no specific and
bacterial decolorization is a fast process. A wide range of aerobic and anaerobic
bacteria have been found beneficial for the degradation purpose, they are Bacillus
subtilis, Pseudomonas sp., Escherichia coli, Rhabdobacter sp., Enterococcus sp.,
Staphylococcus sp, Xenophilussp, Corneybateriumsp, Clostridium sp., Micrococ-
cusdermacoccussp, Acinetobacter sp, Geobacillus, Lactobacillus, Rhizobium,
Proteus sp, Morganella sp., Aeromonas sp, Alcaligenes sp., and Klebsiellla sp.
Some of the aerobic bacteria use azo dye as the sole source of carbon and nitrogen
and others only reduce the azo group by special oxygen-tolerant azo reductases. It
has been concluded by various researchers that the complete degradation of the dye
requires coupled aerobic-anaerobic degradation. In anaerobic conditions, the
azo bond experiences cleavage and aromatic amines are formed. In the aerobic
condition, mineralization by nonspecific enzymes through ring cleavage takes
place. Thus the coupled treatment can efficiently remove azo dye from wastewater.
Pseudomonas sp. can degrade a variety of azo dyes such as Red HE7B, Reactive
Blue172, Reactive Red 22, Reactive Red 2, orange I, and II; thus it is the most
widely used bacterial strain (Wesenberg et al., 2003).
Bacterial degradation can be classified broadly in two ways, using single bacte-
rial cell and using mixed bacterial culture. A strain of Pseudomonas entomophila
BS1 was isolated and was used to degrade Reactive Black 5, a kind of azo dye.
On 120 h incubation, 93% degradation was found. On making a consortium of P.
rettgeri strain HSL1 and Pseudomonas sp. SUK1 to treat certain varieties of azo
dye like Reactive Black 5, Reactive orange 16, Direct Red 81, and Disperse Red
78, complete color removal was noticed in microaerophilic, sequential aerobic/
microaerophilic, and microaerophilic/aerobic conditions with minute time differ-
ences (Wesenberg et al., 2003).
7.1.1 Immobilization of cells for the degradation of dyes

Number of times whole bacterial cells have been used to degrade synthetic dye
effluents. There are various bioreactors that have been used for effective continuous
treatment aerobic and anaerobic treatment, they are fixed film bioreactors, packed
bed bioreactors, aerobic suspended bed activated sludge reactor etc. immobilization
can be defined as a process which prevents cell washing outs and also allows higher
cell density to be maintained in the continuous reactor. Immobilization upgrades the
catalytic stability thus increasing the concentration of degradation of textile dyes. It
has been studied that immobilization results in increased uptake of nutrients due to
the availability of nutrients at the solid-liquid interface. On comparing the immobi-
lized cells of Pseudomonas putinda P8 to the free cells P. putinda, the immobilized
cells have been found to be more efficient in degrading catechol. Immobilization
method using biomass has been found to be really beneficial. It is performed under
aseptic conditions within the bioreactors (Puvaneswari et al., 2006).
7.2 Degradation with algal culture

There hasn’t been much advancement in detecting algae for the degradation of
textile effluents. Azo reductases can degrade azo dye. Few species of Oscillatoria
and Chlorella were able to degrade the azo dye to simpler amines (Banat et al.,
1996; Joshi et al., 2004).
7.3 Degradation with yeast

Degradation of textile dye effluents by biosorption has been much facilitated by
yeast. Yeast has a lot of advantages over bacteria and filamentous fungi. Like bac-
teria, they can grow fast and also like filamentous fungi they have the ability to resist
unfavorable environment. Few yeast species have shown promising result in
dye degrading they are Galacto mycesgeotrichum, Saccharomyces cerevisiae and
Trichosporonbeigelii, etc. NCIM-3326 could decolorize various azo dyes such as
Navy blue HER (100%), Red HE7B (85%), Golden yellow 4BD (60%), Green
HE4BD (70%), and Orange HE2R (50%), among which the decolorization rates
of some dyes were not desirable (Khan et al., 2013).
7.4 Degradation with fungi

Ability to reduce azo dyes by fungi is corelated with the formation of exoenzymes
such as peroxidizes and phenoloxidases. Peroxidizes are a kind of hemoproteins
catalyzing reacting in presence of hydrogen peroxide. Strains like Bjerkanderaa-
dusta, Trametes versicolor and Phanerochaetechrysosporium were proved to
degrade all kinds of azo dyes. Lignin peroxidases were found to oxidize both
phenolic and nonphenolic compounds. But, in order to oxidize phenolic compounds,
manganese peroxidases have to convert Mn2þ to Mn3þ. Phenol oxidases can cata-
lyze the oxidation of phenolic and nonphenolic aromatic compounds without
the help of cofactors. Phenol oxidases are of two types, tyrosinases and laccases
(Dos et al., 2007; Imran et al., 2015).
7.5 Degradation with white-rot fungi

White-rot fungi are a group of fungi that are capable of degrading lignin biologically.
The name white-rot is derived from the white appearance of the wood attacked by
7. Biological methods 17
WRF, removal of lignin giving it a bleached appearance. Based on taxonomical clas-

sification, white-rot fungi are mostly basidiomycete. Exceptionally, few ascomy-
cetes are also capable of white-rot decay. WRF are capable of withstanding a
wide range of pH thus being widely capable of degrading toxic pollutants (Asgher
et al., 2008). WRF are capable of degrading xenobiotics, lignin and other dye stuffs
with their extracellular ligninolytic enzyme system. Few of the extracellular
enzymes which helps predominately in biodegradation of dye stuffs are lignin perox-
idase (Lip), Mn peroxidase (MnP) and H2O2 dependent peroxidases. Some of the
major white-rot fungi strains which are capable of delignification and decolorization
are Phanerochaetechrysosporium, Trametes versicolor, and Coriolusversi color. The
ligninolytic enzymes are substrate nonspecific in nature, thus can degrade a huge
variety of toxic effluents including the complex aromatic pollutants (Kapdan
et al., 2000). Initially, to assay the lignolytic activity of WRF, sulfonated polymeric
dyes were used. Eventually, a number of WRF strains were used to decolorize
distinctive synthetic dyes. In comparison with prokaryotes, WRF are superior dye
decolorizers. Even, in comparison with P. chrysosporium, the lignin transforming
actinomycete Streptomyces chromofuscus is a weak decolorizing microbial strain
(Wesenberg et al., 2003).
7.6 Enzyme system of white-rot fungi

Lignin metabolizing enzymes (LMEs) are essential for lignin degradation. Few of
the major LMEs which are involved in the degradation of lignin and xenobiotics
are lignin peroxidase, Manganese peroxidase, laccase and versatile peroxidase.
Other than these, there are few accessory enzymes which are isolated from certain
WRF strains, they are H2O2-forming glyoxal oxidase, aryl alcohol oxidase, oxalate
producing oxalate decarboxylase (ODC), NAD-dependent formate dehydrogenase
(FDH) and P450 monooxygenase. WRF produces LMEs during secondary meta-
bolism as lignin oxidation yields no net energy to the fungus (Wesenberg et al.,
2003; Asgher et al., 2008).
7.6.1 Lignin peroxidase (LiPs)

These are capable of mineralizing a wide range of aromatic compounds. The molec-
ular mass of various white-rot fungi strain varies from 37 to 50 kDa. It has been
noticed that immobilizing Lip has significantly increased its optimum temperature,
thermo stability as well as catalytic properties. Oxidation by Lip depends on the op-
timum molar ratio of H2O2 to pollutant. At lower concentration, H2O2 acts as an acti-
vator of Phanerochaete chrysosporium LiP and at higher concentration it acts as an
inhibitor rapidly deactivating the enzyme (Asgher et al., 2008).
7.6.2 Manganese peroxidase (MnPs)

This is one of the most common peroxidase produced by almost all white-rot fungi
and few litter decomposing fungi. They are extra cellular glycoprotein having iron
protoporphyrin IX prosthetic group with a molecular weight of around 32 and
62.5 kDa. They are secreted in multiple isoforms. MnPs generally oxidize Mn2þ to
Mn3þ which then is stabilized by chelating compound such as oxalic acid which is
also secreted by the fungi itself. The chelated Mn3þ thus formed is highly reactive
and acts as a diffusible redox mediator. Thus, MnPs oxidizes and depolymerizes
lignin and also xenobiotics like nitroaminotoluenes and toxic textile dyes. The
stability of MnP can be enhanced by immobilizing with sodium alginate, gelatin
or chitosan as carriers and glutaraldehyde as cross-linking (Wesenberg et al.,
2003; Asgher et al., 2008).
7.6.3 Laccases
These are N-glycosylated extracellular blue multicopper oxidases having molecular
mass around 58e90 kDa. The optimum pH and temperature vary from around 2 to
10 and 40e65 C respectively. Two strains of laccase isoenzymes LacI and LacII
have been found in Physisporinus rivulosus T241i, Trametestrogii, Cerrena unicolor
137 and Panustigrinus. Laccases along with oxidizing aphenolic and methoxyphe-
nolic acids, it also decarboxylizes them and attacks their methoxy groups (Asgher
et al., 2008).
Small molecule mediators are low molecular weight highly diffusible redox sub-
stances which help in the interaction between lignin and LME. Due to the random
polymer nature of lignin and bulk of LME, the interaction becomes highly problem-
atic thus these mediators come in action. A few examples of native mediators are P.
chrysosporium, Armillaria mellea, and so forth (Wesenberg et al., 2003).
Most of the dye stuffs that are used in the industries are extensively harmful to
any living being. These dye effluents are highly carcinogenic in nature. White-rot
fungi are used for the purpose of degradation of these harmful dye effluents.
White-rot fungi are preferred over prokaryotic cells due to the presence of LME
system which is nonspecific in nature, thus can degrade a wide range of dyes. P.
chrysosporium and Trametes versicolor are the most used ones. There are other use-
ful WRF too, they are Phellinus gilvus. DIchomitussqualens. Irpex flavus, Gano-
derma sp., etc. The degrading mechanism of dyes varies from the structure and
reactivity of different dyes. For instance, the decolorization of Reactive Blue 15
by chrysosporium follows first order kinetics with respect to initial dye concentra-
tion. Here, MnP plays a major role in decolorization (Asgher et al., 2008).
In an experiment conducted by Kapdan et al. he prepared four separate white-rot
fungi cultures to decolorize five kinds of textile dyes. The strains of white-rot fungi
taken were P. chrysosporium MUCL, P. chrysosporium 671.71, Coriolus versicolor
MUCL, and C. versicolor. The dye stuffs taken were Everzol Yellow 4 GL (Reactive-
monoazo), Everzol Red RBN (Reactive-monoazo), Drimaren Orange K-GL
(Reactive-disazo), Everdirect Supra Yellow PG (Direct-disazo) and Everzol
Turquoise Blue G (Reactive-phtalocyanin). He concluded that the culture of P.
chrysosporium or C. versicolor were highly effective in total removal of dye color
effluents. But, the high incubation temperature around 37 C was marked to be disad-
vantage for the process. Because of lower incubation temperature around 28 C
requirement for C. versicolor cultures, C. versicolor MUCL culture seems to be
more suitable than P. chrysosporium culture for practical applications (Kapdan
et al., 2000).
References 19
8. Conclusion
Different textile industries generate different forms of textile dyes. In this research
article, various physicochemical methods, their advantages and limitations, and
various biological processes with their advantages, have been discussed (Bhatia
et al., 2017). The various physicochemical methods were found to be costly and
reactive only when the effluent volume was less. Many of these processes produced
large volumes of sludge in the end thus not being effective in removal of toxicity
(Ramachandran et al., 2013). Due to the ineffectiveness of the conventional
physicochemical processes, biological processes have been discussed which were
considered to be more ecofriendly and effective for the removal of textile dyes
(Ramachandran et al., 2013). Bioremediation methods use naturally occurring
microbes which are economically friendly and even cost-efficient. Processes like
biosorption is an amalgamation of physical and biological method has been
discussed as an emerging technique which can be used to degrade textile effluent
(Bhatia et al., 2017).
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Title: Trouble Times Two
Author: George O. Smith
Illustrator: Raymond
Release date: May 24, 2022 [eBook #68161]
Language: English
Original publication: United States: Street & Smith Publications,

Incorporated, 1945
Credits: Greg Weeks, Mary Meehan and the Online Distributed

Proofreading Team at http://www.pgdp.net
*** START OF THE PROJECT GUTENBERG EBOOK TROUBLE

TIMES TWO ***
Trouble Times Two
By GEORGE O. SMITH
Illustrated by Raymond
[Transcriber's Note: This etext was produced from

Astounding Science-Fiction December 1945.
Extensive research did not uncover any evidence that
the U.S. copyright on this publication was renewed.]
Thomas Lionel Ph.D., M.M. bounded out of bed with a cheerful bit of
off-tune song. He glanced at the calendar and then the clock and he
grinned because life was just too good to be true.
Everything was according to plan. He'd won his first battle. Up to
now it had been touch and go; at last he had established his right to
co-occupy the mind along with the engineer. No longer could the
engineer claim that he was an expensive detriment. He had forced
the engineer into agreeing that his offering, though not directly
productive, was a causative factor in the development of success.
Then to top it all, he retained enough technology to be a necessary
item. He must be permitted to remain if only for a source of
information.
The engineer's trap had been excellent. But the trap had turned and
caught the engineer. Those reams of data on the poltergeist effect
had been the basis for an entirely new science that only a real
physicist could appreciate—and no engineer could hope to thread
his way through them without a research physicist's assistance.
He stood over the chessboard in the living room for a few minutes.
The engineer was not making any great moves. Therefore the
physicist thought that he might best consolidate his position. He
castled to the queen's side, burying his king behind a bulwark of
defenses that would defy a master chess player to penetrate in less
than ten or fifteen moves.
During breakfast, he perused a thin volume of recent publication. He
did not entirely agree with the theories presented; after all, the book
had been written for the express purpose of getting reader's
viewpoints and Thomas knew it. In fact, the book was not too
interesting to Thomas but he knew that the engineer would fume,
fret, and howl at the idea of having a well-thumbed volume of
"Theory of Multi-Resonant Wave Guides" in the library.
Thomas wouldn't look at the engineer's volume, laying on the table
opposite. It was too un-physical. It was un-erudite. It was "Basic
Theory in Micro-Wave Transmission" and the edges of the pages
were loaded with application formulas, diagrams, and working
sketches.
He was near the end of breakfast when the glint of reflected
sunshine arrowed through the window and caught his eye. He
looked, and wondered who was landing on his lawn in a helicopter.
Then he did a double take.
"Helicopter" stemmed from Greek, the "helix" or screw plus the
"opter" a machine. This contrivance did not. It was not operated with
air screws.
It looked like a three-wheeled coupé. It looked like the industrial
designer's dream of the Plan For Tomorrow, excepting those three
wheels. The Plan For Tomorrow should, by all rights, have four
wheels. And, if the thing is going to fly, it should have some sort of
overhead vanes, or wings, or engines, or jets, or even a skyhook.
But there it was, coming down as light as a feather to make a neat
landing on the back lawn.
By the time the door was open, and the passenger stepped to the
ground, Thomas was standing before the little sky car, looking
somewhat dazed at the name:
POLTERGEIST
"Like a dream," said the driver of the sky car.
"It should," said Thomas, covering his ignorance with monosyllabic
agreement.
"Handles well, too. I think we could stand a bit more positivity of
control, though."
"I'll look into it."
"I wish you would. We've got the jump on the whole world with this.
We'd like to keep it. But the thing doesn't answer to the wheel too
solidly."
"Uh-huh."
"The chief engineer said, 'Jim, take that crate over to Lionel and see
if he will beef up the control force a bit.' So here I am."
"O.K., Jim," said Thomas, offering a prayer for the name that had
been given unwittingly. The engineer must have been a busy boy!
"How are you going to get back?"
Jim looked up into the sky. "Jerry is following in the pilot model. He'll
pick me up and we'll go on back thataway."
Jim nodded skyward, and Thomas looked at the growing speck that
must have been the pilot model.
Thomas forgot about the pilot model. What he wanted to know was
the whereabouts of the five tons of equipment that had been an
integral part of this idea. He looked at the model. He wondered
whether the engineer had installed the whole thing, stepping up the
power and using the main part of the power to support the
equipment. That did not seem possible. Any failure would cause the
little sky car to collapse of its own dead weight. Besides there was
not enough room in the little crate to pack all that equipment-
tonnage.
The engineer had achieved the impossible. He had done away with
the main part while retaining the effect.
The pilot model landed. It was not the finished job of the prototype.
The cabin was squarely functional and the landing wheels were not
faired into the hull. The rear end, instead of tapering gently into a
narrow paraboloid of revolution, was a truncated four-sided pyramid.
Jerry did not emerge. He merely tossed the door open and shouted:
"Come on—we ain't got all day!"
Thomas nodded. "I'll call you when I get it fixed."
Call who? the physicist wondered, and then forgot about it. He
wanted desperately to dig into the sky car. He wanted to find out
where the engineer had packed five tons of equipment. He wanted to
see what made the wheels go around. No doubt the thing could be
returned to its owners without calling in the police. The thing was
probably recorded in the precisely kept engineering notebook of the
physicist's alter ego.
The pilot model was not completely out of sight before Thomas had
the power cowls off, and the whole model stripped of its servicing
doors. They had done an excellent job of design; the sky car without
its servicing panels was but a skeleton frame, with every line, every
connection, and every control rod open for easy servicing.
And it was then and there that the physicist understood what the
engineer had been doing.
Instead of the low-voltage high-current supply lines, with their
attendant heavy busbars, thin pipes ran about the sky car. Seamless
aluminum tubing carried the energizing current. Or, rather the space
inside of the tubing carried it. At the generator end, a ten megawatt
microwave generator supplied high power at ultra high frequency. At
the terminus, rectifiers brought the ultra high frequency down to
direct current for operation of the force-field generators.
Thomas nodded. It was not the final tenth of one percent job. It was
not direct current. The diagravitic force was not constant. It operated
only seven tenths of the time, and was turned off and on fifteen or
twenty million million times per second. Nothing short of high-
definition test equipment would ever tell the difference, however.
Gone were the massive electromagnetic deflection field coils. In their
place was a set of seventy kilovolt electrostatic plates.
Missing entirely were the variable-speed motor generators. In their
place was a simple crystalline formation under permanent magnetic
stress. "Artificial radio-activated crystals," muttered Thomas. "Good
for a couple of years."
But the feed lines. The feed lines. The current carrying ability of
space itself—not the metallic conductor—did the trick. Using the ultra
high frequency bands, the busbars had been replaced with
cylindrical wave guides. The depth of penetration was measured in
microns at those frequencies—and as long as the guides were
properly designed, they offered little loss in power. The current went
down the wave guides by virtue of the magnetic fields created
throughout the guides—magnetic fields generated in the space
inside of the tubular guides.
The generator itself was one of the new crystal microwave
generators and the rectifiers at the receiving end were of the same
ilk.
And the five tons of equipment had vanished in a puff of tubular
guides, electrostatic plates, and intermittent operation.
Thomas hit a snag for a moment. The engineer had answered his
challenge. So he'd come up with the answer to the five-ton-per-fifty-
pound answer—and had gone further. Thomas knew that there was
no apparent limit to the maximum power or lift. It merely set a fifty
pound minimum—actually it was 49.87 pounds by measurement—
under which limit no amount of tinkering would produce the effect.
He smiled. There must be something beyond. After all, small stones
moved quietly in natural poltergeist manifestation; they would be
able to reproduce that eventually. But for now, the engineer was
willing to accept the limitation whereas the physicist would not.
He knew now. And he'd leave the sky car until the engineer returned.
Let him beef up the control force. It was his baby.
Thomas put the panels back on the sky car and stood off to admire
it. It was a neat job, just what the public wanted. The urge to get in
and drive was a most compelling one, and Thomas succumbed. He
sat for a moment, inspecting the dashboard until he had the pattern
well set. Then he snapped on the power, took the wheel and pulled
back gently. The sky car lifted its nose slightly, and as Thomas
pressed the foot pedal, it took off on a side-line straight into the sky.
He leveled off at a thousand feet and he did some scurrying back
and forth in midair. It did handle a little sloppy but not enough to
make the physicist uncomfortable. Yet it wouldn't stand any hedge-
hopping or bridge-undercutting without a prayer on the part of the
driver. Butter the controls a bit and you could thread a needle with it
on the first try.
Yes, the engineer had done it again—all of which made Thomas
chuckle. A bit more of this and the engineer would have such an
income that he'd no longer worry himself into engineering. Then—
Thomas turned the sky car and drove across the city toward Dr.
Hamilton's place. He landed on the psychiatrist's lawn and startled
the doctor out of a week's growth.
"I've won," he told the doctor.
"Good," laughed Hamilton. "Mind if I ask which you are today—and
how do you know you've won?"
"I'm Thomas Lionel, Ph.D. And the engineer has worked himself out
of a job."
"Interesting. But how?"
"He dropped me a mess of cockeyed data, remember? Well, I
unraveled it into a most interesting field of science. From it I handed
him a slab full of theories and experiments that are just inefficient
enough to make him fume. He's come up with several things that
make money in vatfuls."
"That, I know and understand. Go on."
"Remember, I am his ideal personality, I am a physicist, a type of
person he has always wanted to be. He couldn't be a physicist
because of financial reasons and so he went into the engineering
field to bolster up his bank account. That was eminently practical.
But now that the worry about the bankroll is over, he can turn to
theoretical physics and physical research. That's me—and I've won!"
"Suppose he, himself, takes the gradual retreat from engineering into
physical research?"
"Um—I don't think he's capable of it. He's been too well conditioned."
"Might well be," admitted the doctor. "Well, as I said before, I'm just a
referee. Both of you are well adjusted and good, worthy additions to
society. Either one of you that wins will be a credit to civilization."
"You're a great help," laughed Thomas. "But I don't mind. This is my
round, and it's my game. He's licked himself."
"I'll tell him that when I see him," said Dr. Hamilton. "But there is one
thing that I must know. I want to know what makes that little tungsten
box work."
"I cast the tungsten in—"
"I don't care how you made it," said Hamilton flatly, "unless it has a
definite bearing on how it works."
"I made it of tungsten because the engineer would rip it apart if it
weren't too tough," grinned Thomas. "Being of tungsten it doesn't
matter how it works excepting it would have been more efficient if I'd
made it of silver."
"Look, Thomas, stay on the subject. I want to know what's with the
works."
Lionel laughed. "What's so important?"
"Look, man, I'm a psychiatrist. The functioning of the human mind is
my baby. Or," he added bitterly, "it should be. But, darn it, all we can
do is to surmise, theorize, hope and pray. We don't know what
makes schizophrenics, or manic-depressives or any of the other
mental quirks. We aren't even certain why some people are well
liked while others, of almost identical get-together are heartily
disliked. But you've come up with a little dingus that causes a switch-
over from one personality to another merely by pushing a button.
Find out why and we psychiatrists may some day get to first base in
psychoanalysis."
"Um—I suppose a real pathophone would be a help."
"Pathophone is a good word," smiled the psychiatrist, "but to dig into
a warped mind without having the erroneous impressions and false
evaluation clouding the only entry ... we'd be able to clear up almost
any mental condition. Now, how does it work?"
"I am not prepared to say. I was seeking experimental data on the
'epicenter' of the poltergeist phenomena—the poltergeist usually
manifests in the vicinity of or because of some central influence—
usually a person who is unaware of his potentiality. At any rate, I was
setting up a series of local magnetic and electrostatic fields and then
trying the micro-microwave spectrum for response. I was running up
through the region between long heat radiation and micro-micro
radio waves when—blooey!—I was the engineer. I switched back
eventually and consolidated my findings into that little tungsten box."
"I want the dope on it."
"I'll give it to you," nodded Thomas. "As soon as I make some final
measurements and consolidate my data."
"Fine. Mind telling me what causes the poltergeist?"
"As best I can. The present concept of space is that space itself is
under internal strain. Force vectors in cancellation prevail, resulting
in a stable continuum. Space is warped by electrostatic effects,
magnetic effects, and gravitic effects. These local effects do not
create a discontinuity in the space strain, and therefore no eruption
takes place. Now enters the epicenter. Radiation from his mind or
brain in thinking goes out and starts a very minor sympathetic
oscillation in the warps and strains of space. If these strains are in
the right vectorial situation, the minor oscillation builds up the
response amplitude—"
"That doesn't make sense," objected the doctor. "Mental radiation
must be weak. How can it induce high power?"
"It can't. But if you know radio at all, you'll recall that a high 'Q' circuit
will develop very high voltages across the terminals with a very small
driving voltage. Well, this is analogous to the epicenter effect. The
epicenter wave causes instability in the space strains because the
brain wave is not a natural phenomena of space. Then—like two
sticks end to end under compression, it takes very little sidewise
thrust to make the compression-force collapse, forcing the sticks out
at right angles. Follow?"
"But where did this energy or force come from?" puzzled Hamilton.
"Isn't that a violation of the Law of Conservation of Energy?"
"Not at all. The law is still valid. It does state that you cannot get
more out of anything than is put into it. The guesswork comes in
deciding how the energy got there. Coal, for instance, is just a black
stone. It has potential energy which was put into it by the eons of
solar energy shining on the carboniferous forests. A stone has
potential energy for falling. Where did it get it? It may have been
carried up the hill; it may have been dropped from space—put out
there by the cosmic eruption that caused Creation. Or it may have
been on the edge of a gully and the potential drop made by the
stream eroding the ground out from under it."
"How about atomic power?"
"You mean, how did the power get locked in the atom?"
"Yes."
"The power in the atom was put there by the universe's atom
factories. Sol, and the other suns," explained Hamilton.
"But where did the earth—?"
"Creation," murmured Thomas. "Who knows? I don't. Every time
somebody comes up with a perfect answer, someone else comes up
with perfect data that proves that the answer couldn't be anything
that anybody has ever used before.
"The atom factory is the Solar Phoenix. You start with hydrogen and
carbon. The solar heat is such that they combine atomically to an
unstable isotope of nitrogen which immediately becomes a stable
isotope of nitrogen. More hydrogen gets in, making it unstable
oxygen and so forth. Oxygen breaks down, releasing energy, helium,
and, what do you know, carbon again, which begins to take on
hydrogen again, and here we go again. But the thing is uncontrolled
hell on wheels. Things go wrong due to the variances of pressure
and temperature, and the oxygen doesn't always break down into
helium and carbon. It takes offshoots and sidetracks. It'll add
hydrogen and become fluorine, for instance, which then adds more
and becomes something else, some of which trails off like the
branches of a tree and do not break down into recurrent reactions.
Hence the other atoms."
"I'll read about it and get the real picture. Know a good book?"
Thomas scratched his chin. "If you can find a copy of 'The Days of
Creation,' by Willy Ley, the first part of the book has a description of
the Solar Phoenix."
"Well, good enough," said Dr. Hamilton. "But just bear one thing in
mind. You think you've beaten the engineer. Your basic trouble is just
that the engineer is you, too. He has your ability and your knowledge
and your experience upon which to work. He is no fool, and you can
take that as a back-handed compliment if you want to. He is just as
capable an engineer as you are a physicist. He thinks in different
channels, I will admit. But, Thomas, remember that his extra-
channellar thinking is done with the same thinking equipment as
yours is, and it is no less efficient because of being divergent from
your own thought-track. Your battle was won too easily to be
conclusive."
"What do you expect?"
"I wouldn't know. I'm no scientist in physics." Hamilton held up a
hand as Thomas started to protest. "I use 'scientist' despite your
dislike of the word only because there is no term that describes both
of the attributes of practical engineer and research physicist. Frankly,
I'm hoping for an eventual coalition, but I fear not."
"Why view no-coalition with distaste?" demanded Thomas.
"Because both personalities offer much to the world, to science in
general, and to the body that houses both of them."
"I heartily dislike all aspects of practical engineering," stated Thomas
flatly. "To be everlastingly forced to retrace your own steps, again
and again and again, working out the most insignificant details—
bah!"
"The engineer has another viewpoint."
"I know. But the engineer in this case is here only because of his
own necessity—which he himself has removed. I am the real entity; I
am the desire of the engineer. I am what he wants to be. I am what
he will become!"
"Good morning, Frank."

"Morning, Miss Elaine. Mr. Lionel isn't here."
"He'll be back?" asked the girl.
"Oh yes. He went over to see Dr. Hamilton."
"Oh. Frank, the usual question?"
"This morning he is Thomas Lionel, Ph.D., M.M."
"Oh."
"He went to bed Tom Lionel, Consulting Engineer."
"I wonder if he remembers," smiled Elaine.
The Poltergeist landed on the lawn. It was silent, but a flash of
sunshine caught the sleek side and attracted Elaine's attention.
"Hi," she called as she emerged from the house.
"Howdy," he answered. "What brings you out?"
"Never ask a girl a question like that," she laughed. "You'll never get
the right answer."
"Why?"
"If she says 'you' it's either a lie or she's the kind of girl your mother
tried to protect you from. If she says anything else, it's either a lie or
she's the kind of girl your mother tried to protect you from."
"A man can't win," snorted Thomas.
"Does a man really want to win?"
"Nope," admitted Thomas. "I won't ask questions, Elaine. I'll just be
glad you came."
"I'm glad you're glad."
Elaine flirted with him shamelessly, and then turned toward the
laboratory building. He followed, and they kept up a running fire of
light talk all the way.
"The first thing I have to do is to see what the engineer was doing
last," remarked Thomas as he opened the laboratory door.
"You are a strange fellow," smiled Elaine. "You respect each other's
possessions and beliefs, though you argue madly through
impersonal mediums. Still writing nasty letters?"
"Uh-huh. And playing chess."
"What's he been doing?" asked Elaine innocently.
"Don't really know. Aside from some experiments on the poltergeist
effect—reducing them to practice—I wouldn't know. I doubt that he's
been doing much else. I do happen to know that he's deeply
interested in the epicenter effect. He may find the key to it, too."

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Development in Wastewater Treatment

Research and Processes: Innovative

Development in Wastewater Treatment Research and

Development in Wastewater Treatment Research and

Development in Wastewater Treatment Research and

An Innovative Role of Biofiltration in Wastewater

Modular Treatment Approach for Drinking Water and

Electrochemical water and wastewater treatment 1st

Current Trends and Future Developments on (Bio-)

Valorization of Microalgal Biomass and Wastewater

Riti Thapar Kapoor

Copyright © 2022 Elsevier Inc. All rights reserved.

Library of Congress Cataloging-in-Publication Data

British Library Cataloguing-in-Publication Data

For information on all Elsevier publications visit our website at

Publisher: Joe Hayton

Typeset by TNQ Technologies

CHAPTER 1 A comparative study between physicochemical and

CHAPTER 2 An approach toward developing clean green

4 Types of bioremediation technologies ...................................50

4 Microbial degradation of micropollutants ............................ 111

6 Types of pollutants remediated by biofilms.......................... 164

CHAPTER 9 Microbial degradation of lignin: conversion,

6 Manganese peroxidase (MnP).......................................... 227

CHAPTER 13 Bioremediation strategies to overcome heavy

CHAPTER 15 Biological methods for degradation of textile

CHAPTER 17 Removal of pesticides from water and waste

3 Microorganisms for waste management ............................ 406

CHAPTER 20 Azo dyes: a notorious class of water pollutant,

Development in Wastewater Treatment Research and Processes. https://doi.org/10.1016/B978-0-323-85657-7.00003-1 1

2. Types of dyes and their toxicity

2.1 Azo dyes

2.2 Acid dye

2.3 Basic dye

2.4 Direct dye

2.5 Vat dye

Table 1.1 Classification of dyes.

Nitro dye Acid yellow 24

Indigo dye Acid blue71

Anthraquinone dye Reactive blue 4

Pthalein dye Phenolphthalein

Triphenyl Methane dye Malachite green

Nitroso dye Fast green O

Sulfur dye Indophenol

Xanthene dye Rhodamine B

3.1.1 Adsorption by clay particles

3.1.2 Adsorption by activated carbon

3.1.3 Adsorption by wood chips

3.1.4 Adsorption by silica gel

3.2 Coagulation and flocculation

3.3 Emerging physical method for the treatment of textile dye

4.2 Reverse osmosis

5.1 Fenton method

5.4 Sodium hypochlorite

5.4.1 Ion exchange

5.5 Recent biochemical trend

6. Advanced oxidation process

7.1 Degradation with bacteria