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Biosurfactants for a Sustainable Future
Biosurfactants for a Sustainable Future
Production and Applications in the Environment

and Biomedicine
Edited by
Hemen Sarma
Department of Botany
Nanda Nath Saika College
Titabar, Assam, India
Majeti Narasimha Vara Prasad

School of Life Sciences
University of Hyderabad (an Institution of Eminence)
Hyderabad, Telangana, India
This edition first published 2021
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Library of Congress Cataloging‐in‐Publication Data

Names: Sarma, Hemen, editor. | Prasad, Majeti Narasimha Vara editor.
Title: Biosurfactants for a sustainable future : production and
applications in the environment and biomedicine / Hemen Sarma,
Nanda Nath Saika College, Department of Botany, 785630, Titabar, India;
Majeti Narasimha Vara Prasad, University of Hyderabad, School of Life Sciences,
500046 Hyderabad, India.
Description: Hoboken, NJ : Wiley, 2021. | Includes bibliographical
references and index.
Identifiers: LCCN 2020051310 (print) | LCCN 2020051311 (ebook) | ISBN
9781119671008 (cloth) | ISBN 9781119671039 (adobe pdf) | ISBN
9781119671053 (epub)
Subjects: LCSH: Biosurfactants.
Classification: LCC TP248.B57 B565 2021 (print) | LCC TP248.B57 (ebook) |
DDC 668/.1–dc23
LC record available at https://lccn.loc.gov/2020051310
LC ebook record available at https://lccn.loc.gov/2020051311
Cover Design: Wiley
Cover Image: @Kennia Barrantes
Set in 9.5/12.5pt STIXTwoText by SPi Global, Pondicherry, India
10 9 8 7 6 5 4 3 2 1
v
Contents
List of Contributors xii

Preface xvii
1 Introduction to Biosurfactants 1
José Vázquez Tato, Julio A. Seijas, M. Pilar Vázquez-Tato, Francisco Meijide,
Santiago de Frutos, Aida Jover, Francisco Fraga, and Victor H. Soto
1.1 Introduction and Historical Perspective 1
1.2 Micelle Formation 5
1.3 Average Aggregation Numbers 14
1.4 Packing Properties of Amphiphiles 18
1.5 Biosurfactants 20
1.6 Sophorolipids 25
1.7 Surfactin 28
1.8 Final Comments 31
Acknowledgement 32
References 32
2 Metagenomics Approach for Selection of Biosurfactant Producing Bacteria

from Oil Contaminated Soil: An Insight Into Its Technology 43
Nazim F. Islam and Hemen Sarma
2.1 Introduction 43
2.2 Metagenomics Application: A State-of-the-Art Technique 44
2.3 Hydrocarbon-Degrading Bacteria and Genes 46
2.4 Metagenomic Approaches in the Selection of Biosurfactant-Producing Microbes 47
2.5 Metagenomics with Stable Isotope Probe (SIP) Techniques 48
2.6 Screening Methods to Identify Features of Biosurfactants 50
2.7 Functional Metagenomics: Challenge and Opportunities 52
2.8 Conclusion 53
Acknowledgements 54
References 54
3 Biosurfactant Production Using Bioreactors from Industrial Byproducts 59

Arun Karnwal
3.1 Introduction 59
3.2 Significance of the Production of Biosurfactants from Industrial Products 60
vi Contents
3.3 actors Affect Biosurfactant Production in Bioreactor

F 61
3.4 Microorganisms 61
3.5 Bacterial Growth Conditions 63
3.6 Substrate for Biosurfactant Production 65
3.7 Conclusions 71
Acknowledgement 71
References 72
4 Biosurfactants for Heavy Metal Remediation and Bioeconomics 79

Shalini Srivastava, Monoj Kumar Mondal, and Shashi Bhushan Agrawal
4.1 Introduction 80
4.2 Concept of Surfactant and Biosurfactant for Heavy Metal Remediation 81
4.3 Mechanisms of Biosurfactant–Metal Interactions 82
4.4 Substrates Used for Biosurfactant Production 82
4.5 Classification of Biosurfactants 85
4.6 Types of Biosurfactants 85
4.7 Factors Influencing Biosurfactants Production 88
4.8 Strategies for Commercial Biosurfactant Production 89
4.9 Application of Biosurfactant for Heavy Metal Remediation 90
4.10 Bioeconomics of Metal Remediation Using Biosurfactants 93
4.11 Conclusion 94
References 94
5 Application of Biosurfactants for Microbial Enhanced Oil Recovery (MEOR) 99

Jéssica Correia, Lígia R. Rodrigues, José A. Teixeira, and Eduardo J. Gudiña
5.1 Energy Demand and Fossil Fuels 99
5.2 Microbial Enhanced Oil Recovery (MEOR) 101
5.3 Mechanisms of Surfactant Flooding 102
5.4 Biosurfactants: An Alternative to Chemical Surfactants to Increase Oil Recovery 103
5.5 Biosurfactant MEOR: Laboratory Studies 104
5.6 Field Assays 112
5.7 Current State of Knowledge, Technological Advances, and Future Perspectives 113
Acknowledgements 114
References 114
6 Biosurfactant Enhanced Sustainable Remediation of Petroleum Contaminated Soil 119

Pooja Singh, Selvan Ravindran, and Yogesh Patil
6.1 Introduction 119
6.2 Microbial-Assisted Bioremediation of Petroleum Contaminated Soil 121
6.3 Hydrocarbon Degradation and Biosurfactants 122
6.4 Soil Washing Using Biosurfactants 124
6.5 Combination Strategies for Efficient Bioremediation 126
6.6 Biosurfactant Mediated Field Trials 129
6.7 Limitations, Strategies, and Considerations of Biosurfactant-Mediated
Petroleum Hydrocarbon Degradation 130
6.8 Conclusion 132
References 133
Contents vii
7 Microbial Surfactants are Next-Generation Biomolecules for Sustainable Remediation

of Polyaromatic Hydrocarbons 139
Punniyakotti Parthipan, Liang Cheng, Aruliah Rajasekar, and Subramania Angaiah
7.2 Biosurfactant-Enhanced Bioremediation of PAHs 144
7.3 Microorganism’s Adaptations to Enhance Bioavailability 151
7.4 Influences of Micellization on Hydrocarbons Access 151
7.5 Accession of PAHs in Soil Texture 152
7.6 The Negative Impact of Surfactant on PAH Degradations 152
7.7 Conclusion and Future Directions 153
References 153
8 Biosurfactants for Enhanced Bioavailability of Micronutrients in Soil:

A Sustainable Approach 159
Siddhartha Narayan Borah, Suparna Sen, and Kannan Pakshirajan
8.2 Micronutrient Deficiency in Soil 161
8.3 Factors Affecting the Bioavailability of Micronutrients 161
8.4 Effect of Micronutrient Deficiency on the Biota 163
8.5 The Role of Surfactants in the Facilitation of Micronutrient Biosorption 166
8.6 Surfactants 166
8.7 Conclusion 173
References 174
9 Biosurfactants: Production and Role in Synthesis of Nanoparticles for

Environmental Applications 183
Ashwini N. Rane, S.J. Geetha, and Sanket J. Joshi
9.1 Nanoparticles 183
9.2 Synthesis of Nanoparticles 184
9.4 Biosurfactant Mediated Nanoparticles Synthesis 191
9.5 Challenges in Environmental Applications of Nanoparticles and
Future Perspectives 196
References 197
10 Green Surfactants: Production, Properties, and Application in Advanced

Medical Technologies 207
Ana María Marqués, Lourdes Pérez, Maribel Farfán, and Aurora Pinazo
10.1 Environmental Pollution and World Health 207
10.2 Amino Acid-Derived Surfactants 208
10.4 Antimicrobial Resistance 219
10.5 Catanionic Vesicles 223
10.6 Biosurfactant Functionalization: A Strategy to Develop Active
Antimicrobial Compounds 234
10.7 Conclusions 235
References 235
viii Contents
11 Antiviral, Antimicrobial, and Antibiofilm Properties of Biosurfactants: Sustainable Use

in Food and Pharmaceuticals 245
Kenia Barrantes, Juan José Araya, Luz Chacón, Rolando Procupez-Schtirbu,
Fernanda Lugo, Gabriel Ibarra, and Víctor H. Soto
11.2 Antimicrobial Properties 246
11.3 Biofilms 252
11.4 Antiviral Properties 255
11.5 Therapeutic and Pharmaceutical Applications of Biosurfactants 256
11.6 Biosurfactants in the Food Industry: Quality of the Food 258
References 261
12 Biosurfactant-Based Antibiofilm Nano Materials 269

Sonam Gupta
12.2 Emerging Biofilm Infections 270
12.3 Challenges and Recent Advancement in Antibiofilm Agent Development 272
12.4 Impact of Extracellular Matrix and Their Virulence Attributes 273
12.5 Role of Indwelling Devices in Emerging Drug Resistance 274
12.6 Role of Physiological Factors (Growth Rate, Biofilm Age, Starvation) 274
12.7 Impact of Efflux Pump in Antibiotic Resistance Development 275
12.8 Nanotechnology-Based Approaches to Combat Biofilm 276
12.9 Biosurfactants: A Promising Candidate to Synthesize Nanomedicines 277
12.10 Synthesis of Nanomaterials 278
12.11 Self-Nanoemulsifying Drug Delivery Systems (SNEDDs) 282
12.12 Biosurfactant-Based Antibiofilm Nanomaterials 283
12.13 Conclusions and Future Prospects 283
Acknowledgement 285
References 285
13 Biosurfactants from Bacteria and Fungi: Perspectives on Advanced

Biomedical Applications 293
Rashmi Rekha Saikia, Suresh Deka, and Hemen Sarma
13.2 Biomedical Applications of Biosurfactants: Recent Developments 295
13.3 Conclusion 307
References 307
14 Biosurfactant-Inspired Control of Methicillin-Resistant

Staphylococcus aureus (MRSA) 317
Amy R. Nava
14.1 Staphylococcus aureus, MRSA, and Multidrug Resistance 317
14.2 Biosurfactant Types Commonly Utilized Against S. aureus and Other Pathogens 318
14.3 Properties of Efficient Biosurfactants Against MRSA and Bacterial Pathogens 319
14.4 Uses for Biosurfactants 320
Contents ix
14.5 iosurfactants Illustrating Antiadhesive Properties against MRSA Biofilms 320

B
14.6 Biosurfactants with Antibiofilm and Antimicrobial Properties 322
14.7 Media, Microbial Source, and Culture Conditions for Antibiofilm
and Antimicrobial Properties 323
14.8 Novel Synergistic Antimicrobial and Antibiofilm Strategies Against MRSA
and S. aureus 326
14.9 Novel Potential Mechanisms of Antimicrobial and Antibiofilm Properties 328
14.10 Conclusion 330
References 332
15 Exploiting the Significance of Biosurfactant for the Treatment of Multidrug-Resistant

Pathogenic Infections 339
Sonam Gupta and Vikas Pruthi
15.2 Microbial Pathogenesis and Biosurfactants 340
15.3 Bio-Removal of Antibiotics Using Probiotics and Biosurfactants Bacteria 342
15.4 Antiproliferative, Antioxidant, and Antibiofilm Potential of Biosurfactant 343
15.5 Wound Healing Potential of Biosurfactants 344
15.6 Conclusion and Future Prospects 345
References 346
16 Biosurfactants Against Drug-Resistant Human and Plant Pathogens:

Recent Advances 353
Chandana Malakar and Suresh Deka
16.2 Environmental Impact of Antibiotics 354
16.3 Pathogenicity of Antibiotic-Resistant Microbes on Human and Plant Health 356
16.4 Role of Biosurfactants in Combating Antibiotic Resistance: Challenges
and Prospects 360
16.5 Conclusion 364
References 365
17 Surfactant- and Biosurfactant-Based Therapeutics: Structure, Properties, and Recent

Developments in Drug Delivery and Therapeutic Applications 373
Anand K. Kondapi
17.2 Determinants and Forms of Surfactants 374
17.3 Structural Forms of Surfactants 377
17.4 Drug Delivery Systems 381
17.5 Different Types of Biosurfactants Used for Drug Delivery 384
References 392
18 The Potential Use of Biosurfactants in Cosmetics and Dermatological Products:

Current Trends and Future Prospects 397
Zarith Asyikin Abdul Aziz, Siti Hamidah Mohd Setapar, Asma Khatoon, and Akil Ahmad
x Contents
18.2 roperties of Biosurfactants 399

P
18.3 Biosurfactant Classifications and Potential Use in Cosmetic Applications 401
18.4 Dermatological Approach of Biosurfactants 406
18.5 Cosmetic Formulation with Biosurfactant 409
18.6 Safety Measurement Taken for Biosurfactant Applications in Dermatology
and Cosmetics 412
18.7 Conclusion and Future Perspective 415
Acknowledgement 415
References 415
19 Cosmeceutical Applications of Biosurfactants: Challenges and Prospects 423

Káren Gercyane Oliveira Bezerra and Leonie Asfora Sarubbo
19.2 Cosmeceutical Properties of Biosurfactants 424
19.3 Other Activities 429
19.4 Application Prospects 432
19.5 Biosurfactants in the Market 433
19.6 Challenges and Conclusion 434
References 436
20 Biotechnologically Derived Bioactive Molecules for Skin and Hair-Care Application 443
Suparna Sen, Siddhartha Narayan Borah, and Suresh Deka
20.2 Surfactants in Cosmetic Formulation 445
20.3 Biosurfactants in Cosmetic Formulations 445
20.4 Conclusion 457
References 457
21 Biosurfactants as Biocontrol Agents Against Mycotoxigenic Fungi 465

Ana I. Rodrigues, Eduardo J. Gudiña, José A. Teixeira, and Lígia R. Rodrigues
21.1 Mycotoxins 465
21.2 Aflatoxins 466
21.3 Deoxynivalenol 467
21.4 Fumonisins 468
21.5 Ochratoxin A 468
21.6 Patulin 470
21.7 Zearalenone 470
21.8 Prevention and Control of Mycotoxins 471
21.10 Glycolipids 473
21.11 Lipopeptides 474
21.12 Antifungal Activity of Glycolipid Biosurfactants 474
21.13 Antifungal and Antimycotoxigenic Activity of Lipopeptide Biosurfactants 475
Contents xi
21.14 pportunities and Perspectives

O 482
References 483
22 Biosurfactant-Mediated Biocontrol of Pathogenic Microbes of Crop Plants 491

Madhurankhi Goswami and Suresh Deka
22.2 Biosurfactant: Properties and Types 492
22.3 Biosurfactant in Agrochemical Formulations for Sustainable Agriculture 502
22.4 Biosurfactants for a Greener and Safer Environment 503
22.5 Conclusion 503
References 504
Index 510
xii
List of Contributors
Shashi Bhushan Agrawal Kenia Barrantes

Department of Botany Nutrition and Infection Section
Institute of Science Health Research Institute
Banaras Hindu University University of Costa Rica
Varanasi San Jose
Uttar Pradesh Costa Rica
India
Káren Gercyane Oliveira Bezerra
Akil Ahmad Northeastern Network of Biotechnology
School of Industrial Technology Federal Rural University of Pernambuco
Universiti Sains Malaysia Recife
Gelugor Pernambuco
Penang Brazil
Malaysia
Advanced Institute of Technology and
Subramania Angaiah Innovation (IATI)
Electro-Materials Research Lab Recife
Centre for Nanoscience and Technology Pernambuco
Pondicherry University Brazil
Puducherry
India Catholic University of Pernambuco
Recife
Juan José Araya Pernambuco
Escuela de Química Brazil
Centro de Investigaciónen Electroquímica y
Energía Química (CELEQ) Siddhartha Narayan Borah
Universidad de Costa Rica Royal School of Biosciences
San José Royal Global University
Costa Rica Guwahati
Assam, India
Zarith Asyikin Abdul Aziz
School of Chemical and Energy Engineering Luz Chacón
Faculty of Engineering Nutrition and Infection Section
University Teknologi Malaysia Health Research Institute
Johor Bahru University of Costa Rica
Johor Malaysia San Jose, Costa Rica
List of Contributors xiii
Liang Cheng S. J. Geetha

School of Environment and Safety Department of Biology
Engineering College of Science
Jiangsu University Sultan Qaboos University
Zhengjiang Muscat
China Oman
Jéssica Correia
Madhurankhi Goswami
CEB – Centre of Biological Engineering
Environmental Biotechnology Laboratory
University of Minho
Resource Management and
Braga
Environment Section
Portugal
Life Sciences Division
Institute of Advanced Study in Science and
Suresh Deka
Technology (IASST)
Guwahati
Resource Management and Environment
Assam
Section
India
Technology (IASST) Eduardo J. Gudiña
Guwahati CEB – Centre of Biological Engineering
Assam University of Minho
India Braga
Portugal
Santiago de Frutos
Departamento de Química Física Sonam Gupta
Facultad de Ciencias Department of Biotechnology
Universidad de Santiago de Compostela National Institute of Technology, Raipur
Lugo Chhattisgarh
Spain India
Maribel Farfán
Department of Biology Gabriel Ibarra
Healthcare and the Environment Department of Public Health Sciences
Section of Microbiology College of Health Sciences
University of Barcelona University of Texas at El Paso
Barcelona El Paso
Spain TX
USA
Francisco Fraga
Departamento de Física Aplicada Nazim F. Islam
Facultad de Ciencias Department of Botany
Universidad de Santiago de Compostela N N Saikia College
Lugo Assam
Spain India
xiv List of Contributors
Sanket J. Joshi Fernanda Lugo

Oil & Gas Research Center Department of Public Health Sciences
Central Analytical and Applied Research Unit College of Health Sciences
Sultan Qaboos University University of Texas at El Paso
Muscat El Paso
Oman TX, USA
Aida Jover Chandana Malakar

Departamento de Química Física Institute of Advanced Study in Science and
Facultad de Ciencias Technology (IASST)
Universidad de Santiago de Compostela Garchuk
Lugo Assam
Spain India
Arun Karnwal Ana María Marqués

Department of Microbiology Department of Biology
School of Bioengineering and Biosciences Healthcare and the Environment
Lovely Professional University Section of Microbiology
Phagwara University of Barcelona
Punjab Barcelona
India Spain
Asma Khatoon Francisco Meijide

Centre of Lipids Engineering and Applied Departamento de Química Física
Research (CLEAR) Facultad de Ciencias
Universiti Teknologi Malaysia Universidad de Santiago de Compostela
Johor Bahru Lugo
Johor Spain
Malaysia
Monoj Kumar Mondal
Anand K. Kondapi Department of Chemical Engineering and
Laboratory for Molecular Therapeutics Technology
Department of Biotechnology and Indian Institute of Technology
Bioinformatics (Banaras Hindu University)
School of Life Sciences, University of Varanasi
Hyderabad Uttar Pradesh
Hyderabad India
India
Amy R. Nava
Current address: Department of Microbiology Department of Interdisciplinary Health
Immunology and Pathology Sciences
Colorado State University College of Health Sciences
Fort Collins University of Texas
CO El Paso
USA TX, USA
List of Contributors xv
Kannan Pakshirajan Aruliah Rajasekar

Department of Biosciences and Environmental Molecular Microbiology
Bioengineering Research Laboratory
Indian Institute of Technology Guwahati Department of Biotechnology
Guwahati Thiruvalluvar University
Assam Vellore
India Tamilnadu
India
Punniyakotti Parthipan
Electro-Materials Research Lab Ashwini N. Rane
Centre for Nanoscience and Technology Department of Environmental Science
Pondicherry University Savitribai Phule Pune University
Puducherry Pune
India Maharashtra
India
Yogesh Patil
Symbiosis Centre for Research and Selvan Ravindran
Innovation Symbiosis School of Biological Sciences
Symbiosis International University Symbiosis International University
Pune Maharashtra Pune
India Maharashtra
India
Lourdes Pérez
Department of Surfactant and
Ana I. Rodrigues
Nanobiotechnology
IQAC, CSIC
University of Minho
Barcelona
Braga
Spain
Portugal
Aurora Pinazo
Lígia R. Rodrigues
Department of Surfactant and
Nanobiotechnology
University of Minho
IQAC, CSIC
Braga
Barcelona
Portugal
Spain
Rolando Procupez-Schtirbu Rashmi Rekha Saikia

General Chemistry Department of Zoology
Department of Chemistry Jagannath Barooah College
University of Costa Rica Jorhat
San Jose Assam
Costa Rica India
Vikas Pruthi Hemen Sarma

Department of Biotechnology Department of Botany
Indian Institute of Technology Roorkee N N Saikia College
Roorkee Titabar
Uttarakhand Assam
India India
xvi List of Contributors
Leonie Asfora Sarubbo SHE Empire Sdn., Jalan Pulai Ria

Advanced Institute of Technology and Bandar Baru Kangkar Pulai
Innovation (AITI) Skudai
Recife Johor
Pernambuco Malaysia
Brazil
Pooja Singh
Catholic University of Pernambuco Symbiosis School of Biological Sciences
Recife Symbiosis International University
Pernambuco Pune
Brazil Maharashtra
India
Julio A. Seijas Victor H. Soto

Departamento de Química Orgánica School of Chemistry
Facultad de Ciencias Research Center in Electrochemistry and
Universidad de Santiago de Compostela Chemical Energy (CELEQ)
Lugo University of Costa Rica
Spain Costa Rica
Shalini Srivastava
Suparna Sen
Department of Botany
Institute of Science
Resource Management and
Banaras Hindu University
Environment Section
Varanasi
Uttar Pradesh
India
Technology
Guwahati
José A. Teixeira
Assam
India
University of Minho
Braga
Siti Hamidah Mohd Setapar Portugal
School of Chemical and Energy Engineering
Faculty of Engineering, Universiti Teknologi José Vázquez-Tato
Malaysia Departamento de Química Física
Johor Bahru Facultad de Ciencias
Johor Universidad de Santiago de Compostela
Malaysia; Lugo
Spain
Department of Chemical Processes
Malaysia-Japan
M. Pilar Vázquez-Tato
International Institute of Technology
Departamento de Química Orgánica
University Teknologi Malaysia
Facultad de Ciencias
Skudai
Universidad de Santiago de Compostela
Johor
Lugo
Malaysia
Spain
xvii
Preface
This book is useful for the petrochemical industry (enhanced oil recovery from sludge), the
pharmaceutical industry (developed technology for controlling multidrug-resistant pathogens),
and the agro-industry (using byproducts), as well as environmental scientists and engineers
(developing sustainable remediation technologies). As bioremediation is becoming green and a
sustainable approach to environmental pollution control, the articles in this book will be relevant
for future research that could benefit our stakeholders. The chapters in this reference book may be
a unique collection that has been covered by most of the recent studies and provides systematic
material produced by contemporary experts in the field. Focusing on research and development
over the last 10 years, the study highlights relevant developments in the field. We hope that this
book will support researchers by adding a new dimension to environmental studies and the reme-
diation of emerging pollutants. A further benefit would be the understanding of the processes
involved from the production to the sustainable use of biosurfactants in the environment and
biomedicine.
●● This book explains how various methods can be used to recognize and classify microorganism-
producing biosurfactants in the environment. In addition, the various aspects of biosurfactants,
including structural characteristics, developments, production, bioeconomics and their sustain-
able use in the environment, and biomedicine, are addressed. It presents metagenomic strategies
to facilitate the discovery of novel biosurfactants (mechanistic understanding and future pros-
pects) for the sustainable remediation of emerging pollutants.
●● The use of microbes for human well-being is a prospective challenge, as they have developed
novel chemicals and their metabolic pathway could be altered through omics approaches to the
production of high-value chemicals (HVCs), including biosurfactants. These chemicals may be
used in sustainable remediation techniques such as the regulation of the antibiotic resistance
gene (AGR) and microbe-enhanced oil recovery (MEOR). We continue to face new and difficult
challenges in the restoration of the environment, because current methods of remediation require
so many chemicals that have again polluted the environment. There is a need to turn to more
efficient alternative approaches and to find environmentally friendly chemicals for sustainability.
As a result, the microbial world has the option of offering a replacement for green high-value
chemicals to replace certain hazardous compounds already used in environmental reclamation.
This book opens a window on the rapid development of microbiology sciences by explaining
how microbes and their products are used in advanced medical technology and in the sustainable
remediation of emerging environmental contaminants. The authors concentrate on the environ-
ment as well as the biomedical field and highlight the role of microbes in the real world. This book
will be updated to reflect current knowledge, the latest developments in the field of biosurfactants,
xviii Preface
sustainable remediation applications, and applied medical sciences, and the biotechnological strat-
egies being developed to improve production processes. The most important goal of writing this
book will be to communicate current advances and challenges in biosurfactant research. This will
allow the reader to understand the dynamics of applied science that underlie microbially derived
surfactants, called biosurfactants, and their use in sustainable remediation technology. The basic
aim is to include updated content throughout in order to keep pace with this advancing field.
Key features:
●● Addresses the applications of biosurfactants in sustainable remediation technology, for example,

as agents to form emulsions and biofilm formation for desorption of hydrophobic pollutants.
●● Discusses the current state of understanding of the different microbial surfactants, their classifi-
cations, properties, how to achieve higher yields, and new applications.
●● There is a substantial research result on biosurfactants that envisages our capacity to build a
consolidated framework for further development of applications. Biosurfactants for sustainable
remediation technology should fill this need, covering the latest trend on biosurfactant research
and their applications.
The book was contributed by 56 authors from leading surfactants research groups from Brazil,
Costa Rica, China, India, Malaysia, Oman, Portugal, Spain, and the United States, comprising 22
chapters.
1) Introduction to Biosurfactants
2) Metagenomics Approach for Selection of Biosurfactant Producing Bacteria from Oil
Contaminated Soils: An Insight into Its Technology
3) Biosurfactant Production Using Bioreactors from Industrial Byproducts
4) Biosurfactants for Heavy Metal Remediation and Bioeconomics
5) Application of Biosurfactants for Microbial Enhanced Oil Recovery (MEOR)
6) Biosurfactant Enhanced Sustainable Remediation of Petroleum Contaminated Soil
7) Microbial Surfactants Are Next-Generation Biomolecules for Sustainable Remediation of
Polyaromatic Hydrocarbons
8) Biosurfactants for Enhanced Bioavailability of Micronutrients in Soil: A Sustainable Approach
9) Biosurfactants: Production and Role in Synthesis of Nanoparticles for Environmental
Applications
10) Green Surfactants: Production, Properties, and Application in Advanced Medical Technologies
11) Antiviral, Antimicrobial, and Antibiofilm Properties of Biosurfactants: Sustainable Use in
Food and Pharmaceuticals
12) Biosurfactant-Based Antibiofilm Nano Materials
13) Biosurfactants from Bacteria and Fungi: Perspectives on Advanced Biomedical Applications
14) Biosurfactant-Inspired Control of Methicillin-Resistant Staphylococcus aureus (MRSA)
15) Exploiting the Significance of Biosurfactant for the Treatment of Multidrug-Resistant
Pathogenic Infections
16) Biosurfactants Against Drug-Resistant Human and Plant Pathogens: Recent Advances
17) Surfactant- and Biosurfactant-based Therapeutics: Structures, Properties, and Recent
Developments in Drug Delivery and Therapeutic Applications
18) The Potential Use of Biosurfactants in Cosmetics and Dermatological Products: Current
Trends and Future Prospects
Preface xix
19) Cosmeceutical Applications of Biosurfactants: Challenges and Perspectives

20) Biotechnologically Derived Bioactive Molecules for Skin and Hair-Care Application
21) Biosurfactants as Biocontrol Agents Against Mycotoxigenic Fungi
22) Biosurfactant-Mediated Biocontrol of Pathogenic Microbes of Crop Plants
The book explores how these twenty-first century multifunctional biomolecules improve or
replace chemically synthesized surface-active agents with the aid of the industrial application of
biosurfactant production based on renewable resources. This book is also useful for scholars, acad-
emicians in bioengineering and biomedical sciences, undergraduate and graduate students in
microbiology, environmental biotechnology, health, clinical, and pharmaceutical sciences.
1
Introduction to Biosurfactants
José Vázquez Tato1, Julio A. Seijas2, M. Pilar Vázquez-Tato2, Francisco Meijide1,
Santiago de Frutos1, Aida Jover1, Francisco Fraga3, and Victor H. Soto4
1
Departamento de Química Física, Facultad de Ciencias, Universidad de Santiago de Compostela, Avda, Lugo, Spain
2
Departamento de Química Orgánica, Facultad de Ciencias, Universidad de Santiago de Compostela, Avda, Lugo, Spain
3
Departamento de Física Aplicada, Facultad de Ciencias, Universidad de Santiago de Compostela, Avda, Lugo, Spain
4
Escuela de Química, Centro de Investigación en Electroquímica y Energía Química (CELEQ), Universidad de Costa Rica,
San José, Costa Rica
CHAPTER MENU
1.1 Introduction and Historical Perspective, 1
1.2 Micelle Formation, 5
1.3 Average Aggregation Numbers, 14
1.4 Packing Properties of Amphiphiles, 18
1.5 Biosurfactants, 20
1.6 Sophorolipids, 25
1.7 Surfactin, 28
1.8 Final Comments, 31
Acknowledgement, 32
References, 32
1.1 Introduction and Historical Perspective
Surface tension is a property that involves the common frontier (boundary surface) between two
media or phases. Strictly speaking, the surface tension of a liquid should mean the surface tension
of the liquid in contact and equilibrium with its own vapor. However, as the gas phase has nor-
mally a small influence on the surface, the term is generally applied to the liquid–air boundary.
The phases can also be two liquids (interfacial tension) or a liquid and solid. According to IUPAC,
the surface tension is the work required to increase a surface area divided by that area [1]. This is
the reversible work required to carry the molecules or ions from the bulk phase into the surface
implying its enlargement and corresponds to the increase in Gibbs free energy (G) of the system
per unit surface area (A),
G
(1.1)
A T ,P
Biosurfactants for a Sustainable Future: Production and Applications in the Environment and Biomedicine,
First Edition. Edited by Hemen Sarma and Majeti Narasimha Vara Prasad.
© 2021 John Wiley & Sons Ltd. Published 2021 by John Wiley & Sons Ltd.
2 1 Introduction to Biosurfactants
where γ is the interfacial tension. Therefore, the units of γ are J/m2 or N/m, but it is normally
recorded in mN/m (because it coincides with the value in dyn/cm of the cgs system). In 1944,
Taylor and Alexander [2] collected some representative published (1885–1939) values for the sur-
face tension of water at 20 °C. Their own value was 72.70 ± 0.07 mN/m (calculated by extrapola-
tion) in agreement with more recent determinations, the accepted value being 71.99 ± 0.36 mN/m
at 25 °C [3]. This is a rather high value when it is compared with those of other common solvents
as ethanol (22.39 ± 0.06 mN/m), acetic acid (27.59 ± 0.09 mN/m), or acetone (29.26 ± 0.05 mN/m)
(values from [4]) at 20 °C.
The decrease in the surface tension of water has been traditionally achieved by using soaps or
soap-like compounds. According to IUPAC a “soap is a salt of a fatty acid, saturated or unsaturated,
containing at least eight carbon atoms or a mixture of such salts. A neat soap is a lamellar structure
containing much (e.g. 75%) soap and little (e.g. 25%) water. Soaps have the property of reducing the
surface tension of water when they are dissolved in soap-like compounds in water.” This reduction
facilitates personal care, washing of clothes and other fabrics, etc. The early documents with
descriptions of soaps and their uses are typically related with medicinal aspects, and nowadays
there is almost a specific type of soap for each requirement. Levey [5] has reviewed the early his-
tory of “soaps” used in medicine, cleansing, and personal care. For instance, he mentions that “in
a prescription of the seventh century bc, soap made from castor oil (source of ricinoleic [12-hydroxy-
9-cis-octadecenoic] acid) and horned alkali is used. . . as a mouth cleanser, in enemata, and also to
wash the head.” However, Levey concludes that a true soap using caustic alkali was probably not
produced in antiquity but “evidence has been adduced to indicate that salting out was in use in
early Sumerian times.” In his Naturalis Historia, Pliny the Elder [6] refers to soap (sapo) as prodest
et sapo, Galliarum hoc inventum rutilandis capillis. fit ex sebo et cinere, optimus fagino et caprino,
duobus modis, spissus ac liquidus, uterque apud Germanos maiore in usu viris quam feminis, which
may be translated as “There is also soap, an invention of the Gauls for making their hair shiny (or
glossy). It is made from suet and ashes, the best from beechwood ash and goat suet, and exists in
two forms, thick and liquid, both being used among the Germans, more by men than by women.”
Hunt [7] indicates that centers of soap production by the end of the first millennium were in
Marseilles (France) and Savona (Italy), while in Britain some references appear in the literature
around 1000 ad. For instance, in 1192 the monk Richard of Devizes referred to the number of
“soap makers in Bristol and the unpleasant smells which their activities produced.” Hunt also
resumed other aspects as the chemistry of soap, the British alkali industry, the expansion of soap
production, soap manufacturers, and manufacturing methods. As early as 1858, Campbell pre-
sented a USA patent [8] for the production of soaps. He described the process as consisting in “the
use of powdered carbonate of soda for saponifying the fatty acids generally, and more particularly
the red oil or ‘red (oleic) acid oil’ and converting them, by direct combination, into soap in open
pans or kettles, at temperatures between 32 and 500 °F.” Mitchell [9] revised the Jabón de Castilla
or Castile soap (named from the central region of Spain), probably the first white hard soap. It was
an olive oil-based soap and soaps with this name can still be bought today. Traditional recipes and
videos can be easily found on the Internet. In the paper “Literature of Soaps and Synthetic
Detergents”, Schulze [10] recorded the literature (including books, periodicals, abstracts, indexes,
information services, patent publications, association publications, conference proceedings) on
soaps, surfactants, and synthetic detergents up to 1966.
Nowadays descriptions for soap-making from fats and oils are frequent for teaching purposes. For
instance, Phanstiel et al. [11] have described the saponification process (basic hydrolysis of fats). It
involves heating either animal fat or vegetable oil in an alkaline solution. The alkaline solution
hydrolyses the triglyceride into glycerol and salts of the long-chain carboxylic acids (Scheme 1.1).
1.1 Introduction and Historical Perspectiv 3
O O
R1 O HO
R1 O–Na+
R2 NaOH O
HO +
H2O R2 O–Na+
O O
R3 O HO
O R3 O–Na+
Scheme 1.1 Alkaline hydrolysis of a triglyceride to obtain soaps.
To overcome the shortcomings of the carboxylic group of soaps, during the first decades of the
twentieth century, new surface-active agents were obtained in chemistry laboratories. Kastens and
Ayo [12] and Kosswig [13] reviewed the main achievements of these decades. The first result of
this search was Nekal, an alkyl naphthalene sulfonate, although it probably was a mixture of vari-
ous homologs [14]. Other pioneer compounds were Avirol series (sulfuric acid esters of butyl ricin-
oleic acid), Igepon A series (fatty acid esters of hydroxyethanesulfonic acid), Igepon T series
(amide-derivatives of taurine). All these products represented different approaches to the elimina-
tion of the carboxylic group of soaps. IUPAC defines a surfactant as a substance that lowers the
surface tension of the medium in which it is dissolved and/or the interfacial tension with other
phases, and, accordingly, is positively adsorbed at the liquid/vapor and/or at other interfaces. By
detergent, IUPAC refers to a surfactant (or a mixture containing one or more surfactants) having
cleaning properties in dilute solutions. Thus, soaps are surfactants and detergents.
It is not easy to whom the use of the word surfactant should be ascribed for the first time. A
search in SciFinder® suggests that the word was first used by Bellon and LeTellier in a French pat-
ent (1943) [15]. The SciFinder abstract of this patent indicates that “Surfactants such as wetting
agents, detergents, emulsifiers, and stickers are prepared by treating by-product materials contain-
ing starches, cellulose, amino acids, and smaller quantities of inedible fats with NaOH and neu-
tralizing the reaction product.”
Because of their physicochemical properties, surfactants have found applications in almost any
kind of industry. A list of the relevant ISO and DIN regulations for a utility evaluation of sur-
factants has been provided by Kosswig [13]. For instance, in 1950 Lucas and Brown [16] meas-
ured the wetting power of 13 surfactants to find a wetting agent that would enable sulfuric acid to
wet peaches quickly and uniformly so as to permit acid peeling. Anionic, cationic, and neutral
surfactants were tested. In the Application Guide appendix of the book Chemistry and Technology
of Surfactants [17] there is a list that illustrates the variety of surfactants and their versatility in a
wide range of applications. Among others the following are mentioned: Agrochemical formula-
tions, Civil engineering, Cosmetics and toiletries, Detergents, Household products, Miscellaneous
industrial applications, Leather, Metal and engineering, Paints, inks, coatings, and adhesives,
Paper and pulp, Petroleum and oil, Plastics, rubber, and resins, and Textiles and fibers. For
instance, their wetting properties have been early used in food technology. We have already men-
tioned the early connection of soap and medicine and correspondingly the use of surfactants in
pharmacy in the formulation (as emulsifying agents, solubilizers, dispersants, for suspensions)
and as wetting agents, which cannot be a surprise [18]. Nursing care makes a continuous use of
surface-active agents.
The soaps of Scheme 1.1 show the most important structural characteristic of surfactants: the
coexistence of one lyophilic group (alkyl chain) and one lyophobic group (carboxylate ion). In
aqueous solutions, it is more frequent to use the terms hydrophilic and hydrophobic. A graphical
representation head–tail (hydrophobic group–hydrophilic group) is widely used, the alkyl chain
being the tail and the carboxylate group the head (Figure 1.1). This structure gives the amphiphile
character to surfactant compounds.
More generally, the head can be any polar group and the tail any apolar group, leading to a wide
range of structures and types of surfactants. Among anionic heads, typical groups are carboxylate,
sulfate, sulfonate, and phosphate, while the most frequent counterions are monovalent and diva-
lent cations. Polycharged heads are also common, EDTA derivatives being well-known exam-
ples [19]. Cyclopeptides constitute another important group [20]. Among cationic heads, typical
groups are tetralkylammonium, N,N-dialkylimidazolinium and N-alkylpyridinium ions, while
chloride and bromide are the most common counterions. Among neutral heads, polyethylene gly-
col ethers, polyglycol ethers, and carbohydrates can be mentioned. Zwitterionic heads are very
important as phospholipids belong to this group, as well as sulfobetaines and trialkylamine oxides.
Many examples can be found elsewhere [13].
However, the structures of surfactants may be more complex than the head–tail model suggests.
For instance, the number of polar and non-polar groups can be higher than one, the phospholipid
phosphatidylcholine with two alkyl–allyl chains and a zwitterion as the head being an example.
Gemini surfactants are dimeric surfactants [21] carrying two charged groups and two alkyl groups.
The two amphiphilic moieties are connected at the level of the head groups, which are separated by
a spacer group. They are characterized by critical micelle concentrations that are one to two orders
of magnitude lower than those corresponding to conventional (monomeric) surfactants [22].
Bolaamphiphilic molecules contain a hydrophobic skeleton (e.g. one, two, or three alkyl chains,
a steroid, or a porphyrin) and two water-soluble groups on both ends [23]. They can be symmetric
or asymmetric [24, 25]. Recent examples of bolaamphiphilic, Y-shaped and divalent surfactants
have been published by Baccile et al. [26] (Figure 1.1).
Some surfactants, instead of the mentioned head–tail structure, present a bifacial polarity with
the hydrophilic and hydrophobic characteristics at two opposite sides of the molecule. The best-
known examples are bile salts (see Figure 1.2) [27, 28]. Many membrane-active compounds are
facial amphiphiles including cationic peptide antibiotics [29]. The facial amphiphilic conforma-
tion adopted by these peptides is a consequence of their secondary and tertiary structures, allowing
Classic Bolaamphiphile Gemini Phospholipid Divalent Y-shaped Hybrid
Hydrophilic
head
Hydrophobic
tail
Figure 1.1 Schematic representation of the structure of some surfactants.
O Figure 1.2 Bifacial structure of cholic acid.
OH
OH
OH
OH
1.2 Micelle Formatio 5
that one face of the molecule presents cationic groups (protonated amines or guanidines) and the
other face contains hydrophobic groups. An example may be magainin I [30]. Among other sur-
factant structures, diblock copolymers and polymeric surfactants, fluorosurfactants and silicone-
based surfactants can be mentioned [13].
1.2 Micelle Formation
The necessity of a quantitative measurement of the surface tension of soap solutions was soon
evident. By the time that I. Traube published his earliest paper in 1884, significant theories of capil-
larity from La Place, Poisson, or Gauss were known [31]. Early measurements of the surface ten-
sion only imply inorganic salts, acids, and bases. In 1864 Guthrie [32, 33] measured some organic
liquids. At the same time, Musculus [34] studied the capillarity of aqueous solution of alcohol
observing that “the capillarity of the water decreases considerably with the addition of the least
amount of alcohol, in the beginning, much faster than in the presence of more alcohol.” He also
noticed that “all derivatives of ethyl alcohol which are soluble in water (as acetic acid) behave like
this, and probably this is also the case with the other alcohols,” but substances such as “sugars, and
salts if they are not present in a great amount, almost do not influence the capillarity of water.” He
proposed the use of capillarity for measuring the concentration of alcohol and acetic acid in water,
among other reasons, because “it offers the advantage that one needs only very little fluid for
analysis, one drop being enough.” He continued that, as “the animal fluids, such as blood serum,
urine, have a capillarity which is equal to that of water, it is possible to detect and quantify sub-
stances in the urine,” making reference, for instance, to bile.
Traube started the measurement of the influence of many organic substances on the surface ten-
sion of water in the period 1884–1885 [31] and observed that “the surface tension of capillary-
active compounds belonging to one homologous series decreased with each additional CH2 group
in a constant ratio which is approximately 3:1,” leading him to propose Traube’s Rule.
A nice historical paper was published by Traube [31] in 1940, in which he mentioned previous
works related to the investigation of aqueous solutions of inorganic salts, acids, and bases, employ-
ing the method of capillary tubes, and, particularly, the dropping method applied by Quinke.
Traube developed this method and designed a simple instrument, the stalagmometer – together
with the stagonometer – which found general application in science and industry. In the men-
tioned paper, Traube refers mainly to his publications that appeared in the period 1886–1887. By
1906, the measurement of the surface tension by the capillary rise was so important that it was
included in the book Practical Physical Chemistry by A. Findlay. The use of Traube’s stalagmometer
for such a purpose was proposed in the 3rd edition of the book, published in 1915. The experiment
is still proposed in recent textbooks on practical Physical Chemistry [35].
Seventeen of the more important methods of measuring surface tension were described in 1926
by Dorsey [36]. According to his own words, “The list of references does not pretend to be com-
plete but is intended merely to direct the reader to one or more of the sources from which the
required information can be obtained most satisfactorily.” Even so, the number of cited papers was
greater than 110, while the number of citations corresponding to the nineteenth century was 63
(56%). Eminent scientists such as Bohr, Rayleigh, Thomson, Kelvin, Maxwell, Laplace, and Poisson
were among them. Tate [37] published his famous law in 1864 and Wilhelmy in 1863.
Even at low concentrations, surfactants reduce the surface tension of water due to its tendency to
migrate toward the air–water interface, forming a monolayer. This was first suggested in 1907 by
Milner [38] and, previously, Marangoni in 1871 “suggested that this capability [local variation in the
tension of its surface] is due to the presence on the surface of the film of a pellicle, composed of
matter having a smaller capillary tension than that of water.” Milner clearly established that “in
several organic solutions the surface tension is less than that of water, and there is consequently an
excess of solute in the surface.” Later, Langmuir [39] indicated that “the -COOH, -CO, and –OH
groups have more affinity for water than for hydrocarbons. . . [and] when an oil is placed on water,
the –COO– groups combine with the water, while the hydrocarbon chains remain combined with
each other.” In other words, the tail of a surfactant (the hydrocarbon chain) must be located at the
air interface, with the tail upwards oriented and the head (hydrophilic groups) at the water interface.
Rising the surfactant concentration, the surface concentration increases as well until the full
coverage of the interface by the molecules or ions. If the interface is completely covered, further
increment of the surfactant concentration does not (almost) modify the surface tension.
Furthermore, the additional surfactant molecules (or ions) have to remain in the bulk solution,
and following Langmuir “hydrocarbon chains remain combined with each other, thus forming
micelles” (or other aggregates).
The term micelle was commonly used by the first years of the twentieth century [40, 41] in rela-
tion to colloid solutions (frequently inorganic gels). In 1920 McBain and Salmon [42] (see also [43])
described a brief résumé of previous work, citing, for instance, Krafft’s work. From the summary of
this paper we extract the following sentences:
• 3. These colloidal electrolytes are salts in which one of the ions has been replaced by an
ionic micelle.
• 5. This is exemplified by any one of the higher soaps simply on change of concentration.
Thus, in concentrated solution there is little else present than colloid plus cation, whereas
in dilute solution both undissociated and dissociated soap are crystalloids of simple
molecular weight.
• 8. The ionic micelle in the case of soaps exhibits an equivalent conductivity quite equal to
that of potassium ion. Its formula may correspond to Pnn .mH2O but more probably it is
NaP x Pnn .mH2O , where P− is the anion of the fatty acid in question.
Therefore, the essential definition of the present concept of a micelle was established. IUPAC
indicates that “Surfactants in solution are often association colloids, that is, they tend to form
aggregates of colloidal dimensions, which exist in equilibrium with the molecules or ions from
which they are formed. Such aggregates are termed micelles.”
In 1922, McBain and Jenkins [44] studied solutions of sodium oleate and potassium laurate by
ultrafiltration, using this technique for separating the ionic micelle from the neutral colloid. For
both surfactants they showed that the proportion (simple potassium laurate or sodium oleate)/
(ionic micelle) increases fast at low concentrations and reached a plateau at high concentrations
(see graphs of the paper). They also concluded that the diameter of the ionic micelle is only a few
times the length of the molecule and “the particles of sodium oleate are about ten times larger than
those of potassium laurate.”
By the end of the twenties and the beginning of thirties of the twentieth century, the research
activity on micelle-forming substances experienced an extraordinary blooming spring. The paper by
Grindley and Bury [45] is a landmark on the subject, being particularly illustrative for the purposes
of this review. They represented the formation of micelles by butyric acid in solution by the equation
nC3 H 7CO2 H  C3 H 7CO2 H n

(1.2)
where n is “the number of simple molecules in a micelle” or aggregation number (which is a rela-
tively large number) and write the equilibrium constant as
Kn s n /mn (1.3)
where s and m are the concentrations of butyric acid as monomers and as micelles, respectively.
The previous equation can be written as
mn ( s /K ) n (1.4)
from which they deduced that if s/K is appreciably smaller than unity, the concentration of micelles
will be negligible. Only when s approaches the value K does the concentration of micelles become
appreciable, and “will rapidly increase as the total concentration increases.” From this analysis
they conclude that “if any physical property of aqueous butyric acid solutions be plotted against
the concentration, the slope of the curve will change abruptly near this point.” A few months later,
Davies and Bury [46] named that concentration as the critical concentration for micelles.
Previous analysis constitutes the basis of all experimental techniques so far used for determining the
critical concentration for micelles (from here cmc). For instance, the association of monomers in
micelles reduces the number of particles in the solution and, consequently, colligative properties
(freezing point, vapor pressure. . .) also drastically change at this concentration. Other properties such
as solubilization of solutes as dyes or the conductivity of the solution also change significantly. As an
example, we shall mention the paper by Powney and Addison [47] who measured the surface tension
of aqueous solutions of sodium dodecyl, tetradecyl, hexadecyl, and octadecyl sulfates and plotted the
results in the form of vs log (concentration), as we do nowadays. The curves showed breaks at critical
concentrations, which correspond to transitions from single ions to micelles, these single ions consti-
tuting the surface-active species. Figure 1.3 shows a typical plot for an unspecified surfactant. Powney
and Addison noticed that the magnitude of the surface activity and the critical concentration for
micelles were governed by chain length, temperature, and the valency of the added cation.
In 1895, Krafft and Wiglow [48] observed the formation of crystals at 60°, 45°, 31.5°, 11°, 35°, and
0° with hot aqueous solutions (1%) of stearate, palmitate, myristate, laurate, and elaidate sodium
salts, respectively. Each of these temperatures is now known as the Krafft point (Tk). IUPAC defines
it as the temperature (more precisely, narrow temperature range) above which the solubility of a
surfactant rises sharply. At this temperature the solubility of the surfactant becomes equal to the cmc.
In 1955, Hutchinson et al. [49] published the paper “A new interpretation of the properties of
colloidal electrolyte solutions” in which “the formation of micelles was treated as a phase separa-
tion rather than as an association governed by the law of mass action.” Seven years later, Shinoda
and Hutchinson [50] used this model to interpret the Krafft point, associated to the micellization
process. These authors proposed micellization as a “similar phase separation, with the important
distinction that micellization does not lead to an effectively infinite aggregation number, such as
corresponds to true phase separation.” If correct, the model requires that the activity of
Figure 1.3 Typical surface tension vs ln

γ/(mN/m)
(surfactant) plot showing the break point
corresponding to cmc.
cmc In St
micelle-forming compounds should be practically constant above the cmc. Among others, the
authors invoke Nilsson results [51] with radiotracers as evidence for their proposition. In a fre-
quently reproduced graph, Shinoda and Hutchinson [50] plotted the concentration vs temperature
for sodium decyl sulfonate near the Krafft point. The plot resembles the phase diagram of water
near its triple point. If micelles are considered as a phase, by the phase rule, the system should
become invariant at constant temperature and pressure [49, 50]. In other words, “the equilibrium
hydrated solid monomers micelles is univariant, so that at a given pressure the point is fixed.” As
temperature increases, the solubility also increases until Tk where the cmc is reached. Above this
temperature, the surfactant is dissolved in the form of micelles.
Let us go back to 1915. In this year, Allen [52] published a paper in which he showed the use of
the surface tension measurement for the determination of bile salts in urine. In the introduction of
his paper, Allen refers to Hay’s method of testing the presence of bile salts in the urine. That
method consists in “shaking flowers of sulphur upon the surface of the urine. . . When the surface
tension of the urine is lowered [by bile salts] the powdered sulphur sinks to the bottom, and the
lower the surface tension the more rapidly this takes place. [But] The method is very unsatisfactory. . .
and if possible, a quantitative method, would be very desirable.” Thus Allen proposed a very accu-
rate measurement of the surface tension of a solution by the stalagmometric method “to determine
the feasibility of estimating the amount of bile salts present in pathological urines from measure-
ments of the surface tension taken with a portable Traube stalagmometer.” He computed the sur-
face tension value of a solution in per cent of that of distilled water according to the formula
Number of drops of distillided water
specific gravity of the solution (1.5)
Number of drops of solution
The method relies on the fact that bile salts possess the property of lowering the surface tension
of a solution very markedly, even when present in small concentrations. Table I of his paper shows
some results for sodium glycocholate (NaGC) in distilled water. In a reanalysis of these data by
plotting the surface tension vs ln(concentration), it is possible to determine a value of 0.0117 M for
the cmc of NaGC, a value in perfect agreement with recent measurements. From the Reis et al. [53]
compilation, an average value of (1.04 ± 0.29) × 10−2 M may be estimated for the cmc of this bile salt.
The “excess of solute [surfactant] in the surface” indicated by Milner has traditionally been ana-
lyzed through the Gibbs equation [54]
1
(1.6)
nRT ln c
where Γ is the surface excess, ( γ/ ln c) is the slope of the dependence of γ with the logarithm of the
concentration of the surfactant (frequently being linear), R is the ideal gas constant, T the tempera-
ture, and n a factor that depends on the nature of the surfactant. The equation allows the determina-
tion of the area occupied per molecule at the interface, which is the inverse of the surface excess, i.e.
1
a (1.7)
NA
where NA is Avogadro’s number.
Recently Menger et al. [55] have questioned the validity of the Gibbs equation on the basis that
in the region of concentration where the equation is applied the adsorption at the interface does
not generally reach saturation. This criticism has been supported by measurements from a radioac-
tive surfactant [56], results that suggest that the γ-ln c linearity is not indicative of surface satura-
tion, a hypothesis required for the deduction of the Gibbs equation. Neutron reflection
measurements also support the fact that there are serious limitations in applying the Gibbs equa-
tion accurately to surface tension data [57, 58].
In the late nineteen thirties, other important papers were published. Wright and co-workers [59–61]
measured the conductivity, density, viscosity, and solubility of several sodium alkyl (decyl, dodecyl,
and hexadecyl) sulfonates at several temperatures. In all cases breaks at the curves or linear depend-
ences of the property with the sulfonate concentration were observed. They also reported that the
addition of sodium chloride to solutions of sodium dodecyl sulfonate lowered the cmc and that the
lowering becomes less marked with a rise in temperature. Hartley [62] demonstrated that paraffin
chain salts behave as strong electrolytes at low concentrations. For cetane sulfonic acid, a value of
about 0.008 N in water at 60 °C was given for cmc and that it increased by about 2% per degree. This
is an important question since the formation (or not) of premicellar aggregates is still under debate.
By the end of this decade Hartley [63] reviewed (36 references) the subject, the title of the paper
being Ion aggregation in solutions of salts with long paraffin chains. In the abstracts we can read about
the structure of micelles which are “aggregates of paraffin-chain ions with some adsorbed opposite
ions,” micelles are spherical with a radius equal to the length of a completely stretched paraffin-chain
and have a liquid interior and the strong dependence of cmc with the length of the hydrocarbon
chain and nature of the ionized terminal groups and opposite ions (counterions), and with tempera-
ture (in less extension). He also affirmed that “the spherical micelle is more stable than ion pairs.”
Thus, by this time, the essential parameters that define a micelle were introduced or established:
change of properties at the cmc, variables that influence the cmc, shape and size, internal and
peripheral structures, and the essential thermodynamics (mass action law).
In the period 1946–1947, immediately after the Second World War, the activity on surfactant
research experiences an important enhancement.
Corrin and Harkins [64] proposed the equation log(cmc) = − A × log(counterion±) − B to relate the
dependence of the cmc with the concentration of added salts (the sign at the superscript of the coun-
terion is opposite to that of the surfactant ion). Table 1.1 resumes the values for the constants A and
B for several surfactants. They also noticed that urea has a negligible effect in lowering the cmc.
Three years later, Lange [65] applied the mass action law to ionic micelles and wrote the equilib-
rium of formation of the micelle as
pK qA  K p Aq q p (1.8)
where K is the counterion, A the surfactant ion, and p and q the stoichiometric coefficients.
Although Lange considered the activity coefficients of the different species, for simplicity we will
ignore them and write the equilibrium constant as
p q
K A K eq K p Aq (1.9)
Table 1.1 Parameters A and B of the Corrin–Harkins equation.

The number of figures on the values of A and B has been reduced.
Surfactant A B
Potassium laurate 0.570 2.62

Sodium dodecyl sulfate 0.458 3.25
Dodecyl ammonium chloride 0.562 2.86
Decyltrimethylammonium bromide 0.343 1.58
Source: Corrin and Harkins [64], p. 683.

Writing [A] = ck and [K] = ck + N, where N is the equivalent concentration of added salt, it is finally
found that
p 1
log ck log ck N log L (1.10)
q q
where L = [KpAq]. Thus with logck as ordinate and log(ck + N) as abscissa, this is the equation of a
straight line with the slope –p/q, which corresponds to the empirical one found by Corrin and
Harkins. This point has been discussed in detail by Hall [66] in his theory for dilute solutions of
polyelectrolytes and of ionic surfactants.
The effects of solvents (alkyl alcohols CnH2n+1OH, n = 1–4; HOCH2CH2OH, glycerol, 1,4-diox-
ane, and heptanol) on the critical concentration for micelle formation of cationic soaps was stud-
ied by Corrin and Harkins [67], Herzfeld et al. [68], and Reichenberg [69]. Klevens [70] found that
increasing the temperature causes an apparent decrease in the cmc, as determined by spectral
changes in various dyes. However, this same author found the opposite effect when the micelles
formation was determined by refraction [71].
Simultaneously, other experimental techniques, mainly spectroscopic ones, were introduced for
the determination of the cmc. After a paper published by Sheppard and Geddes [72], in which the
authors reported that by the addition of cetyl pyridinium chloride, the absorption spectrum of aque-
ous pinacyanol chloride was shifted from that exhibited in aqueous solutions to that in non-polar
solvents, Corrin et al. [73] used this property to determine the cmc of laurate and myristate potas-
sium salts, giving values of 6 × 10−3 M and 0.023–0.024 M, respectively. The concentration of soap at
which this spectral change occurs was taken as the cmc, proposing that the dye is solubilized in a
non-polar environment within the micelle. Klevens [74] performed a similar work by studying the
changes in the spectrum of pinacyanol chloride in solutions of myristate, laurate, caprate and
caprylate potassium salts, and sodium lauryl sulfate. These studies were extended to other sur-
factants [75] and other dyes as p-dimethylaminoazobenzene [76]. By using suitable dyes (Rhodamine
6G, Fluorescein, Acridine Orange, Acridine Yellow, Acriflavine, and Dichlorofluorescein) fluores-
cence spectroscopy was soon adopted [77, 78].
In 1950, Klevens [79] studied the solubility of some polycyclic hydrocarbons in water and in
solutions of potassium laurate (at 25 °C). For all the polycyclic hydrocarbons, he showed that by
increasing the concentration of the surfactant, their solubility also increased. Particularly, for pyr-
ene he measured solubilities of 0.77 × 10−6 and 2.24 × 10−3 M in water and potassium laurate
(0.50 M), respectively.
One year later, Ekwall [80] studied the sodium cholate association by measuring the fluorescence
intensity, and determined that the lowest concentration at which polycyclic hydrocarbons
(3,4-benzopyrene included) are solubilized is 0.018 M. This corresponds to the beginning of the
micelle formation, although “at first relatively small amounts of cholate ion aggregates and the actual
micelle formation occurs at about 0.040 to 0.044 M.” Foerster and Selinger [81] observed that in
micelles of cetyldimethylbenzylammonium chloride, pyrene forms dimers in excited states (excimers).
In the period 1971–1980, the number of papers on solubilized pyrene in micelle solutions
increased very quickly. The fluorescence decay of the excited state of pyrene received an important
attention. The aggregation number and microviscosities of the micellar interior [82], the permea-
bility of these micelles with respect to nonionic and ionic quenchers [83], oxygen penetration of
micelles [84], or the environmental effects on the vibronic band intensities in pyrene monomer
fluorescence in micellar systems [85, 86] were published. Kalyanasundaram and Thomas carefully
analyzed the lifetime of the monomer fluorescence and the ratio I3/I1 of the third and first vibronic
band intensities of pyrene in sodium lauryl sulfate as a function of its concentration. Both curves
have a sigmoidal shape (see Figure 1.3 of the paper). A value of 8 × 10−3 M for the cmc of the sur-
factant was given.
However, Nakajima [86] plotted the ratio I1/I3 and accepted the cmc as the concentration at which
the first break is observed (point A in Figure 1.4). At low concentrations of the surfactants the values
of the I1/I3 ratio are high, typical of a hydrophilic environment for pyrene, the value in water being
1.96 [87] while at high surfactant concentrations the I1/I3 ratio tend to typical values of non-polar
solvents. For instance, at high surfactant concentrations of sodium cholate and sodium deoxycho-
late, the I1/I3 ratio is around 0.7 [88] while the value in cyclohexane is 0.61 [89]. This suggests that the
polarity of the microenvironment of pyrene is a lipophilic one. Andersson and Olofsson [90], when
performing a calorimetric study of nonionic surfactants, also made use of Nakajima’s approach.
Other authors have proposed the inflection point of the curve (point B) as cmc [91]. As such it
fulfills the condition
d
0 (1.11)
dSt St cmc
where φ would be the I1/I3 ratio. The expression is also valid for any other property that exhibits a
sigmoidal behavior as the obtained enthalpograms from isothermal titration calorimetry (ITC) [92].
The plot of (dφ/dSt) vs St is shown in Figure 1.4 (right) and the cmc is easily obtained from the peak.
Aguiar et al. [93] have analyzed both points (A and B) for several surfactants and proposed an
approach for choosing between one or the other point. Occasionally, both A and C points have been
accepted as an indication that the system has two cmc values. We consider that this is not correct.
These different points of view introduce an important question related to the determination of the
cmc from sigmoidal curves, which are frequently found when using some experimental techniques.
By now, some different approaches to determine the cmc have already been introduced.
Rusanov [94] has reviewed the definitions of cmc based on the application of the mass action law
to the aggregation process in surfactant solutions. Among them, we must mention the definition
given by the equation
d3
0 (1.12)
dSt3 St cmc
φA A
φ dφ/dSt
φB
B
φC
C
SA SB SC St cmc St
Figure 1.4 Typical plot of a sigmoidal curve. Example φ = I1/I3 (ratio of the intensities of the first and third
vibronic peaks of pyrene) vs increasing concentration of a surfactant (left) and its first derivative (right). The
shape of curves from isothermal titration calorimetry are similar in shape (see below for a description).
which was proposed by Phillips [95] in 1955 for determining the cmc for an ideal measured prop-
erty (φ)-concentration (St) relationship. Phillips pretended that Eq. (1.12) corresponds to the point
of maximum curvature, but this is not the case. Nakajima’s approach fulfills this condition as well
as the methodology proposed by Olesen et al. [96] for determining the aggregation number of
aggregates from ITC curves. However, the definition of cmc as corresponding to the inflection
point in the φ vs St curve has been recommended [97] for the determination of the cmc from ITC
curves. For large absolute values of the slope at the inflection point (= (φC − φA)/(SC − SA)) in previ-
ous sigmoidal curves, the difference in the values obtained from any of the two previous equations
may be considered negligible for practical purposes. If the cmc is fairly sharp, Hall [98] has pro-
posed that it can be regarded approximately as a second order phase transition.
Among the other definitions for cmc analyzed by Rusanov we would like to remark on the fol-
lowing one. The focus is on a system in which micelles are composed of a single sort of particle.
For further details and the analysis of more complex systems, the two papers by Rusanov [94] are
recommended.
Let us redefine the Grindley and Bury [45] equilibrium constant as
K K on 1
mn /s n (1.13)
The equilibrium constant Ko would correspond to a hypothetical single step in which a virtual
aggregate mj is formed by the binding of an additional monomer to a virtual aggregate of size mj-1
according to
s mj 1  mj (1.14)
its equilibrium constant being
Kj m j / m j 1s (1.15)
The isodesmic model accepts that all Kj constants are equal to Ko. The difference with the
Grindley and Bury equilibrium constant comes from the fact that only (n – 1) steps are required to
form a micelle with n monomers. Interestingly, in 1935 Goodeve [99] have pointed out that form-
ing micelles of, say, about 20 molecules must pass through all the intermediate stages of associa-
tion. The formation of the micelles from the monomer in one stage is, of course, highly improbable
as it requires “a collision of 20 molecules at one time.” Goodeve presented Eq. (1.14) as represent-
ing the equilibrium according to this point of view.
Equation (1.13) is better understood in the form
mn n 1
Ko s (1.16)
s
where Ko and s are both positive, n is usually large, and, independently of the value of n, for
Ko × s = 1, the concentration of micelles and monomers are the same. Deviations of the product
Ko × s from that value lead to either mn < s or mn > s. For instance, for n = 50, the ratio mn/s changes
by a factor 1.86 × 104 when Ko × s varies from 0.9 to 1.1. This is in fact the analysis by Grindley and
Bury [45].
This suggests a definition of cmc by the condition
scmc Ko 1 (1.17)
and from the conservation of material (St = s + n × mn) it follows that at cmc St,cmc = (n + 1)scmc =
(n + 1)mn.
From Eq. (1.16) it also follows that
mn 1 mn
(1.18)
s n s
or
ln mn
n (1.19)
ln s
In a monodisperse system, this equation may be simplified to
mn
n (1.20)
s cmc
since at cmc, mn = s and n is constant for the whole range of surfactant concentrations. Thus, the
larger the rate of change of the micelle concentration is with respect to the change in the monomer
concentration, the higher the aggregation number will be.
Once the equilibrium constant and the aggregation number are known, all the thermodynamic
functions may be obtained. These thermodynamic quantities have traditionally been determined
from the measurement of cmc at different temperatures, the range of temperatures being around
40 °C (or less). The problem is that the dependence of the cmc with temperature is usually low for
most of the surfactants and, as the dependence of ΔGo with the concentration is logarithmic, the
range of experimental values is even shorter. This introduces an error in the determination of the
thermodynamic amounts, which is necessarily rather high.
The commercial introduction of high quality isothermal titration calorimeters has provided a
routine way for the determination of previous amounts, which have a much higher precision. In a
typical measurement a sample cell is filled with water (or any other appropriate solvent). A sur-
factant solution is placed in a syringe, which allows the injection of small aliquots into the sample
cell at different intervals of time. The solvent of this solution and the one filling the sample cell
must be identical to prevent some effects as the dilution heat of inert salts or buffers. The concen-
tration of the surfactant ranges from 10 to 30 times the cmc value. Each injection increases the
surfactant concentration in the sample cell from zero to a concentration clearly above the cmc. The
heat involved in the process (the concentration in the syringe is always higher than in the sample
cell) after each injection is measured and plotted vs the increasing concentration in the sample cell.
Figure 1.4 imitates a typical enthalpogram and its derivative. In this case, φ = ΔH (in kJ/mol of
injectant) is the involved heat after each injection.
An ideal enthalpogram can be subdivided into three concentration ranges. In region I (first injections,
till point A in Figure 1.4) the increasing concentration in the cell is still below the cmc. Here, the large
enthalpic effects observed are mainly due to breaking micelles into monomers (demicellization pro-
cess) and dilution of monomers [92]. In region III (final injections, after point C in Figure 1.4), the
increasing concentration in the cell is above the cmc. Here, the low enthalpic effects observed are
mainly due to dilution of micelles. In the central region (between A and C in Figure 1.4), a sharp
decrease is observed and corresponds to a transition from reaching the cmc and exceeding it.
Therefore, the cmc corresponds to the inflection point of the curve, which can easily be determined
as the first derivative of the curve (Eq. (1.11), Figure 1.4, right). The heat of demicellization ΔHdemic
is equal to the enthalpy difference between the two extrapolated lines in Figure 1.4. Thus, the cmc
and the enthalpy of micellization are simultaneously determined, but independently to each other.
Repetition of the ITC experiment at other temperatures allows the determination of the change
in the heat capacity of the demicellization process, ∆CPo . The interval of temperatures used in these
studies is rarely larger than 30–40 °C. Within this interval, the dependence of ΔHdemic for most of
the surfactants is linear with T, meaning that ∆CPo is constant ([100] and references therein). While
ΔHdemic may be either positive (endothermic) or negative (exothermic), ∆CPo for the demicelliza-
tion process is always positive. This means that the hydrophobic surface of monomers, being
exposed to water, increases upon demicellization. For this reason, it is frequently observed that
ΔHdemic is negative at low temperatures and positive at high ones. The formation of a micelle
requires that some water molecules surrounding each monomer must be lost in the aggregation
process to form the final aggregate. The process also contributes to a favorable entropy term for
micellization. Thus, the transfer of surfactant monomers from an aggregate to the bulk water has
many facts in common with the dissolution of liquid alkanes into water [101]. Gill et al. [102] have
noticed that the experimental heat capacity difference between gaseous and dissolved non-polar
molecules in water is correlated with the number of water molecules in the first solvation shell.
They concluded that a two-state model, in which each water molecule in the solvation shell
behaves independently, provides a satisfactory basis to quantitatively describe the heat capacity
properties of the solvation shell. For a series of solutes (most of them being hydrocarbon com-
pounds), an average value of ~13.3 J/mol K (see the theoretical line shown in Figure 1.1 at 25 °C of
that paper) was estimated for the contribution of each water molecule to ∆CPo .
Calorimetric measurements of vapor equilibrium of the system cyclohexane-heptane were per-
formed almost 70 years ago by Crutzen et al. [103]. These authors observed that between 40 and
60 °C, the increase in the molar free Gibbs energy becomes small because of the partial compensa-
tion of the heat of mixing and the entropy of mixing. Since then, many papers have been published
in which the concept enthalpy–entropy compensation (EEC) has been taken into consideration.
Arguments for or against EEC have been published and, for surfactant systems, EEC has been
reviewed several times [100, 104–106]. For demicellization (or equivalently micellization), the
relationship is written linearly as
H Hc Tc S (1.21)
where Tc = ( ∆H/ ∆S)P is known as the compensation temperature.

Recently, Vázquez-Tato et al. [100] have shown that “it is possible to obtain as many compensa-
tion temperature values as the number of temperature intervals in which the dependencies of
enthalpy and entropy changes with temperature are analyzed.” Furthermore, “the value of each Tc
will agree with the central value To of each temperature interval.” These authors concluded that
“Tc is simply such experimental To” without any physical meaning and concluded that it “does not
provide any additional information about the systems.” In other words, any physical interpretation
derived from Tc (and by extension from ΔHc) is meaningless.
1.3 Average Aggregation Numbers

Recently Olesen et al. [96] have published a method for analyzing the ITC curves that allows the
determination of the aggregation number. As in previous cases, we will limit the presentation to
neutral surfactants. The mass-action model and monodispersity of micelles are assumed.
Other significant papers for determining the aggregation number of micelles are those by Debye
and by Turro and Yekta. After his landmark paper published in 1947 for the molecular weight
determination by light scattering, Debye [107] immediately published the first determinations of
1.3 Average Aggregation Number 15
micellar molecular weights by this technique in 1949 [108], the surfactants being alkyl quaternary
ammonium salts and amine hydrochlorides. A few years later, Tartar and Lelong [109] determined
the micellar molecular weights of some paraffin chain salts by the same technique. Nowadays, the
technique is almost routinely applied in laboratories for determination of molecular weights of
polymers, micelles, and so on.
In 1978, Turro and Yekta [110] presented a simple procedure for determination of the mean
aggregation number of micelles by measuring the steady-state fluorescence quenching of a lumi-
nescent probe by a hydrophobic quencher. The Poisson statistics to describe the distribution of the
luminescent molecule D and the quencher Q in a solution that contains a well-defined but
unknown micelle concentration [M] was accepted. Both D and Q are selected in such a way that
they reside exclusively in the micellar phase. D will partition itself both among micelles containing
Q and among “empty” micelles. They also assumed that only excited micelles of D, D*, emit in the
micelles containing no Q, i.e. D* is completely quenched when it occupies a micelle containing at
least one Q. Under these conditions a “very simple expression” for obtaining the aggregation num-
ber is deduced.
In 1899 Biltz [111] published the book Practical Methods for Determining Molecular Weights. The
book is a summation of the practical methods for determining molecular weights by vapordensity
and other methods based on colligative properties, mainly from the measurement of the increase
of the boiling point of a solution with respect to a pure solvent, and the freezing-point method.
Both methods, together with the Nernst method (based upon the principle of lowering of solubil-
ity) are the only ones that “find practical application in the laboratory.” When the freezing-point
method is applied to some salts, Biltz accepts the Arrhenius dissociation theory of electrolyte solu-
tions. Otherwise “the electrolytic dissociation in aqueous solutions can lead to smaller molecular
weights than would be expected from the formula of the substance.” Earlier in 1896, Krafft [112]
noticed that the sodium salts of the shorter fatty acids exist in the “molecular state” (meaning that
they are in the state of single molecules, and not in that of molecular aggregates) in aqueous solu-
tion and that they give twice the normal rise of boiling point which in fact it would correspond
with a hydrolytic decomposition into sodium (hydroxide, in the original) and the fatty acid.
Following Biltz, the reverse can take place if “. . . by condensation several simple molecules form a
more complex molecule. . .. The term association has recently been proposed for this kind of con-
densation.” Similarly, Kahlenberg and Schreiner [113] observed that the reduction of the freezing
point of solutions of sodium oleate resulted in a molecular weight, which is nearly twice as large,
like the theoretical formula. Botazzi and d’Errico [114] investigated glycogen of different concen-
trations by viscosity, freezing point, and electrical conductivity and observed that when the con-
centration of glycogen solutions reaches a certain maximum it appears that the colloidal particles
combine with one another to form micelles. McBain et al. [115] measured the freezing point and
the conductivity of sodium and potassium salts of saturated fatty acids that remain liquid at 0°.
From this paper we must remark the comment that “free ions of charge equal and opposite to that
of the charged colloid are present in the sol or gel.” In 1935 McBain and Betz [116] measured the
freezing point of undecyl and lauryl sulfonic acids, expressing the results in terms of the osmotic
coefficient. They concluded that in dilute solutions they behave as simple moderately weak elec-
trolytes but with increased concentration molecules and ions associate into neutral and ionic
micelles. They also considered that micelles “owing to the wide spacing of their charges, have the
ionic strength similar to uni-univalent electrolytes.” McBain and Betz [116] and Johnston and
McBain [117] carried out careful freezing point measurements on colloidal electrolytes (potassium
and sodium oleate, sodium decyl and dodecyl sulphate, sodium decyl sulfonate and sodium deoxy-
cholate) and, in 1947, Gonick and McBain [118] showed a “cryoscopic” evidence of a micellar
association in aqueous solution of nonionic detergents. Following their own words, “since the
depression of the freezing point is determined primarily by the number of solute particles per unit
weight of solvent, the osmotic coefficient expresses directly, to a first approximation, the ratio of
the true number of solute particles to that obtaining at complete dispersion of the solute and thus
gives a measure of the average degree of association into micelles or other aggregates.” The critical
concentration of micelles formation was evident from an abrupt drop in the coefficient.
Furthermore, they noticed that the addition of potassium chloride to a solution of nonaethylene
glycol (mono) laurate caused no significant change in the degree of association of the detergent.
Herrington and Sahi [119] studied the nonionic surfactants sucrose monolaurate and sucrose
monooleate in aqueous solution by the freezing point and vapor pressure methods. For the analysis
of the results they accepted that micelles are monodisperse and observed that the aggregation
numbers do not increase significantly with temperature.
Starting from a model published by Burchfield and Woolley [120], Coello et al. [121, 122] com-
bined freezing point and sodium ion activities measurements to obtain both the aggregation num-
ber and the fraction of bound counterions of aqueous bile salt solutions. The mass action model for
an ionic micelle and the fraction of bound counterions to the micelles were considered. The theory
of the freezing point depression is very well known [54] and does not need further attention.
While in a monodisperse system (as we have analyzed above) the aggregation number is a single-
value variable [123], in a polydisperse self-aggregation system the aggregation number can assume
all possible values from 2 to . For these systems only the average aggregation number of the
aggregates is obtained. However, different experimental techniques lead to aggregation numbers
that are not the same. For instance, the average aggregation number measured from colligative
properties is the number average aggregation number, n, given by
iXi
n (1.22)
Xi
where Xi is the concentration of the ith species. Similarly, the average aggregation numbers
obtained from static light scattering and viscosity techniques are the weight average and the
z-average aggregation numbers, nw and nz , respectively, given by
i 2 Xi
nw (1.23)
iXi
i 3 Xi
nz (1.24)
i 2 Xi
For analyzing the experimental results from the experimental techniques, the hypothesis of a
constant average aggregation number with concentration is frequently used. Let us examine this
hypothesis by following the landmark paper by Israelachvili et al. [124]. In this presentation we are
following the notation of this original paper.
Each aggregate in the solution is characterized by its own free energy. They also accepted that
the dilute solution theory holds. Equilibrium thermodynamics requires that in a system of mole-
cules that form aggregated structures in solution, the chemical potential of all identical molecules
in different aggregates is the same [125]. For an aggregate containing i monomer molecules, this
statement is expressed as
o o kT Xi
1 kT ln X1 i ln (1.25)
i i
1.3 Average Aggregation Number 17
where µio is the standard part of the chemical potential (the mean interaction free energy per mol-
ecule) in aggregates of aggregation number i and Xi is the concentration of the ith aggregate. The
subscript “1” corresponds to monomers in solution. That is to say, the left-side term corresponds to
the chemical potential of the free monomer and the right-side term is the chemical potential per
monomer incorporated in a micelle.
From the previous equation and the material balance in a polydispersed micellar solution, it fol-
lows that the number average aggregation number is [124]
ln St X1
n (1.26)
ln X1
which relates the rate of change of micelle concentration with monomer concentration to the
mean micelle aggregation number. It should be noticed that this equation reduces to Eq. (1.19) for
a monodisperse system. In such a case, the various average aggregation numbers are identical to
n [123].
The standard deviation is given by
2 ln n
ln St X1 (1.27)
which relates it to the rate of change of the mean aggregation number with respect to micelle con-
centration and means that if the standard deviation is close to zero, the aggregation number must
almost be independent of the total surfactant concentration.
Well above cmc, Eq. (1.27) may be approximated by [125]
2 ln n
(1.28)
ln(St )
which shows that if the system is highly polydisperse the average aggregation number will be very
sensitive to the total surfactant concentration. Therefore, a rapid change of concentration is evi-
dence of a large distribution of polydispersity in micelle size. Nagarajan [123] remarks that “this is
a general thermodynamic result applicable to any self-assembling system” and that “interpreting
any experimental data, one must ensure that this equation is not violated.” Similar comments have
been provided by Rusanov [126].
A well-known approach to study a fast process is to rapidly perturb a system in equilibrium
and measure the relaxation time required by the system to adjust to the new equilibrium. Typical
techniques are temperature jump, pressure jump, and ultrasonics [127]. The time scale depends
on the relaxation technique [128]. The mathematical analysis to study simple A↔B and A+B↔C
systems by temperature jump has been didactically published by Finholt [129]. Kresheck
et al. [130] applied the temperature-jump (jump 5.2 °C) technique to study the dissociation of
the dodecylpyridinium iodide micelle. Pressure-jump studies have also been carried out by
several authors [131–133].
In the ultrasonic methods (Ultrasonic Absorption Spectrometry) the perturbation of equilibrium
is due to the periodic variation of pressure and temperature due to the passage of the sound wave
through the system. Relaxation of the equilibrium gives rise to changes in the quantity α/f2 (where
α is the sound absorption coefficient at frequency f ) and from its dependence with frequency the
relaxation time is obtained [128].
Platz [134] has presented a simplified analysis of the Aniansson and Wall [135] isodesmic model
for analyzing the kinetics of micelle association and dissociation in surfactant solutions. The the-
ory is nowadays commonly known as the Teubner–Kahlweit–Aniansson–Wall theory [136, 137],
the key equation being Eq. (1.14), characterized by forward (kif ) and back (kib ) kinetic constants,
which correspond to the exchange of monomers between micelles. The ratio between the kinetic
constants is directly the equilibrium constant. Further developments of the theory are due to Lang
et al. [133] and Telgmann and Kaatze [138, 139]. In relaxation experiments two characteristic
times are observed. The so-called “fast process” corresponds to the kinetic analysis of Aniansson
and Wall and the “slow process” is assumed to be a change of the total number of micelles.
After several assumptions (micellar distribution is Gaussian-like, reactions between aggregates
do not have to be considered, and the change in the number of the micelles is much slower than
the interchange of the monomer) Eq. (1.29) is deduced [140]:
1 1 1 St s
kb 2 (1.29)
1 n s
where τ1 is the relaxation time (associated to kb), σ2 is the variance of the micellar distribution,
kb knb is the backward rate constant at micelle sizes around the mean micelle size n, and s is the
average monomer concentration. In addition, s is identified with the concentration at cmc, scmc, at
concentrations above the cmc. When s , kb, the aggregation number and the standard deviation (see
above) do not appreciably change with the concentration, and the previous equation suggests a
linear relationship between 1 1 and the concentration. This fact has been verified, for instance, for
alkyl sulfates [128].
The backward kinetic constant depends on the length of the alkyl chain of the surfactant. For
instance, values in the interval 10–0.8 × 109/s have been measured by Kaatze [136] for ammonium
chloride surfactants CH3Cx−1H2(x−1)NH3+Cl− (x = 5–8).
1.4 Packing Properties of Amphiphiles

In 1964 Reiss-Husson and Luzzati [141] used small-angle X-ray scattering methods to study micel-
lar solutions of several amphiphiles in water, without added electrolytes. The amphiphiles were
sodium salts of lauryl sulfate (SLS), laurate (NaC12), myristate, palmitate, stearate and oleate, and
cetyltrimethylammonium chloride (CTACl) and bromide. The studies were performed at different
concentrations and temperatures. SLS (67, 25 °C), NaC12 (25, 70 °C), and CTACl (84, 27 °C) form
spherical micelles with the aggregation numbers given in parenthesis. However, at high surfactant
concentrations the spheres become rods and the concentration at which the sphere–rod transition
takes place was also determined.
The sphere–rod transition was observed by Hayashi and Ikeda [142] for SLS when the concentra-
tion of NaCl is increased. The transition is accompanied by a large increment of the aggregation
number. For instance, at NaCl 0.01 M the apparent aggregation number is 70 while at NaCl 0.80 M
the value is 1630 and the length of the rod 597 Å. A simple version of this experiment was provided
by Coello et al. [143].
The monomer packing of an alkyl surfactant in a given geometrical shape, for instance, a sphere,
will depend on parameters such as the micellar radius, R, chain monomer volume, v, maximum
1.4 Packing Properties of Amphiphile 19
length that the chain can assume (critical chain length, lc), and the interfacial area per monomer,
ao. For spherical micelles, simple geometry gives [125],
4 R3
n (1.30)
3v
(from the volume) and
4 R2
n (1.31)
ao
(from the surface). It follows that
3v
R (1.32)
ao
According to Tanford [144], for an alkyl chain with nc carbon atoms the maximum chain length
is given by
lmax 1.5 1.265nc A (1.33)
and the volume of the alkyl chain as
v 27.4 26.9nc A (1.34)
Only with the condition (lc critical length)
v 1
(1.35)
aolc 3
will the amphiphiles be able to fill the whole volume of the spherical micelle without leaving an
empty space in the micelle core. For two surfactants having identical ao values, previous equations
predict that both the radius (R2 = (v2/v1)2R1) and the aggregation number (n2 = (v2/v1)2n1) will
increase with the alkyl chain length. Figure 1.5 shows the plot of ln n vs ln v for sodium alkyl sul-
phates at 25 °C [145] and sodium alkyl sulfonates at 23–60 °C [76], the values of the slopes being
2.06 ± 0.05 and 2.20 ± 0.11, respectively, in good agreement with the theoretical expected one equal
to 2. For the original measurements of Debye the value of the slope is 3.53. The value of ao depends
not only on the structure of the hydrophilic head group but also on the environmental properties
(presence of inert salts, pH, additives, temperature).
The ratio (v/aolc) of Eq. (1.35) is named as packing parameter P.
Surfactants can self-aggregate in other well-defined structures, different from spherical, ellipsoi-
dal or rod-like micelles. Flat lamellar or disk-like structures are also common. Jung et al. [146]
have studied the origins of stability of spontaneous vesicles. The cryo-TEM images provided by
these authors, unequivocally show the formation of vesicles with a well-defined limiting mem-
brane [147]. Similarly, Terech and Talmon [148] have demonstrated the formation of long single-
walled tubes, and mechanisms for the formation of these tubes have been proposed for other bile
acid derivatives [149, 150].
O CH3
Rx O P O (CH2)2 N+ Ry Cx– Cy x,y = 8 – 18
O CH3
Scheme 1.2 Structure of synthetized gemini surfactants. Source: Peresypkin and Menger [153].
The determining factor of the surfactant molecules to self-organize in one or another structure
is the packing parameter P. Israelachvili et al. [124] have indicated that the critical conditions for
cylindrical micelles and planar bilayers are
v 1
(cylindrical micelles) (1.36)
aolc 2
v
1(planar bilayers) (1.37)
aolc
The structure of gemini surfactants [151, 152] provides different ways of changing their physico-
chemical properties. The ionic groups may be positive, negative, or zwitterionic, the spacer may be
rigid or flexible allowing for different separation lengths between the hydrophilic groups, and the
hydrophobic chains may be identical or not. In this last case, the gemini is clearly asymmetric.
Thus, different values can be obtained for the packing parameter. A particularly instructive exam-
ple is the family of zwitterionic geminis synthesized by Peresypkin and Menger [153] in which the
negative group is a phosphodiester and the other is a cationic quaternary ammonium salt
(Scheme 1.2).
Dynamic light scattering experiments suggest that only the C8–C8 derivative originates micelles
exclusively, while others (for example, C8–C12, C12–C8, C14–C8, C10–C14, and C10–C16) self-
assemble in structures such as tubules or vesicles of different sizes, forming also coacervate drop-
lets (C8–C10).
1.5 Biosurfactants
Although phospholipids or bile acids are of a biological origin, the term “biosurfactant” concerns
those amphiphilic derivatives that are produced by microorganisms. In general, they have excel-
lent surface-active properties.
Different classifications have been purposed for biosurfactants [154–156]. For instance, Otzen
indicates that, based on their overall structure, biosurfactants fall into four classes: glycolipids,
lipopeptides, saponins, and all the rest. Glycolipids can be subdivided into rhamnolipids,
sophorolipids, trehalolipids . . ., in which the head group are different saccharides (rhamnose,
sophorose, trehalose, . . .). Similarly, lipopeptides can be divided into several families as surfactin,
iturin or fengycin [157, 158], some of them having a peptide-cycle structure. Gemini type biosur-
factants are also common [159]. The hydrophobic part is commonly one or more unsaturated or
saturated hydrocarbon chains. The saponin Aescin or glycyrrhic acid are examples of biosurfactant
complex structures [159, 160].
The number of biosurfactants can enormously be enlarged by chemical modifications, which
can be carried out in laboratories. Sugar surfactants are well-known examples in which glucose,
1.5 Biosurfactant 21
galactose, xylose, sucrose, or lactose are common hydrophilic heads [161]. They can easily be
obtained chemically [162] or enzymatically [163]. Baccile et al. [26] have obtained and character-
ized the self-assembly properties of a broad family of amino derivatives of sophorolipid biosur-
factants, including asymmetric (sophorose–ammonium and sophorose–amine oxide) and
symmetric (sophorose–sophorose bearing three and four hydrophilic centers) bolaamphiphiles.
Previous naturally found biosurfactants and similar derivatives will all together show a broad
range of physicochemical properties. Let us analyze some significant examples.
The surface tension vs concentration curves of biosurfactants are similar to those of classical
surfactants. The break points in these plots, corresponding to cmc values, are normally well
defined, and the surface tension values at cmc, γcmc are frequently found to be <30 mN/m. However,
differences in reported cmc values are not uncommon, probably due to the presence of impurities
or the use of mixtures instead of pure samples. For instance, Saini et al. [164] reported a value of
54 mg/l for the cmc of viscosin (Figure 1.6), a cyclic oligopeptide lipid which contains nine amino
acids: two (leu-glu) are linked to the fatty acid tail, and the remaining seven [val-leu-ser-leu-ser-ile-
allo(thr)] form a cycle. However, other values have also been reported for cmc, as Saini et al. have
noticed, with values ranging from 4 to 9 mg/l [165] to 150 mg/l [166].
General rules observed in classical surfactants are also observed for biosurfactants. Figure 1.5
shows a linear relationship between the aggregation number and the alkyl chain volume in classi-
cal surfactants. This is just an example of linear relationships for different properties in a series of
homologous surfactants. For instance, Garofalakis et al. [161] have observed a reduction in the
critical aggregation concentration (cac) of the surfactants when increasing the carbon chain length
for a series of monoesters of xylose, galactose, sucrose, and lactose with different hydrophobic
chain lengths (C12–C16). These authors also observed that the more hydrophilic head groups, the
higher is cac, though this trend was moderated by the alkyl chain length. Some observed differ-
ences between maltose and glucose derivatives have been ascribed to a higher degree of hydration
of maltose compared to that of the glucose head group as molar heat capacities for these two sugars
suggest [167]. Another example corresponds to the standard free energy change. For nc-alkyl-D-
maltosides (nc, number of carbon atoms of the alkyl chain, = 8, 10, 12, and 14), Varga et al. [168]
have shown that the dependence of the standard free energy change (and from here the cmc as
well) with the length of the alkyl chain for these sugar surfactants is parallel to the one for alkyltri-
methylammonium bromides and sodium alkylsulfates.
Figure 1.5 Plot of logarithm ln n 5

(aggregation number) vs logarithm
(alkyl chain volume) of sodium alkyl
sulfates (o) and sodium alkyl sulfonates
(•). Source: Original data from Aniansson
4
et al. [145] and Tartar and Lelong [109],
respectively.
2
5 5.4 5.8 6.2
ln v
Leu
O
Ser
HN
H HN H O
O
Leu Val OH
H
NH OH O
H NH Glu
O NH O H O H
H NH N
O N
Ser O H H O
O OH
HN Leu
OH O H
H
allo(thr)
ile
Figure 1.6 Structure of viscosin. Source: Saini et al. [164]. Reproduced by permission of the American
Chemical Society.
Similarly, Ribeiro et al. [169] have obtained diacetylated lactonic sophorolipids with different
hydrophobic chains (C18:0, C18:1, C18:2, and C18:3). The cmc values (in mg/l) increase linearly
with the number of double bonds from 29.2 (C18:0) to 39.1 (C18:3), the slope being 3.35 (r2 = 0.979).
The surface tension at cmc (γcmc) also increases with that number (from 35.7 to 38.8 mN/m).
Sugar alkyl surfactants also display additivities with the number of carbon atoms of the alkyl
chain. For instance, Angarten and Loh [170] studied two series of surfactants CncGx using ITC,
where x = 1,2 is the number of glucoside units (G) of the hydrophilic head and nc = 7–12 (the limits
depending on x). Both families exhibit a linear ΔCp vs nc plot, the slope representing the contribu-
tion of each methylene group to the heat capacity for micellization. The obtained values for the
slope are 63 ± 2 J/(K mol) and 59 ± 2 J/(K mol) for x = 1 and x = 2, respectively. Therefore, within
experimental error, there are not significant differences between them, suggesting that the contribu-
tions of the mono or diglucoside head groups are essentially the same. Let us remember that a value
of 33 J/(K mol) was found for the contribution of each hydrogen atom of each methylene group for
the removal of alkanes from water to air [171]. This also has been related to the number of water
molecules in the first solvation shell that contribute to the thermodynamics of hydrophobic solva-
tion [102]. Other ITC measurements for alkyl sugar surfactants have been carried out by Blume
et al. [92, 172]. Similar studies [173] were carried out for monorhamnose and dirhamnose rham-
nolipids (R1, R2) (and their mixtures) (Figure 1.7). The cmc values are shown at Table 1.2.
Another example of the general behavior of biosurfactants corresponds to the kinetics of the
micelle formation (see above). For instance, Haller and Kaatze [140] have studied the kinetics of
micelle formation in aqueous solution of sugar surfactants as hexyl-, heptyl-, octyl-, nonyl-, and
decyl-β-D-maltopyranoside (CxG2, x = 6, 8–10) as well as of decyl-β-D-maltopyranoside C10G2. As
for other alkyl surfactants, there is a general tendency in the backward rate constant to increase
with increasing cmc and with decreasing length of the alkyl chain.
From small angle neutron scattering (SANS) experiments, Chen et al. [174] found that, in dilute
solution (<20 mM), R1 and R2 form small globular micelles (aggregation numbers being about 50
and 30, respectively), while at higher concentrations, R1 aggregates have a predominantly planar
structure (unilamellar or bilamellar vesicles) whereas R2 remains globular, with an aggregation
number that increases with increasing surfactant concentration. Over the concentration range
1.5 Biosurfactant 23
OH OH
O O HO OH
HO HO O
O OH O OH
O
OH
OH O
O
Figure 1.7 Structure of biosurfactants monorhamnose and dirhamnose rhamnolipids (R1 -left- and R2
right). Source: Chen et al. [174], p. 18281.
measured, for both surfactants the mean thickness of the adsorbed monolayer is about
21 ± 1 Å. From neutron reflectivity (NR) data and the Langmuir isotherm, the molecular areas at
the surface are 62 ± 2 Å2 (R1) and 77 ± 2 Å2 (R2).
Gouzy et al. [24] have obtained two series of asymmetric bipolar surfactants with lactose as
one of the hydrophilic groups. Their structures resemble those of asymmetric gemini surfactants
but without a second hydrophobic moiety (Figure 1.1). These surfactants are known as diva-
lent [26]. They have a long hydrocarbon chain, a nonionic polar head (lactose), a hydrocarbon
spacer (of length nc), and a second polar head (of length mc) at the end of the spacer. At constant
nc, the results evidence a linear variation of log (cmc) with mc but with a positive slope, i.e. the
largest the hydrophobic alkyl side chain, the larger the cmc, which is the opposite trend observed
for classical alkyl surfactants. These results are in line with those observed for gemini surfactants.
Menger and Littau affirm that they are “counter to all previously reported trends in surfactant
chemistry” [151], are anomalous as “in the protected duchies of academia, it is taught that a
longer hydrocarbon tail always lowers the cmc” [152], while Rosen et al. [181] described this
behavior as “aberrant.” These authors accept that this unconventional behavior is indicative of
substantial premicellar aggregation.
As indicated, low critical aggregation values have been measured and they favorably compare
with both ionic and nonionic surfactants. The values obtained for the sucrose hexadecyl and dode-
cyl derivatives by Garofalakis et al. [161] (see Table 1.2) canbe compared with the cmc values for
sodium hexadecyl sulfate (4.5 × 10−4 M) and sodium dodecyl sulfate (8.2 × 10−3 M) [145], as well as
with those for polyoxyethylenated nonionic surfactants of structure CncHnc+1(OC2H4)xOH (CnEx).
For instance, for the series nc = 12, x = 2, Rosen et al. [182] have measured values in the interval
3.3 × 10−5 (x = 2) to 1.09 × 10−4 M (x = 12) (all data at 25 °C), where it is obvious that the larger the
hydrophilic head, the higher the cmc. Similar values for other members of this type of surfactant
have been published elsewhere [183–185].
Some observed differences between maltose and glucose derivatives [161] have been ascribed to
a higher degree of hydration of maltose compared to that of the glucose head group, as molar heat
capacities for these two sugars suggest [167]. Significative differences in the hydration shell of the
uncharged head groups were observed by Tyrode et al. [186, 187]. These authors have studied the
OH stretching region of water molecules in the vicinity of nonionic surfactant monolayers by
using vibrational sum frequency spectroscopy, thus allowing the study of the hydration shell
around the head groups. Compounds such as dodecanol, sugar surfactants (n-decyl-β-D-
glucopyranoside and n-decyl-β-D-maltopyranoside), and polyoxyethylene surfactants (C12E4 and
Table 1.2 Examples of cmc values of biosurfactants.
Compound Cmc/M γ/mN/m References
Sucrose hexadecyl 4.1 × 10−6 31.0–43.0 Garofalakis et al. [161]

Sucrose dodecyl 2.1 × 10−4
R1 monorhamnose rhamnolipid (3.6 ± 0.2) × 10−4 31.2 ± 0.2 Chen et al. [173]
L-rhamnosyl-β-hydroxydecanoyl- 30 °C
β-hydroxydecanoate (RhaC10C10).
R2 L-rhamnosyl-L-rhamnosyl- (1.8 ± 0.2) × 10−4 37.4 ± 0.2
β-hydroxydecanoyl- 30 °C
β-hydroxydecanoate (Rha2C10C10);
The surface tension, NR, and
SANS measurements were all
made at pH 9 (buffer consisted of
0.023 M borax and 0.008 M HCl).
Rhamnolipid A 6.22 × 10−5 pH 7.35 Ishigami et al. [175]
−4
Rhamnolipid B 1.50 × 10 pH 7.35
Rhamnolipid 50 mg/l Whang et al. [176]
−5
Surfactin 4.72 × 10 pH 8.0̧ 20 mM Onaizi et al. [177]
phosphate buffer
Surfactin 45 mg/l <30 Whang et al. [176]
Sophorolipid (lactonic form) 2.8 × 10−5 potassium 36.1 Otto et al. [178]
C18:1 LS phosphate buffer (0.1 M,
pH 7.4) at 25 °C
Diacetyl LS (Sophorolipid) 6 × 10−5 36 Chen et al. [179]
Diacetyl AS (Sophorolipid) 6.7 × 10−4 38.5
Nonacetyl AS (Sophorolipid) 6.2 × 10−4 39
LAS (Sophorolipid) 1.6 × 10−3
SL-p (palmitic) >200 mg/l 35 Ashby et al. [180]
SL-s (stearic) 35 mg/l 35
SL-o (oleic) 140 mg/l 36
SL-l (linoleic) 250 mg/l 36
−3
Glycyrrhizic acid 2.9 × 10 pH 5 55.2 Matsuoka et al. [160]
5.3 × 10−3 pH 6 56.8
No clear cmc at pH 7
C12E8) were studied. For all the surfactants, it was detected that the water molecules located in
proximity to the surfactant hydrocarbon tail phase have both hydrogen atoms free from forming
hydrogen bonds. For the two sugar surfactants, the strength of the hydrogen bonds in the hydra-
tion shell was found to be similar to those observed for tetrahedrally coordinated water molecules
in ice. Despite being itself disordered, the polyoxyethylene head group induces a significant order-
ing and structuring of water at the surface. The orientation of the C12E5 molecules changes with
concentration from lying on the surface with their hydrocarbon tails close to the surface plane (the
observed results being consistent with the formation of disk-like “surface micelles” with a flat
1.6 Sophorolipid 25
orientation of the amphiphiles at low surface concentrations) to a more upright configuration as

the surface covering liquid layer is formed [187].
The formation of surface micelles was discussed in another paper [188] in which surface tension
measurements were used to study the adsorption isotherms for sugar surfactants (n-decyl-β-D-
glucopyranoside (Glu), n-decyl β-D-maltopyranoside (Mal), and n-decyl-β-D-thiomaltopyranoside
(S-Mal)). A gradual change in molecular areas is observed when the surfactant concentration is
increased. As the area/molecule is comparatively large, the resulting surface phase cannot be a
coherent hydrocarbon film and should include a large portion of unperturbed air–water interface.
The formation of surface micelles can account for this observation. A hard-disk simulation allowed
the calculation of the number of molecules per micelle as a function of bulk surfactant concentra-
tion for Mal (values in the interval 9–12) and Glu (values in the interval 10–14), the surfactant
molecules strongly favoring an orientation in the plane of the surface.
1.6 Sophorolipids
The head hydrophilic group of sophorolipids is the disaccharide sophorose (2-O-β-d-glucopyranosyl-

β-d-glucopyranose). It is linked to a hydroxy fatty acyl moiety by a glycosidic bond between the
1′-hydroxy group of the sophorose-sugar and the ω or (ω−1) carbon atom of the fatty acid.
Sophorolipids were first isolated by Gorin et al. [189] from the oil formed during fermentation by
a strain of Torulopsis magnoliae. It has been demonstrated that they have taste-sensory proper-
ties [190]. Sophorolipids may have a free carboxylic acid (AS) structure or a lactone (LS) one.
Figure 1.8 shows both structures for the derivative with a C18:1 (oleic) chain.
Ashby et al. [180] have obtained other derivatives by fed-batch fermentation of Candida bom-
bicola on glucose and several fatty acids as palmitic acid (SL-p), stearic acid (SL-s), oleic acid (SL-o),
and linoleic acid (SL-l). The cmc values obtained by these authors are shown in Table 1.2. The exact
composition can vary with the type of hydrocarbon substrate used in the sophorolipid production
and the production conditions [178], and correspondingly different cmc values have been pub-
lished. For a pure diacetylated C18:1 LS, Otto et al. [178] have reported a cmc value of 2.8 × 10−5 M
(Table 1.2). Higher values have been published by Chen et al. [179] for diacetyl LS, diacetyl AS, and
nonacetyl AS.
AcO
HO HO HO
AcO
AcO HO O O OH
O O
HO O O OH O O OH
O O
HO OH O
OAc
Figure 1.8 Chemical structures of the acidic (AS) and lactonic (LS) C18:1 sophorolipids.
Penfold et al. [191] have studied sophorolipids by SANS. At low surfactant concentrations
(0.2–3 mM), data for LS are consistent with the formation of small unilamellar vesicles, with inner
and outer radii increasing with concentration. The shell thickness also increases from about 15 to
24 Å. At high concentrations (30 mM), dynamic light scattering measurements are consistent with
large aggregates (~300 nm). The solutions of AS with one and two acetyl groups have a hazy appear-
ance, indicating the presence of large aggregates, while the solution of AS with no acetyl groups
are consistent with small micellar structures, the aggregation number increasing steadily from 28
to 40 at the concentration range 5–50 mM. These results are consistent with predictions from the
packing parameter.
At the air–solution interface, NR measurements were also carried out by Chen et al. for the deu-
terated surfactants (d-LS and d-AS) [179]. This technique provides different parameters as adsorbed
amounts, composition, thickness of the adsorbed layer, and structure at the surface. The adsorp-
tion obtained values are consistent with a Langmuir isotherm (Eq. (1.38))
max C
(1.38)
C k
where Γ and Γmax are the adsorbed amounts and the maximum adsorption, C is the surfactant
concentration, and k is the adsorption coefficient. AS and LS have similar k values (2.2 × 10−6), sug-
gesting that both sophorolipids have similar affinities for the air–water interface. Above cmc, the
thickness is around 23 Å while the area/molecule is around 74 Å2. For the less hydrophobic AS, the
authors obtained a value of 85 Å2. These results for the adsorbed amount are in good agreement
with the values obtained from surface tension data.
Studies by Manet et al. [192] have shown that the micellar morphology of no acetylated C18:1 AS
is a prolate ellipsoid. Depending on experimental conditions (the salts cause an increase of the
aggregation number and an elongation of the micellar aggregates), the equatorial radius of the
ellipsoid varies between 6.1 and 8.0 Å, the axial core ratio varies between 4.7 and 9.4, and the aggre-
gation number between 24 and 73. The fraction of CH2 groups inserted in the dry core of the
micelle is in the interval 0.5–0.7, meaning that the core/shell interface is located far from the sugar
head group. However, the equatorial shell thickness is almost constant (12.0 ± 0.5 Å). The shell
thickness that best describes the sophorolipid micelles is a variable one from the equatorial value
given above to zero, i.e. the hydrophilic shell has a nonhomogeneous distribution of matter con-
taining carboxylic groups, sophorose, salt, water, and part of the aliphatic chain. This is an atypical
result since most of surfactant systems are described by a homogeneous shell thickness. The area
per sophorolipid between the alkyl chain and the sugar/carboxylate head group has been esti-
mated between 102 and 141 Å2 for the most ionized micelles. For nonacetyl AS, Chen et al. [179]
reported a value of 104 ± 8 Å2 for the area at the air−water interface.
Previously, Cecutti et al. [193] had noticed that the sugar rings represent a major part of the
molecular volume for glycolipids and, consequently, they differentiate between the micelle-solvent
interface and the hydrophobic core-sugar head group interface. The best result for interpreting
neutron and X-ray small-angle scattering intensity curves for -dodecyl maltoside in water (6% w/v,
310 K) is by a short ellipsoid model with an ellipticity of 1.2. The difference between the total short
radius of the micelle (24 Å), and the short radius of the apolar hydrophobic core (18 Å) allows
enough space for the sugar head groups. Other obtained parameters are the areas per surfactant
head at the water-micelles interface (87 Å2), and at the chain-head group interface (50 Å2), the
aggregation number (82) and the number of water molecules per surfactant molecules (10).
AS has a bolaamphiphile asymmetrical structure with cis-9-octadecenoic chain linking the
sophorose disaccharide and the carboxylic acid groups. Zhou et al. [194] have observed that, at a
1.6 Sophorolipid 27
concentration of AS of 2.0 mg/ml and pH 2.0, ribbon formation may be noticed by a light micro-
scope. The length and the width of the ribbons grow with time, and after two hours, twisted rib-
bons with lengths of a few hundred micrometers and a width of ~5 m had formed. Ribbon formation
is slowed by increasing the pH. After a time, which depends on pH, precipitates were observed.
After 28 hours, at pH 4.1 dynamic light scattering measurements of the solution showed that the
large aggregates coexisted with small micelles. Small-angle X-ray scattering (SAXS) and wide-
angle X-ray scattering (WAXS) measurements were carried out in aqueous solutions and dried
solid ribbons formed at pH 4.1. All WAXS diffractograms indicate high crystallinity of the hydro-
carbon chains and the disaccharide head groups inside the ribbons. At pH 5.1 individual ribbons
were rarely found. At 0.02 mg/ml and pH 7.8 (at which the carboxylic groups are in their negative
carboxylate form), small aggregates are formed, the hydrodynamic radius Rh being 37 nm (meas-
ured at a scattering angle of 15°). At concentrations of 0.97–1.78 mg/ml, nearly monodisperse
micellar aggregates were formed with Rh about 100 nm. An apparent radius of gyration Rg of
175 nm was estimated for the radius of gyration by measuring the scattering intensity at the angle
range of 15–35° at a surfactin concentration of 1.40 mg/ml. The Rg/Rh ratio is about 1.75, a rather
large value which indicates that the large micellar aggregates have a very anisotropic geometry.
As described in the previous paragraph, the carboxylic acid group of C18:1 AS makes that its
aggregation behavior is sensitive to pH. Baccile et al. [195] have studied the system by SANS at dif-
ferent amounts of NaOH and other bases (NH3, KOH, and Ca(OH)2). A core–shell prolate micelle
structure with an interaction potential (which combines hard-sphere and screened Coulomb
potentials) was used to fit SANS data. The total effective cross-radius of the micelle (core
radius + shell thickness) decreases (from 21.8 to 18.2 Å) with increasing NaOH, the core radius
contributing the most to this reduction as the shell is practically constant (~8 Å). The core size and
the length of the oleic acid chain suggest that the chain is partially folded and that the formation
of more carboxylate/Na+ pairs favors such a bending. The eccentricity of the prolate also reduces
(from 3.3 to 2.6) and micelles become more negatively charged as the effective micellar charge var-
ies from about −0.5 to −5.3 with increasing NaOH concentration. Baccile et al. have also noticed
that in the presence of Ca(OH)2, the system evolves toward a better surface charge screening,
which has the effect of reducing the repulsive potential (between micelles) and the effective sur-
face charge. The micellar length is also elongated.
In a nice piece of work, Baccile et al. [26] prepared a broad range (38 new compounds) of amino
derivatives of sophorolipid biosurfactant, comprising quaternary ammonium salts, amine oxides,
and symmetric and asymmetric bolaamphiphiles with three or four hydrophilic centers, as well as
divalent and Y-shaped derivatives. The compounds are constituted by at least two hydrophilic head
groups, different in nature and charge since sophorose is neutral and the nitrogen atom is charged,
being separated by a spacer. SAXS experiments were used for the estimation of the aggrega-
tion number.
Bolaform derivatives with two (sophorose and ammonium) hydrophilic groups either do not
form aggregated or exhibit a poor tendency to self-aggregate. Correspondingly, the aggregation
numbers are small (<20) and the aggregates are highly hydrated. The behavior of compounds with
three hydrophilic groups (sophorose–ammonium–sophorose) depends on the nature of the ammo-
nium group. If this is small, the compounds behave as the bolaform derivatives with two groups. If
the group is charged and bulky, small hydrated aggregates are observed, coexisting with fibrilary
systems. Tetra-center compounds (sophorose–ammonium–ammonium–sophorose) with small
and/or charged ammonium groups, have a poor tendency for self-aggregation, and again small
aggregation numbers and highly hydrated micelles are observed. However, divalent and Y-shaped
surfactants tend to form larger micellar aggregates as a function of the size of the hydrophobic
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the acquirement of knowledge; he sought to prepare the vase rather
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But in these days of revolutionary thought, when all along the line
—in geology and anthropology, chemistry, philology, and biology—
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for, supposing that education really did consist in systematised
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develop on the same lines, be as like as “two peas,” and (should we
not?) die of weariness of one another! Some of us have an uneasy
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indeed, the fear is groundless. We may believe that the personality,
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The problem of education is more complex than it seems at first
sight, and well for us and the world that it is so. “Education is a life;”
you may stunt and starve and kill, or you may cherish and sustain;
but the beating of the heart, the movement of the lungs, and the
development of the faculties (are there any “faculties”?) are only
indirectly our care. The poverty of our thought on the subject of
education is shown by the fact that we have no word which at all
implies the sustaining of a life: education (e, out, and ducere, to lead,
to draw) is very inadequate; it covers no more than those occasional
gymnastics of the mind which correspond with those by which the
limbs are trained: training (trahere) is almost synonymous, and upon
these two words rests the misconception that the development and
the exercise of the “faculties” is the object of education (we must
needs use the word for want of a better). Our homely Saxon
“bringing up” is nearer the truth, perhaps because of its very
vagueness; any way, “up” implies an aim, and “bringing” an effort.
The happy phrase of Mr. Matthew Arnold—“Education is an
atmosphere, a discipline, a life”—is perhaps the most complete and
adequate definition of education we possess. It is a great thing to
have said it; and our wiser posterity may see in that “profound and
exquisite remark” the fruition of a lifetime of critical effort. Observe
how it covers the question from the three conceivable points of view.
Subjectively, in the child, education is a life; objectively, as affecting
the child, education is a discipline; relatively, if we may introduce a
third term, as regards the environment of the child, education is an
atmosphere.
We shall examine each of these postulates later; at present we
shall attempt no more than to clear the ground a little, with a view to
the subject of this paper, “Parents as Inspirers”—not “modellers,” but
“inspirers.”
It is only as we recognise our limitations that our work becomes
effective: when we see definitely what we are to do, what we can do,
and what we cannot do, we set to work with confidence and courage;
we have an end in view, and we make our way intelligently towards
that end, and a way to an end is method. It rests with parents not
only to give their children birth into the life of intelligence and moral
power, but to sustain the higher life which they have borne. Now that
life, which we call education, receives only one kind of sustenance; it
grows upon ideas. You may go through years of so-called
“education” without getting a single vital idea; and that is why many a
well-fed body carries about a feeble, starved intelligence; and no
society for the prevention of cruelty to children cries shame on the
parents. Only the other day we heard of a girl of fifteen who had
spent two years at a school without taking part in a single lesson,
and this by the express desire of her mother, who wished all her time
and all her pains to be given to “fancy needlework.” This, no doubt, is
a survival (not of the fittest), but it is possible to pass even the
Universities’ Local Examinations with credit, without ever having
experienced that vital stir which marks the inception of an idea; and,
if we have succeeded in escaping this disturbing influence, why we
have “finished our education” when we leave school; we shut up our
books and our minds, and remain pigmies in the dark forest of our
own dim world of thought and feeling.
What is an idea? A live thing of the mind, according to the older
philosophers, from Plato to Bacon, from Bacon to Coleridge. We say
of an idea that it strikes us, impresses us, seizes us, takes
possession of us, rules us; and our common speech is, as usual,
truer to fact than the conscious thought which it expresses. We do
not in the least exaggerate in ascribing this sort of action and power
to an idea. We form an ideal—a, so to speak, embodied idea—and
our ideal exercises the very strongest formative influence upon us.
Why do you devote yourself to this pursuit, that cause? “Because
twenty years ago such and such an idea struck me,” is the sort of
history which might be given of every purposeful life—every life
devoted to the working out of an idea. Now is it not marvellous that,
recognising as we do the potency of an idea, both the word and the
conception it covers enter so little into our thought of education?
Coleridge brings the conception of an “idea” within the sphere of
the scientific thought of to-day; not as that thought is expressed in
Psychology—a term which he himself launched upon the world with
an apology for it as an insolens verbum,[1] but in that science of the
correlation and interaction of mind and brain, which is at present
rather clumsily expressed in such terms as “mental physiology” and
“psycho-physiology.”
In his method he gives us the following illustration of the rise and
progress of an idea:—
“We can recall no incident of human history that impresses the
imagination more deeply than the moment when Columbus, on an
unknown ocean, first perceived that startling fact, the change of the
magnetic needle. How many such instances occur in history, when
the ideas of nature (presented to chosen minds by a Higher Power
than Nature herself) suddenly unfold, as it were, in prophetic
succession, systematic views destined to produce the most
important revolutions in the state of man! The clear spirit of
Columbus was doubtless eminently methodical. He saw distinctly
that great leading idea which authorised the poor pilot to become a
‘promiser of kingdoms.’”
Notice the genesis of such ideas—“presented to chosen minds by
a Higher Power than Nature;” notice how accurately this history of an
idea fits in with what we know of the history of great inventions and
discoveries, with that of the ideas which rule our own lives; and how
well does it correspond with that key to the origin of “practical” ideas
which we find elsewhere:—
“Doth the plowman plow continually to ... open and break the
clods of his ground? When he hath made plain the face thereof, doth
he not cast abroad the fitches, and scatter the cummin, and put in
the wheat in rows, and the barley in the appointed place, and the
spelt in the border thereof? For his God doth instruct him aright, and
doth teach him....
“Bread corn is ground; for he will not ever be threshing it.... This
also cometh forth from the Lord of hosts, which is wonderful in
counsel and excellent in wisdom.”[2]
Ideas may invest as an atmosphere, rather than strike as a
weapon. “The idea may exist in a clear, distinct definite form, as that
of a circle in the mind of a geometrician; or it may be a mere instinct,
a vague appetency towards something, ... like the impulse which fills
the young poet’s eyes with tears, he knows not why.” To excite this
“appetency towards something”—towards things lovely, honest, and
of good report, is the earliest and most important ministry of the
educator. How shall these indefinite ideas which manifest
themselves in appetency be imparted? They are not to be given of
set purpose, nor taken at set times. They are held in that thought-
environment which surrounds the child as an atmosphere, which he
breathes as his breath of life; and this atmosphere in which the child
inspires his unconscious ideas of right living emanates from his
parents. Every look of gentleness and tone of reverence, every word
of kindness and act of help, passes into the thought-environment,
the very atmosphere which the child breathes; he does not think of
these things, may never think of them, but all his life long they excite
that “vague appetency towards something” out of which most of his
actions spring. Oh! the wonderful and dreadful presence of the little
child in the midst.
That he should take direction and inspiration from all the casual
life about him, should make our poor words and ways the starting-
point from which, and in the direction of which, he develops—this is
a thought which makes the most of us hold our breath. There is no
way of escape for parents; they must needs be as “inspirers” to their
children, because about them hangs, as its atmosphere about a
planet, the thought-environment of the child, from which he derives
those enduring ideas which express themselves as a life-long
“appetency” towards things sordid or things lovely, things earthly or
divine.
Let us now hear Coleridge on the subject of those definite ideas
which are not inhaled as air, but conveyed as meat to the mind:—[3]
“From the first, or initiative idea, as from a seed, successive ideas
germinate.”
“Events and images, the lively and spirit-stirring machinery of the
external world, are like light, and air, and moisture to the seed of the
mind, which would else rot and perish.”
“The paths in which we may pursue a methodical course are
manifold, and at the head of each stands its peculiar and guiding
idea.”
“Those ideas are as regularly subordinate in dignity as the paths
to which they point are various and eccentric in direction. The world
has suffered much, in modern times, from a subversion of the natural
and necessary order of Science ... from summoning reason and faith
to the bar of that limited physical experience to which, by the true
laws of method, they owe no obedience.”
“Progress follows the path of the idea from which it sets out;
requiring, however, a constant wakefulness of mind to keep it within
the due limits of its course. Hence the orbits of thought, so to speak,
must differ among themselves as the initiative ideas differ.”
Have we not here the corollary to, and the explanation of, that law
of unconscious cerebration which results in our “ways of thinking,”
which shapes our character, rules our destiny? Thoughtful minds
consider that the new light which biology is throwing upon the laws of
mind is bringing to the front once more the Platonic doctrine, that “An
idea is a distinguishable power, self-affirmed, and seen in its unity
with the Eternal Essence.”
The whole subject is profound, but as practical as it is profound.
We absolutely must disabuse our minds of the theory that the
functions of education are, in the main, gymnastic. In the early years
of the child’s life it makes, perhaps, little apparent difference whether
his parents start with the notion that to educate is to fill a receptacle,
inscribe a tablet, mould plastic matter, or nourish a life; but in the end
we shall find that only those ideas which have fed his life are taken
into the being of the child; all the rest is thrown away, or worse, is
like sawdust in the system, an impediment and an injury to the vital
processes.
This is, perhaps, how the educational formula should run:
Education is a life; that life is sustained on ideas; ideas are of
spiritual origin; but,
“God has made us so,”
that we get them chiefly as we convey them to one another. The duty
of parents is to sustain a child’s inner life with ideas as they sustain
his body with food. The child is an eclectic; he may choose this or
that; therefore, in the morning sow thy seed, and in the evening
withhold not thy hand, for thou knowest not which shall prosper,
whether this or that, or whether they both shall be alike good.
The child has affinities with evil as well as with good; therefore,
hedge him about from any chance lodgment of evil suggestion.
The initial idea begets subsequent ideas; therefore, take care that
children get right primary ideas on the great relations and duties of
life.
Every study, every line of thought, has its “guiding idea;” therefore
the study of a child makes for living education, as it is quickened by
the guiding idea “which stands at the head.”
In a word, our much boasted “infallible reason”—is it not the
involuntary thought which follows the initial idea upon necessary
logical lines? Given, the starting idea, and the conclusion may be
predicated almost to a certainty. We get into the way of thinking such
and such manner of thoughts, and of coming to such and such
conclusions, ever further and further removed from the starting-point,
but on the same lines. There is structural adaptation in the brain
tissue to the manner of thoughts we think—a plan and a way for
them to run in. Thus we see how the destiny of a life is shaped in the
nursery, by the reverent naming of the Divine Name; by the light
scoff at holy things; by the thought of duty the little child gets who is
made to finish conscientiously his little task; by the hardness of heart
that comes to the child who hears the faults or sorrows of others
spoken of lightly.
FOOTNOTES:
[1] “We beg pardon for the use of this insolens verbum, but it is
one of which our language stands in great need.”—S. T.
Coleridge.
[2] Isaiah xxviii.
[3] Method—S. T. Coleridge.
CHAPTER V
PARENTS AS INSPIRERS
Part III
It is probable that parents as a class feel more than ever before the
responsibility of their prophetic office. It is as revealers of God to
their children that parents touch their highest limitations; perhaps it is
only as they succeed in this part of their work that they fulfil the
Divine intention in giving them children to bring up—in the nurture
and admonition of the Lord.
How to fortify the children against the doubts of which the air is
full, is an anxious question. Three courses are open—to teach as we
of an older generation have been taught, and to let them bide their
time and their chance; to attempt to deal with the doubts and
difficulties which have turned up, or are likely to turn up; or, to give
children such hold upon vital truth, and, at the same time, such an
outlook upon current thought, that they shall be landed on the safe
side of the controversies of their day, open to truth, in however new a
light presented, and safeguarded against mortal error.
The first course is unfair to the young: when the attack comes,
they find themselves at a disadvantage; they have nothing to reply;
their pride is in arms; they jump to the conclusion that there is no
defence possible of that which they have received as truth; had there
been, would they not have been instructed to make it? They resent
being made out in the wrong, being on the weaker side—so it seems
to them,—being behind their times; and they go over without a
struggle to the side of the most aggressive thinkers of their day.
Let us suppose that, on the other hand, they have been fortified
with “Christian Evidences,” defended by bulwarks of sound dogmatic
teaching. Religion without definite dogmatic teaching degenerates
into sentiment, but dogma, as dogma, offers no defence against the
assaults of unbelief. As for “evidences,” the rôle of the Christian
apologist is open to the imputation conveyed in the keen proverb, qui
s’excuse, s’accuse; the truth by which we live must needs be self-
evidenced, admitting of neither proof nor disproof. Children should
be taught Bible history with every elucidation which modern research
makes possible. But they should not be taught to think of the
inscriptions on the Assyrian monuments, for example, as proofs of
the truth of the Bible records, but rather as illustrations, though they
are, and cannot but be subsidiary proofs.
Let us look at the third course; and first, as regards the outlook
upon current thought. Contemporary opinion is the fetish of the
young mind. Young people are eager to know what to think on all the
serious questions of religion and life. They ask what is the opinion of
this and that leading thinker of their day. They by no means confine
themselves to such leaders of thought as their parents have elected
to follow; on the contrary, the “other side” of every question is the
attractive side for them, and they do not choose to be behind the
foremost in the race of thought.
Now, that their young people should thus take to the water need
not come upon parents as a surprise. The whole training from
babyhood upward should be in view of this plunge. When the time
comes, there is nothing to be done; openly, it may be, secretly if the
home rule is rigid, the young folk think their own thoughts; that is,
they follow the leader they have elected; for they are truly modest
and humble at heart, and do not yet venture to think for themselves;
only they have transferred their allegiance. Nor is this transfer of
allegiance to be resented by parents; we all claim this kind of
“suffrage” in our turn when we feel ourselves included in larger
interests than those of the family.
But there is much to be done beforehand, though nothing when
the time comes. The notion that any contemporary authority is
infallible may be steadily undermined from infancy onwards, though
at some sacrifice of ease and glory to the parents. “I don’t know”
must take the place of the vague wise-sounding answer, the random
shot which children’s pertinacious questionings too often provoke.
And “I don’t know” should be followed by the effort to know, the
research necessary to find out. Even then, the possibility of error in a
“printed book” must occasionally be faced. The results of this kind of
training in the way of mental balance and repose are invaluable.
Another safeguard is in the attitude of reservation, shall we say?
which it may be well to preserve towards “Science.” It is well that the
enthusiasm of children should be kindled, that they should see how
glorious it is to devote a lifetime to patient research, how great to find
out a single secret of nature, a key to many riddles. The heroes of
science should be their heroes; the great names, especially of those
who are amongst us, should be household words. But here, again,
nice discrimination should be exercised; two points should be kept
well to the front—the absolute silence of the oracle on all ultimate
questions of origin and life, and the fact that, all along the line,
scientific truth comes in like the tide, with steady advance, but with
ebb and flow of every wavelet of truth; so much so, that, at the
present moment, the teaching of the last twenty years is discredited
in at least half a dozen departments of science. Indeed, it would
seem to be the part of wisdom to wait half a century before fitting the
discovery of to-day into the general scheme of things. And this, not
because the latest discovery is not absolutely true, but because we
are not yet able so to adjust it—according to the “science of the
proportion of things”—that it shall be relatively true.
But all this is surely beyond children? By no means; every walk
should quicken their enthusiasm for the things of nature, and their
reverence for the priests of that temple; but occasion should be
taken to mark the progressive advances of science, and the fact that
the teaching of to-day may be the error of to-morrow, because new
light may lead to new conclusions even from the facts already
known. “Until quite lately, geologists thought ... they now think ... but
they may find reason to think otherwise in the future.” To perceive
that knowledge is progressive, and that the next “find” may always
alter the bearings of what went before; that we are waiting, and may
have very long to wait, for the last word; that science also is
“revelation,” though we are not yet able fully to interpret what we
know; and that ‘science’ herself contains the promise of great
impetus to the spiritual life—to perceive these things is to be able to
rejoice in all truth and to wait for final certainty.
In another way we may endeavour to secure for the children that
stability of mind which comes of self-knowledge. It is well that they
should know so early, that they will seem to themselves always to
have known, some of the laws of thought which govern their own
minds. Let them know that, once an idea takes possession of them,
it will pursue, so to speak, its own course, will establish its own place
in the very substance of the brain, will draw its own train of ideas
after it. One of the most fertile sources of youthful infidelity is the fact
that thoughtful boys and girls are infinitely surprised when they come
to notice the course of their own thoughts. They read a book or listen
to talk with a tendency to what is to them “free-thought.” And then,
the “fearful joy” of finding that their own thoughts begin with the
thought they have heard, and go on and on to new and startling
conclusions on the same lines! The mental stir of all this gives a
delightful sense of power, and a sense of inevitableness and
certainty too; for they do not intend or try to think this or that. It
comes of itself; their reason, they believe, is acting independently of
them, and how can they help assuming that what comes to them of
itself, with an air of absolute certainty, must of necessity be right?
But what if from childhood they had been warned, “Take care of
your thoughts, and the rest will take care of itself; let a thought in,
and it will stay; will come again to-morrow and the next day, will
make a place for itself in your brain, and will bring many other
thoughts like itself. Your business is to look at the thoughts as they
come, to keep out the wrong thoughts, and let in the right. See that
ye enter not into temptation.” This sort of teaching is not so hard to
understand as the rules for the English nominative, and is of
infinitively more profit in the conduct of life. It is a great safeguard to
know that your “reason” is capable of proving any theory you allow
yourself to entertain.
We have touched here only on the negative side of the parent’s
work as prophet, inspirer. There are perhaps few parents to whom
the innocence of the babe in its mother’s arms does not appeal with
pathetic force. “Open me the gates of righteousness, that I may go in
unto them,” is the voice of the little unworldly child; and a wish,
anyway, that he may be kept unspotted from the world is breathed in
every kiss of his mother, in the light of his father’s eyes. But how
ready we are to conclude that children cannot be expected to
understand spiritual things. Our own grasp of the things of the Spirit
is all too lax, and how can we expect that the child’s feeble
intelligence can apprehend the highest mysteries of our being? But
here we are altogether wrong. It is with the advance of years that a
materialistic temper settles upon us. But the children live in the light
of the morning-land. The spirit-world has no mysteries for them; that
parable and travesty of the spirit-world, the fairy-world, where all
things are possible, is it not their favourite dwelling-place? And fairy
tales are so dear to children because their spirits fret against the
hard and narrow limitations of time and place and substance; they
cannot breathe freely in a material world. Think what the vision of
God should be to the little child already peering wistfully through the
bars of his prison-house. Not a far-off God, a cold abstraction, but a
warm, breathing, spiritual Presence about his path and about his bed
—a Presence in which he recognises protection and tenderness in
darkness and danger, towards which he rushes as the timid child to
hide his face in his mother’s skirts.
A friend tells me the following story of her girlhood. It so
happened that extra lessons detained her at school until dark every
day during the winter. She was extremely timid, but, with the
unconscious reserve of youth, never thought of mentioning her fear
of “something.” Her way home lay by a river-side, a solitary path
under trees—big trees, with masses of shadow. The black shadows,
in which “something” might lie hid—the swsh-sh, swsh-sh of the
river, which might be whisperings or the rustle of garments—filled
her night by night with unabated terror. She fled along that river-side
path with beating heart; but, quick as flying steps and beating heart,
these words beat in her brain, over, and over, and over, the whole
length of the way, evening by evening, winter after winter: “Thou art
my hiding-place; Thou shalt preserve me from trouble; Thou shalt
compass me about with songs of deliverance.” Years after, when the
woman might be supposed to have outgrown girlish terrors, she
found herself again walking alone in the early darkness of a winter’s
evening under trees by the swsh-sh of another river. The old terror
returned, and with it the old words came to her, and kept time the
whole length of the way with her hasty steps. Such a place to hide
him in should be the thought of God to every child.
Their keen sensitiveness to spiritual influences is not due to
ignorance on the part of the children. It is we, not they, who are in
error. The whole tendency of modern biological thought is to confirm
the teaching of the Bible: the ideas which quicken come from above;
the mind of the little child is an open field, surely “good ground,”
where, morning by morning, the sower goes forth to sow, and the
seed is the Word. All our teaching of children should be given
reverently, with the humble sense that we are invited in this matter to
co-operate with the Holy Spirit; but it should be given dutifully and
diligently, with the awful sense that our co-operation would appear to
be made a condition of the Divine action; that the Saviour of the
world pleads with us to “Suffer the little children to come unto Me,”
as if we had the power to hinder, as we know that we have.
This thought of the Saviour of the world implies another
conception which we sometimes leave out of sight in dealing with
children. Young faces are not always sunny and lovely; even the
brightest children in the happiest circumstances have their clouded
hours. We rightly put the cloud down to some little disorder, or to the
weather, but these are the secondary causes which reveal a deep-
seated discontent. Children have a sense of sin acute in proportion
to their sensitiveness. We are in danger of trusting too much to a
rose-water treatment; we do not take children seriously enough;
brought face to face with a child, we find he is a very real person, but
in our educational theories we take him as “something between a
wax doll and an angel.” He sins; he is guilty of greediness,
falsehood, malice, cruelty, a hundred faults that would be hateful in a
grown-up person; we say he will know better by-and-by. He will
never know better; he is keenly aware of his own odiousness. How
many of us would say about our childhood, if we told the whole truth,
“Oh, I was an odious little thing!” and that, not because we recollect
our faults, but because we recollect our childish estimate of
ourselves. Many a bright and merry child is odious in his own eyes;
and the “peace, peace, where there is no peace,” of fond parents
and friends is little comfort. It is well that we “ask for the old paths,
where is the good way;” it is not well that, in the name of the old
paths, we lead our children into blind alleys, nor that we let them
follow the new into bewildering mazes.
CHAPTER VI
PARENTS AS INSPIRERS
Part IV
“One of the little boys gazing upon the terrible desolation of the scene, so unlike
in its savage and inhuman aspects anything he had ever seen at home, nestled
close to his mother, and asked with bated breath, ‘Mither, is there a God here?’”—
John Burroughs.
The last chapter introduced the thought of parents in their highest
function—as revealers of God to their children. To bring the human
race, family by family, child by child, out of the savage and inhuman
desolation where He is not, into the light and warmth and comfort of
the presence of God, is, no doubt, the chief thing we have to do in
the world. And this individual work with each child, being the most
momentous work in the world, is put into the hands of the wisest,
most loving, disciplined, and divinely instructed of human beings. Be
ye perfect as your Father is perfect, is the perfection of parenthood,
perhaps to be attained only in its fulness through parenthood. There
are mistaken parents, ignorant parents, a few indifferent parents,
even, as one in a thousand, callous parents; but the good that is
done upon the earth is done, under God, by parents, whether directly
or indirectly.
Parents, who recognise that their great work is to be done by the
instrumentality of the ideas they are able to introduce into the minds
of their children, will take anxious thought as to those ideas of God
which are most fitting for children, and as to how those ideas may
best be conveyed. Let us consider an idea which is just now causing
some stir in people’s thoughts.
“We read some of the Old Testament history as ‘history of the
Jews,’ and Job and Isaiah and the Psalms as poetry—and I am glad
to say he is very fond of them; and parts of the Gospels in Greek, as
the life and character of a hero. It is the greatest mistake to impose
them upon children as authoritative and divine all at once. It at once
diminishes their interest: we ought to work slowly up through the
human side.”[4]
Here is a theory which commends itself to many persons because
it is “so reasonable.” But it goes upon the assumption that we are
ruled by Reason, an infallible entity, which is certain, give it fair play,
to bring us to just conclusions. Now the exercise of that function of
the mind which we call reasoning—we must decline to speak of “the
Reason”—does indeed bring us to inevitable conclusions; the
process is definite, the result convincing; but whether that result be
right or wrong depends altogether upon the initial idea which, when
we wish to discredit it, we call a prejudice; when we wish to exalt, we
call an intuition, even an inspiration. It would be idle to illustrate this
position; the whole history of Error is the history of the logical
outcome of what we happily call misconceptions. The history of
Persecution is the tale of how the inevitable conclusions arrived at by
reasoning pass themselves off for truth. The Event of Calvary was
due to no hasty mad outburst of popular feeling. It was a triumph of
reasoning: the inevitable issue of more than one logical sequence;
the Crucifixion was not criminal, but altogether laudable, if that is
right which is reasonable. And this is why the hearts of religious
Jews were hardened and their understanding darkened; they were
truly doing what was right in their own eyes. It is a marvellous thing
to perceive the thoughts within us driving us forward to an inevitable
conclusion, even against our will. How can that conclusion which
presents itself to us in spite of ourselves fail to be right?
Let us place ourselves for one instant in the position of the logical
and conscientious Jew. “‘Jehovah’ is a name of awe,
unapproachable in thought or act except in ways Himself has
specified. To attempt unlawful approach is to blaspheme. As
Jehovah is infinitely great, presumptuous offence is infinitely
heinous, is criminal, is the last crime as committed against Him who
is the First. The blasphemer is worthy of death. This man makes
himself equal with God, the unapproachable. He is a blasphemer,
arrogant as Beelzebub. He is doubly worthy of death. To the people
of the Jews is committed in trust the honoured Name; upon them it is
incumbent to exterminate the blasphemer. The man must die.” Here
is the secret of the virulent hatred which dogged the steps of the
blameless Life. These men were following the dictates of reason,
and knew, so they would say, that they were doing right. Here we
have the invincible ignorance which the Light of the world failed to
illumine; and He,
“Who knows us as we are,
Yet loves us better than He knows,”
offers for them the true plea, “They know not what they do.” The
steps of the argument are incontrovertible; the error lies in the initial
idea,—such conception of Jehovah as made the conception of Christ
inadmissible, impossible. Thus reasoned the Jew upon whom his
religion had the first claim. The patriotic Jew, to whom religion itself
was subservient to the hopes of his nation, arrived by quite another
chain of spontaneous arguments at the same inevitable conclusion:
—“The Jews are the chosen people. The first duty of a Jew is
towards his nation. These are critical times. A great hope is before
us, but we are in the grip of the Romans; they may crush out the
national life before our hope is realised. Nothing must be done to
alarm their suspicions. This Man? By all accounts He is harmless,
perhaps righteous. But He stirs up the people. It is rumoured that
they call Him King of the Jews. He must not be permitted to ruin the
hopes of the nation. He must die. It is expedient that one man die for
the people, and that the whole nation perish not.” Thus the
consummate crime that has been done upon the earth was done
probably without any consciousness of criminality; on the contrary,
with the acquittal of that spurious moral sense which supports with
its approval all reasonable action. The Crucifixion was the logical
and necessary outcome of ideas imbibed from their cradles by the
persecuting Jews. So of every persecution; none is born of the
occasion and the hour, but comes out of the habit of thought of a
lifetime.
It is the primal impulse to these habits of thought which children
must owe to their parents; and, as a man’s thought and action
Godward is—
“The very pulse of the machine,”
the introduction of such primal ideas as shall impel the soul to God is
the first duty and the highest privilege of parents. Whatever sin of
unbelief a man is guilty of, are his parents wholly without blame? Let
us consider what is commonly done in the nursery in this respect. No
sooner can the little being lisp than he is taught to kneel up in his
mother’s lap, and say “God bless ...” and then follows a list of the
near and dear, and “God bless ... and make him a good boy, for
Jesus’ sake. Amen.” It is very touching and beautiful. I once peeped
in at an open cottage door in a moorland village, and saw a little
child in its nightgown kneeling in its mother’s lap and saying its
evening prayer. The spot has ever since remained to me a sort of
shrine. There is no sight more touching and tender. By-and-by, so
soon as he can speak the words,
“Gentle Jesus, meek and mild,”
is added to the little one’s prayer, and later, “Our Father.” Nothing
could be more suitable and more beautiful than these morning and
evening approaches to God, the little children brought to Him by their
mothers. And most of us can “think back” to the hallowing influence
of these early prayers. But might not more be done? How many
times a day does a mother lift up her heart to God as she goes in
and out amongst her children, and they never know! “To-day I talked
to them” (a boy and girl of four and five) “about Rebekah at the well.
They were very much interested, especially about Eliezer praying in
his heart and the answer coming at once. They said, ‘How did he
pray?’ I said, ‘I often pray in my heart when you know nothing about
it. Sometimes you begin to show a naughty spirit, and I pray for you
in my heart, and almost directly I find the good Spirit comes, and
your faces show my prayer is answered.’ O. stroked my hand and
said, ‘Dear mother, I shall think of that!’ Boy looked thoughtful, but
didn’t speak; but when they were in bed I knelt down to pray for them
before leaving them, and when I got up, Boy said, ‘Mother, God filled
my heart with goodness while you prayed for us; and, mother, I will
try to-morrow.’” Is it possible that the mother could, when alone with
her children, occasionally hold this communing out loud, so that the
children might grow up in the sense of the presence of God? It would
probably be difficult for many mothers to break down the barrier of
spiritual reserve in the presence of even their own children. But
could it be done, would it not lead to glad and natural living in the
recognised presence of God?
A mother who remembered a little penny scent-bottle as an early
joy of her own, took three such small bottles home to her three little
girls. They got them next morning at the family breakfast and
enjoyed them all through the meal. Before it ended the mother was
called away, and little M. was sitting rather solitary with her scent-
bottle and the remains of her breakfast. And out of the pure well of
the little girl’s heart came this, intended for nobody’s ear, “Dear
mother, you are too good!” Think of the joy of the mother who should
overhear her little child murmuring over the first primrose of the year,
“Dear God, you are too good!” Children are so imitative, that if they
hear their parents speak out continually their joys and fears, their
thanks and wishes, they too will have many things to say.
Another point in this connection: the little German child hears and
speaks many times a day of der liebe Gott; to be sure he addresses
Him as “Du,” but du is part of his everyday speech; the circle of the
very dear and intimate is hedged in by the magic du. So with the little
French child, whose thought and word are ever of le bon Dieu; he
also says Tu, but that is how he speaks to those most endeared to
him. But the little English child is thrust out in the cold by an archaic
mode of address, reverent in the ears of us older people, but
forbidding, we may be sure, to the child. Then, for the Lord’s Prayer,
what a boon would be a truly reverent translation of it into the
English of to-day. To us, who have learned to spell it out, the present
form is dear, almost sacred; but we must not forget that it is after all
only a translation; and is, perhaps, the most archaic piece of English
in modern use: “which art,”[5] commonly rendered “chart,” means
nothing for a child. “Hallowed” is the speech of a strange tongue to
him—not much more to us; “trespasses” is a semi-legal term, never

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Biosurfactants for a Sustainable

Future: Production and Applications in

Production and Applications in the Environment

Majeti Narasimha Vara Prasad

Limit of Liability/Disclaimer of Warranty

Library of Congress Cataloging‐in‐Publication Data

Set in 9.5/12.5pt STIXTwoText by SPi Global, Pondicherry, India

List of Contributors xii

2 Metagenomics Approach for Selection of Biosurfactant Producing Bacteria

3 Biosurfactant Production Using Bioreactors from Industrial Byproducts 59

3.3 ­ actors Affect Biosurfactant Production in Bioreactor

4 Biosurfactants for Heavy Metal Remediation and Bioeconomics 79

5 Application of Biosurfactants for Microbial Enhanced Oil Recovery (MEOR) 99

6 Biosurfactant Enhanced Sustainable Remediation of Petroleum Contaminated Soil 119

7 Microbial Surfactants are Next-Generation Biomolecules for Sustainable Remediation

8 Biosurfactants for Enhanced Bioavailability of Micronutrients in Soil:

9 Biosurfactants: Production and Role in Synthesis of Nanoparticles for

10 Green Surfactants: Production, Properties, and Application in Advanced

11 Antiviral, Antimicrobial, and Antibiofilm Properties of Biosurfactants: Sustainable Use

12 Biosurfactant-Based Antibiofilm Nano Materials 269

13 Biosurfactants from Bacteria and Fungi: Perspectives on Advanced

14 Biosurfactant-Inspired Control of Methicillin-Resistant

14.5 ­ iosurfactants Illustrating Antiadhesive Properties against MRSA Biofilms 320

15 Exploiting the Significance of Biosurfactant for the Treatment of Multidrug-Resistant

16 Biosurfactants Against Drug-Resistant Human and Plant Pathogens:

17 Surfactant- and Biosurfactant-Based Therapeutics: Structure, Properties, and Recent

18 The Potential Use of Biosurfactants in Cosmetics and Dermatological Products:

18.2 ­ roperties of Biosurfactants 399

19 Cosmeceutical Applications of Biosurfactants: Challenges and Prospects 423

21 Biosurfactants as Biocontrol Agents Against Mycotoxigenic Fungi 465

21.14 ­ pportunities and Perspectives

22 Biosurfactant-Mediated Biocontrol of Pathogenic Microbes of Crop Plants 491

Shashi Bhushan Agrawal Kenia Barrantes

Liang Cheng S. J. Geetha

Sanket J. Joshi Fernanda Lugo

Aida Jover Chandana Malakar

Arun Karnwal Ana María Marqués

Asma Khatoon Francisco Meijide

Kannan Pakshirajan Aruliah Rajasekar

Rolando Procupez-Schtirbu Rashmi Rekha Saikia

Vikas Pruthi Hemen Sarma

Leonie Asfora Sarubbo SHE Empire Sdn., Jalan Pulai Ria

Julio A. Seijas Victor H. Soto

●● Addresses the applications of biosurfactants in sustainable remediation technology, for example,

19) Cosmeceutical Applications of Biosurfactants: Challenges and Perspectives

1.1 ­Introduction and Historical Perspective

Scheme 1.1 Alkaline hydrolysis of a triglyceride to obtain soaps.

Classic Bolaamphiphile Gemini Phospholipid Divalent Y-shaped Hybrid

Figure 1.1 Schematic representation of the structure of some surfactants.

O Figure 1.2 Bifacial structure of cholic acid.

1.2 ­Micelle Formation

nC3 H 7CO2 H  C3 H 7CO2 H n

Figure 1.3 Typical surface tension vs ln

Table 1.1 Parameters A and B of the Corrin–Harkins equation.

Potassium laurate 0.570 2.62

Source: Corrin and Harkins [64], p. 683.

its equilibrium constant being

From Eq. (1.16) it also follows that

In a monodisperse system, this equation may be simplified to

where Tc = ( ∆H/ ∆S)P is known as the compensation temperature.

1.3 ­Average Aggregation Numbers

1.4 ­Packing Properties of Amphiphiles

(from the volume) and

3.3 actors Affect Biosurfactant Production in Bioreactor

14.5 iosurfactants Illustrating Antiadhesive Properties against MRSA Biofilms 320

18.2 roperties of Biosurfactants 399

21.14 pportunities and Perspectives

1.1 Introduction and Historical Perspective

1.2 Micelle Formation

1.3 Average Aggregation Numbers

1.4 Packing Properties of Amphiphiles