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Evidence Based Validation of Herbal Medicine 1st
Edition Pulok K. Mukherjee Digital Instant Download
Author(s): Pulok K. Mukherjee
ISBN(s): 9780128008744, 0128008741
Edition: 1
File Details: PDF, 47.46 MB
Year: 2015
Language: english
EVIDENCE-BASED
VALIDATION OF
HERBAL MEDICINE
Edited by

PULOK K. MUKHERJEE
School of Natural Product Studies, Department of Pharmaceutical Technology,
Jadavpur University, Kolkata, India

AMSTERDAM BOSTON HEIDELBERG LONDON NEW YORK OXFORD

PARIS SAN DIEGO SAN FRANCISCO SINGAPORE SYDNEY TOKYO
Elsevier
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 Contributors

Aijaz Ahmad Department of Pharmaceutical Sciences, Fabio Firenzuoli Center for Integrative Medicine,
Tshwane University of Technology, Pretoria, South Africa Careggi University Hospital of Florence, Florence, Italy;
Shiv Bahadur School of Natural Product Studies, Referring Center for Phytotherapy, Tuscany Region,
Department of Pharmaceutical Technology, Jadavpur School of Medicine, University of Florence, Florence,
University, Kolkata, India Italy
Sanjay K. Banerjee Drug Discovery Research Center, Luigi Gori Referring Center for Phytotherapy, Tuscany
Translational Health Science and Technology Institute Region, School of Medicine, University of Florence,
(THSTI), Faridabad, Haryana, India Florence, Italy
Subhadip Banerjee Bengal Institute of Pharmaceutical Debayan Goswami ICMR Virus Unit, I.D. & B.G. Hospital,
Sciences, Kalyani, West Bengal, India Beliaghata, Kolkata, India; School of Natural Product
Studies, Department of Pharmaceutical Technology,
Santanu Bhadra School of Natural Product Studies,
Jadavpur University, Kolkata, India
Department of Pharmaceutical Technology, Jadavpur
University, Kolkata, India Ranjit K. Harwansh School of Natural Product Studies,
Department of Pharmaceutical Technology, Jadavpur
Sauvik Bhattacharyya School of Natural Product Studies,
University, Kolkata, India
Department of Pharmaceutical Technology, Jadavpur
University, Kolkata, India Michael Heinrich Centre for Pharmacognosy and
Phytotherapy/Research Cluster Biodiversity and
Rajarshi Biswas School of Natural Product Studies,
Medicines, UCL School of Pharmacy, London, UK
Department of Pharmaceutical Technology, Jadavpur
University, Kolkata, India Christian Huck Institute of Analytical Chemistry and
Radiochemistry, CCB e Center for Chemistry and
Anthony Booker Centre for Pharmacognosy and
Biomedicine, Leopold-Franzens University, Innsbruck,
Phytotherapy/Research Cluster Biodiversity and
Austria
Medicines, UCL School of Pharmacy, London, UK
Deborah Johnston Department of Economics, School of
Rainer W. Bussmann William L. Brown Center, Missouri
Oriental and African Studies, University of London,
Botanical Garden, St. Louis, MO, USA
London, UK
Joydeb Chanda School of Natural Product Studies,
Amit Kar School of Natural Product Studies, Department of
Department of Pharmaceutical Technology, Jadavpur
Pharmaceutical Technology, Jadavpur University, Kolkata,
University, Kolkata, India
India
Debprasad Chattopadhyay ICMR Virus Unit, I.D. & B.G.
Sanmoy Karmakar Department of Pharmaceutical
Hospital, Beliaghata, Kolkata, India
Technology, School of Natural Product Studies, Jadavpur
Sushil K. Chaudhary School of Natural Product Studies, University, Kolkata, West Bengal, India
Department of Pharmaceutical Technology, Jadavpur
Deepak Kasote Department of Pharmaceutical
University, Kolkata, India
Sciences, Tshwane University of Technology, Pretoria,
Søren Brøgger Christensen Department of Drug Design and South Africa
Pharmacology, University of Copenhagen, Copenhagen,
Mahnaz Kazemipoor Department of Science &
Denmark
Technology Studies, University of Malaya, Kuala
Geoffrey A. Cordell Natural Products Inc., Evanston, IL, Lumpur, Malaysia
USA
Werner Knöss Federal Institute for Drugs and Medical
Manoj K. Dalai School of Natural Product Studies, Devices, Bonn, Germany
Department of Pharmaceutical Technology, Jadavpur
Gail B. Mahady Department of Pharmacy Practice, College
University, Kolkata, India
of Pharmacy, PAHO/WHO Collaborating Centre for
Pratip K. Debnath Gananath Sen Institute of Ayurvdiya and Traditional Medicine, University of Illinois at Chicago,
Research, Kolkata, India Chicago, IL USA
Parikshit Debnath S.D.M College of Ayurveda and Subir K. Maulik Department of Pharmacology, All India
Hospital, Hassan, India Institute of Medical Sciences, New Delhi, India

xi
xii CONTRIBUTORS

José L. Medina-Franco Facultad de Quı́mica, Departamento Bhushan Patwardhan Interdisciplinary School of Health
de Farmacia, Universidad Nacional Autónoma de México, Sciences, Savitribai Phule Pune University, Maharashtra,
Mexico City, Mexico India
Achintya Mitra National Research Institute of Ayurvedic Mukhlesur Rahman Medicine Research Group, School of
Drug Development, Kolkata, India Health, Sports and Bioscience, University of East London,
Supriya Mondal ICMR Virus Unit, I.D. & B.G. Hospital, Stratford Campus, London, UK
Beliaghata, Kolkata, India Roy Upton R.H. American Herbal Pharmacopoeia, Scotts
Kakali Mukherjee School of Natural Product Studies, Valley, CA, USA
Department of Pharmaceutical Technology, Jadavpur Indra Neil Sarkar University of Vermont, Burlington, VT,
University, Kolkata, India USA
Pulok K. Mukherjee School of Natural Product Studies, Ratul Sarkar Department of Pharmaceutical Technology,
Department of Pharmaceutical Technology, Jadavpur School of Natural Product Studies, Jadavpur University,
University, Kolkata, India Kolkata, West Bengal, India
Lutfun Nahar Medicinal Chemistry and Natural Products Satyajit D. Sarker Medicinal Chemistry and Natural
Research Group, School of Pharmacy and Biomolecular Products Research Group, School of Pharmacy and
Sciences, Faculty of Science, Liverpool John Moores Biomolecular Sciences, Faculty of Science, Liverpool John
University, Liverpool, United Kingdom Moores University, Liverpool, United Kingdom
Neelesh K. Nema School of Natural Product Studies, Tuhinadri Sen Department of Pharmaceutical Technology,
Department of Pharmaceutical Technology, Jadavpur School of Natural Product Studies, Jadavpur University,
University, Kolkata, India Kolkata, West Bengal, India
Durbadal Ojha ICMR Virus Unit, I.D. & B.G. Hospital, Alvaro Viljoen Department of Pharmaceutical Sciences,
Beliaghata, Kolkata, India Tshwane University of Technology, Pretoria, South Africa
 Foreword

EVIDENCE-BASED MEDICINE, A NEED FOR PARADIGM SHIFT!

We are living in a rapidly changing world, with new economic realities, and new challenges for the production
of food and for health care. In the past 60 years a number of novel medicines have been introduced which are used
to treat various diseases, though only a few, mainly the antibiotics, do cure patients. Now for the most important
ailments medicines are available. But to develop better ones, or to develop medicines for minor diseases or diseases
of the poor, the costs are far too high to bring a novel drug to the market for the patients concerned. Estimations are
around 1 billion V for a single novel drug, moreover drug development is a time-consuming process, with
8e12 years from finding a hit to the clinical application. Pharmaceutical industry has no economic incentives in
developing novel drugs for small markets and as a result the pipeline of novel small molecule drugs is stalling, with
every year less real novel small molecule drugs. At the same time globally still up to 80% of the people are using
traditional, mostly herbal, medicines. Some 40,000e70,000 plant species have one or more medical applications in
various systems of traditional medicine. In general they are cheap and locally readily available.
Already since many years a number of governments and international organizations have advocated the use of
such local traditional medicine in primary health care. But to achieve this, one should have at least evidence for
nontoxicity and preferably even know the active compound(s) and their mode of action. With other words there is
an urgent need for evidence-based traditional medicines to be able to meet the needs of the poor. At the same time,
it may, open the way for novel ideas to develop small molecule drug by finding new leads, targets, or even con-
cepts, like the use of synergy between compounds. Moreover, such information is needed for an adequate level of
quality control. Natural products research has the important challenge to deliver the information needed for evi-
dence-based use of traditional medicines.
There are, however, a number of hurdles to overcome. First of all in the present system in the Western world
organizations like EMA and FDA make the rules for registration of novel medicines. These rules are based on a
single target, single compound paradigm. In traditional medicine often mixtures of plants are used, in which each
ingredient does have a certain meaning, and the ingredients and amounts given are part of a personalized
medication. Personalized medicine in the Western world is only used in case of very toxic medicines, e.g., in cancer
therapy, but otherwise more or less all people get exactly the same dose of a medicine. The formulations are made
with very high precision containing the active compound in a certain amount with not more than 1% standard
deviation. Obviously such a system is not at all ready to deal with the problem of every person receiving a different
mixture of plant extracts, using raw materials in which the variability in the content of active ingredient(s) with no
doubt is more than such a 1% standard deviation.
New approaches are thus required to deal with these problems. Particularly in Asia much of the traditional
knowledge has already been recorded in books thousands of years ago and still plays a very important role in
health care. Therefore this is the place for developing novel approaches supported by their rapidly growing
economies: a unique momentum for local pharmaceutical companies.
This book is an effort to bring together the views, expertise and experience of leaders in the field of medicinal plant
research and development with the aim to show what is expected and thus needs to be done to come to evidence-
based medicinal plants. Eventually this should lead to approaches that generate all the necessary information to
register medicinal plants. Evidence that also convinces the regulatory authorities that now advocate the single target,
single compound paradigm. There is a major task for us in the natural product research field. A task which requires the
close collaboration between pharmacology, toxicology, natural products chemistry, and bioinformatics. New ap-
proaches as systems biology to study single patients treated with personalized traditional medicines, instead of large-
scale clinical trials, using the various omics technologies need to be developed and used in conjunction with
reductionist approaches to confirm activity of compounds identified with activity. It requires more in depth studies
and not the more of the same as we see too much happening now, like in vitro screening of many plants at single dose
for one single activity. We need first of all to show pharmacological activity and safety of the traditional medicines,
based on that one may see if there are possibilities to develop novel leads from these plants.

xiii
xiv FOREWORD

To encourage the study of traditional medicine there is also an urgent need for protecting the rights of the
development of an evidence-based traditional medicine, just as for any novel single compound medicine. The
present patent laws will not accept the development of an example, traditional antidiabetic medicine as an inno-
vation, consequently when that evidence has been obtained, anyone may use that information and produce and
market the medicine. With other words there is a need for an economic incentive for developing traditional
medicine.
This book should thus make a major contribution to the global discussion how to explore and exploit the ancient
knowledge to the benefit of mankind. To again make discoveries like morphine, atropine, and artemisinin, evi-
dence-based medicine needs a paradigm shift to get access to the heritage of our ancestors!

Prof. Dr. Rob Verpoorte

Natural Products Laboratory, IBL, Leiden University
The Netherlands
E-mail: [email protected].
 Preface

EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE

In today’s world, the role of herbal medicine has increased manifold. The use of herbal medicine in therapeutics is
becoming more popular. In the era of revolution in herbal medicines, the need of the day is the development of an
evidence base for validation for production, evaluation, regulation, safety, and allied aspects of natural products.
Herbal medicines are evaluated, validated, and regulated in various countries according to their own system.
Globalization in the context of modern drug development will increase the practice and use of natural products
worldwide, and herbal medicine is still open to fascinating realms of research. Development of secondary metabolites
and natural leads by high-throughput screening offers exciting frontiers of future research. This book highlights
several aspects of natural products for validating the quality, safety, and efficacy of herbal medicine, particularly
methods to assess their activity and underlying mechanisms of action with a view to improve standards used in
different systems of medicine. It will provide a current cutting-edge scientific research on natural remedies, and
therefore, reading of this edited volume will be essential for everyone whose professional life impinges on the use of
natural resources.
Development of natural products requires the confluence of modern techniques and integrated approaches in
various fields of science and technology. This book provides state-of-the-art reviews from researchers around the
world on various aspects for evaluation of herbal medicine and will help researchers to know about their validation to
exploit traditional medicines (TMs) for drug discovery and development. It will be a very useful publication, which
will not only serve as a handy tool for students and researchers in this area but will also provide the most recent
methodologies developed for evidence-based validation, phytochemical and pharmacological evaluation of herbal
drugs in all aspects from field to bed side. With the emerging interest, this book will encourage the continuing efforts to
understand TM-inspired drug development as well as the roles of TM in the global health care at large. The main aim
of this book is to improve the level of understanding of various aspects on evaluation of natural products to provide a
comprehensive validation of herbal medicine, so that they can be used with greater confidence, because of improved
quality and raising a scientifically sound evidence base. It will also be an imperative essential reference for those
involved in the fields of herbal medicine, traditional remedies, pharmaceutical sciences, and natural product research.
I am sure it will be meant for a global readership and to provide a structured approach to the evidence-based eval-
uation of herbal medicinal products.

Pulok K. Mukherjee, PhD, FRSC

Editor,
School of Natural Product Studies,
Department of Pharmaceutical Technology,
Jadavpur University, Kolkata, India

xv
 Acknowledgments

EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE

The enormous growth of herbal medicinal products worldwide has been one of the most interesting aspects of
healthcare. Harmonization on the different facets of development of herbal medicine, including their quality, safety,
efficacy, validation, and regulation, is best possible through international coordination.
The intention of this book is to describe and assess various approaches for evidence-based validation of herbal
medicine, which has been described in different chapters of this edited volume by eminent scientists and tech-
nologists from different countries. I would like to express my gratitude to all of them for their valuable
contributions.
I would like to express my heartfelt thanks to Prof. Robert Verpoorte from Leiden, Netherlands, for his support
and encouragement for my works and particularly for writing the Foreword for this book.
It would not be possible to complete this work without the active help from my research group.
I gratefully acknowledge the help and support rendered by my research scholars particularly Mr Sushil K.
Chaudhary, Mr Ranjit K. Harwansh, and Dr. Neelesh K. Nema for their active cooperation and assistance. I also
thank Dr. Santanu Bhadra, Mr Amit Kar, Mr Rajarshi Biswas, Mr Shiv Bahadur, Mr Joydeb Chanda, Sk Milan
Ahmmed and Mr Sankarshan Saha for their active help.
I extremely appreciate the interest taken by Elsevier, USA, for producing this edited volume. I gratefully
acknowledge the cooperation and support received from Jill Cetel, Katey Birtcher, Beth Campbell, and Sharmila
Vadivelan from Elsevier in bringing out this book.
My father, Shri Harihar Mukherjee inspired me a lot in all of my works. Unfortunately, he passed away a few
months before this book was written. A man I intensely admire, he continues to live in my heart.
I am thankful to my family, my mother, my wife Dr. (Mrs) Kakali Mukherjee, daughter Maria and son Manish,
for all the support and love they give to me always, you are the reason behind my every successful step.

Pulok K. Mukherjee PhD, FRSC

Editor,
School of Natural Product Studies,
Department of Pharmaceutical Technology,
Jadavpur University, Kolkata, India

xvii
 C H A P T E R

1
Quality Related Safety Issue-Evidence-Based
Validation of Herbal Medicine Farm to Pharma
Pulok K. Mukherjee, Shiv Bahadur, Sushil K. Chaudhary,
Amit Kar, Kakali Mukherjee
School of Natural Product Studies, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, India

O U T L I N E

1.1 Health Care through Herbal Medicine 2 1.9.2 Boswellia serrata (Family: Burseraceae) 14
1.9.3 Gardenia jasminoids (Family:
1.2 Integrated Approaches for Development of
Rubiaceae) 16
Herbal Medicine 2
1.9.4 Matricaria recutita (Family: Asteraceae) 16
1.2.1 Opportunities and Challenges in Herbal
1.9.5 Echinacea purpurea (Family:
Medicine 2
Asteraceae) 17
1.2.2 Several Aspects for Revitalization
1.9.6 Ginkgo biloba (Family: Ginkgoaceae) 17
of Medicinal Plants 3
1.9.7 Evodia rutaecarpa (Family: Rutaceae) 17
1.3 Use of Herbs in TM 4 1.9.8 Hydrastis Canadensis (Family:
Ranunculaceae) 17
1.4 Globalization of TM 4
1.9.9 Piper methysticum (Family: Piperaceae) 18
1.4.1 Strategies for Globalization of TM 5
1.9.10 Radix pueraria (Family: Fabaceae) 19
1.5 TM Inspired Drug Discovery and Drug 1.9.11 Phyllanthus amarus (Family:
Development 6 Euphorbiaceae) 20
1.9.12 Valeriana officinalis (Family:
1.6 Issues for Quality Control and Quality
Valerianaceae) 20
Assurance of Herbal Medicine 7
1.9.13 Triphala 20
1.6.1 Contamination 8
1.9.14 Trikatu 20
1.6.2 Adulteration 8
1.9.15 Murraya koenigii (Family: Rutaceae) 21
1.6.3 Misidentification 8
1.9.16 Glycyrrhiza glabra (Family: Fabaceae) 21
1.6.4 Nonuniform Chemical Constituents 9
1.6.5 Pharmacopoeial Standards for Evaluation 1.10 Herb-Drug Interactions 21
of Herbal Products 9 1.10.1 Synergistic Effects of Herbs 21
1.7 Marker Analysis and Standardization 1.11 System Biology and Metabolomics 22
of Botanicals 9
1.12 International Harmonization 24
1.7.1 Applications of Marker Profiling 11
1.13 Conclusion 24
1.8 Pharmacovigilance of Herbal Medicine 11
1.8.1 Why Pharmacovigilance for Herbal Drugs? 12 Acknowledgments 25
1.8.2 Steps to Initiate Herbal Pharmacovigilance 12
References 25
1.9 Safety Issues on Herbal Medicine-Cytochrome
List of abbreviations 28
P450 Study 13
1.9.1 Cimicifuga racemosa (Family:
Ranunculaceae) 14

Evidence-Based Validation of Herbal Medicine

http://dx.doi.org/10.1016/B978-0-12-800874-4.00001-5 1 Copyright © 2015 Elsevier Inc. All rights reserved.
2 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

1.1 HEALTH CARE THROUGH from the leaves was 45 times more than that of the pure
HERBAL MEDICINE drug [6]. Thus, the complexity of the plant extract could
have contributed to the increased bioavailability and
Herbal medicines attract the interest of both patients thus the bioactivity. A genuine interest on various tradi-
and scientists, in all aspects of drug development from tional practices now exists among practitioners of
natural products and also for validation of traditional modern medicine and a number of practitioners of tradi-
medicine (TM). Several developing countries rely on tional, indigenous, or alternative systems are beginning
TM because of their accessibility and affordability, and to accept and use some of the modern technologies.
scientists all over the world consider medicinal plants Proper methodologies for the research and development,
as a source of new chemical entities and use them to manufacturing, and quality control of the formulations in
isolate compounds such as digoxin, morphine, taxol, TM and investigations of the therapeutic potentials of
atropine, and vinblastine [1]. Herbal medicines have an plants used in those systems with support of scientific
important position in health care systems worldwide; methods may help to use them with maximum possible
their current assessment and quality control are a major efficacy [7].
bottleneck. Many adverse events of herbal medicines
can be attributed to the poor quality of the raw materials
or the finished products. Quality issues of herbal medi- 1.2 INTEGRATED APPROACHES FOR
cines can be classified into two categories, external and DEVELOPMENT OF HERBAL MEDICINE
internal. External issues include toxic metals, pesticides
residues, microbes, adulteration, and misidentification The international trade in herbal medicine has
of medicinal plants. The internal issues affecting the qual- attracted most of the pharmaceutical companies,
ity of herbal medicines are complexity and nonunifor- including the multinationals. Until a few years ago,
mity of the ingredients. Through the use of modern only small companies had interest in the marketing of
analytical methods and pharmaceutical techniques, pre- herbal medicines. Currently, several large multinational
viously unsolved internal issues have become solvable companies are interested in commercializing herbal
[2]. The increasing search for therapeutic agents derived drugs [8]. The world market for herbal medicine,
from plant species is justified by the emergence of dis- including herbal products and raw materials, has been
eases. Medicinal plants serve as the most valuable source estimated to have an annual growth rate upto 15%.
for curing many diseases. Herbal medicines include herb- Several integrated approaches in herbal research for pro-
al extracts, herbal drug preparations, and herbal drugs. motion and development of natural products are shown
Herbal drugs are unprocessed parts of plants or whole in Figure 1.1.
plants [3]. Herbs include crude plant material such as
leaves, flowers, fruit, seed, stems, wood, bark, roots, rhi-
1.2.1 Opportunities and Challenges in Herbal
zomes, or other plant parts, which may be entire, frag-
mented, or powdered. Herbal preparations include
Medicine
comminuted or powdered materials or extracts, tinctures, With the global increase in the demand for medicinal
and fatty oils of herbal materials, which may be produced plant or plant-derived medicines, there is a call for
by extraction, fractionation, purification, concentration, ensuring the quality and safety of herbal drugs using
or other physical or biological processes [4]. several modern analytical techniques. Chemical constit-
Modern allopathic medicine has developed from uents in herbal medicine may vary depending on har-
ancient medicine, and it is likely that many important vest seasons, plant origins, drying processes, and other
new remedies were discovered and commercialized related factors. Thus, it seems to be necessary to deter-
following the leads provided by traditional knowledge mine most of the phytochemical constituents of herbal
and experiences. The study of these traditions not only products in order to ensure the reliability and repeat-
provides an insight into how the field has developed but ability of pharmacological and clinical research, to un-
it is also a fascinating example of our ability to develop a derstand their bioactivities and possible side effects so
diversity of cultural practices [5]. The administering of a as to enhance the quality of the herbal products [9].
pure chemical or a plant extract containing the same Quality control of herbal medicines aims to ensure their
chemical entity is essentially different. The difference is quality, safety, and efficacy. The lack of chemical markers
mainly due to the complexity of a plant extract that intro- remains a major problem for the quality control of herbal
duces many variables to conventional phytomedicinal medicines. In many cases, we do not have sufficient
research, which could possibly contribute to chemical chemical and pharmacological data of chemical
complexity and bioactivity. On administration of plant markers. Further, there are many technical challenges
material of Artemisia annua versus the pure drug, for in the production of markers. For example, temperature,
example, artemisinin, showed that the bioavailability light, and solvents often cause degradation and/or
 1.2 INTEGRATED APPROACHES FOR DEVELOPMENT OF HERBAL MEDICINE 3

Omic
Technologies
Target
Compound System
Analysis Biology
Approach
Extensive Quality Control
Studies of
Metabolic Metabolic Metabolomics
Metabolic Traditional
Analysis Fingerprinting Study Safety
Profiling Medicine Regulatory Aspects
Evaluation

Efficacy Study
Metabolic
Synergism
Evidence
Based
Approach
Conservations of Traditional Globalising Local Knowledge or
Medicine Localizing Global Technologies

Ethnopharmacology & Ethnopharmacological International Global trade &

Bio-cultural diversity Perspectives Studies Cooperation Commercialization

Phyto-Chemical Pharmacological Clinical Trial

Study Study Study

Herbal
New Chemical Phyto Drug Personalized Reverse Pharmacology
Pharmacovigilance
Entities Delivery Medicine

FIGURE 1.1 Integrated approaches in botanical research.

transformation of purified components; isomers and con- the TM for the betterment health care globally [13].
formations may also cause changes in the markers. How- Development and evaluation of medicinal plant-
ever, a concept of understanding the complex principles derived products are being controlled and implemented
of herbal medicine must be developed through marker through various agencies in different countries. This
profiling and related approaches so as to develop provides unique advantages for researchers and phar-
evidence-based practice of herbal medicine [10]. maceutical industries to enhance drug discovery and
Evidence-based submissions for regulatory approval development [14].
and interlinking of various pharmacopoeial and mono-
graphs would be helpful for herbal manufacturers to
1.2.2 Several Aspects for Revitalization
the regulated markets across the world. A general com-
of Medicinal Plants
parison of the pharmacopoeial standards reveals that
there is a wide variation in plant-specific parameters In order to revitalize herbal medicine in line with
and quality standards of different nations. With respect modern medicine, various strategic areas in medicinal
to Southeast Asia, India is among the leading countries plant research are being considered. Scientists are
with respect to development of pharmacopoeial stan- convinced that the integration of herbal medicine with
dards as well as modification of existing regulatory modern tools will not only benefit their own develop-
guidelines [11]. ment but will also help to fight against many complex
The major challenges for the development and diseases through development of new entities [15]. It is
promotion of TM include the chemoprofiling, safety a fact that a very large number of medicines are derived
evaluations, quality control, and effective regulatory from plants or plant-derived synthetic analogs. Dedi-
guidelines for herbal medicines [12]. Wisdom and cated research would be beneficial only with support
compassion-enhanced global collaboration and leader- from advanced approaches and novel strategies [16].
ship are needed to change the contemporary paradigms Numerous methods exist to evaluate the quality of
and develop new strategies for the enhancement of either natural or synthetic substances. Several in vitro,
TMs and dietary supplements. Research through collab- in vivo, and high-throughput screening methods are
oration and cooperation across the nation can help to a currently involved in traditional drug discovery
large extent in the promotion and development of approaches [17]. During the past decades, public interest
4 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

in natural therapies, namely, herbal medicine, has development from medicinal plants continues, with
increased dramatically not only in developing countries drug manufacturing companies engaged in large-scale
but also mainly in industrialized countries [18]. This has pharmacological screening of herbs. In TM, some popu-
increased the international trade in herbal medicine lar herbs such as Turmeric, Neem, Ginger, Holi Basil,
enormously and has attracted most of the pharmaceu- Ashwagandha, and Rauwolfia., create a revival of inter-
tical companies, including the multinationals. India is est in herbal products at a global level [22]. Around 60%
one of the few countries that are capable of producing of the global health care product market is dominated by
most of the important plants used in modern as well medicinally useful formulations and other health prod-
as in traditional systems of medicine. In the modern ucts, derived or developed from botanicals. In India,
era, combinatorial chemistry and high-throughput around 25,000 traditional and folk medicinal effective
screening are very useful methods, and so many new plant-originated formulations are used. In India, more
drug molecules are emerging from herbal resources. than 1.5 million consultants are using traditional medic-
The traditional use of medicinal plants needs to be sys- inal systems for health care, and more than 7800
tematically investigated and standardized from the manufacturing units are involved in the production of
perspective of quality, safety, and efficacy [14]. Although natural health products (NHP) and traditional plant-
there has been an increase in interest in science-based originated formulations [11]. There is worldwide
research into herbal medicine, much of the research to emerging interest in executing traditional practices in
date has been plagued by studies conducted using the health care system by exploring their therapeutic
unauthenticated, uncharacterized products. One of the as well as preventive potential. In TM, various regula-
most important issues involved in any research study tions and control on the use of botanicals have come
is the quality of the test material. A study cannot be up, which will not only help to cure different aliments
considered scientifically valid if the material tested through indigenous natural resources but will also
was not authenticated and characterized such that the help in the screening and evaluation of the medicinal
material can be reproduced. In the case of botanicals, plants in a better way to use them in traditional health
there may be misidentification of the collected plant, care systems [23].
adulteration with other species, or contamination with
extraneous ingredients [5].
1.4 GLOBALIZATION OF TM
1.3 USE OF HERBS IN TM TM has been defined as skills and a practice based on
the theories, beliefs, and experiences that are indige-
TM generally refers to those medical and health care nous to different cultures. It is used in the maintenance
systems that are practiced in a traditional manner from of health care as well as in the prevention, diagnosis,
ancient times, and this discipline is not considered to and treatment of physical and mental illnesses [24]. Sci-
be a part of conventional modern medicine. Over several entists around the world are highly emphasizing on
years, this system has evolved on the basis of religious medicinal plants as alternative medicine and their com-
beliefs and social edifices of several indigenous peoples mercial potential in health care. Globalization of TM is
by exploiting the natural resources and more recently by necessary for the establishment of evidence-based
developing a scientific method for validating therapeu- health care, based on TM in consideration of its safety,
tic and preventive approaches [19]. However, TM is efficacy, therapeutic, and clinical evidence [25]. Modern
not always documented properly through evidenced- technology and science have developed many tech-
based scientific validation as in conventional modern niques and systems for core disciplines including eth-
medicine. TMs are more easily accepted by most people nomedicine, ethnobotany, ethnopharmacology, and
due to their strong beliefs faith, practical benefits, medical anthropology to promote TM compounds
economical advantage, easy access, and many other rea- globally [10]. Establishment of global and/or regional
sons that have regional, religious, and social bases, etc. regulatory harmonization is obligatory for its develop-
[20]. In almost all TM systems, botanicals as well as me- ment and promotion through scientific validation. The
dicinal plants play a key role and constitute the back- development of TM and natural products requires the
bone of TM. The Indian material medica includes convergence of modern techniques and integrated ap-
approximately 2000 drugs of natural resources, nearly proaches related to their evidence-based research in
all of which are derived from different traditional sys- various fields of science through coordination and
tems of medicine and Indian folklore practices. Many cooperation [26].
conventional modern drugs originate from different nat- To combat the growing market demand, there is an
ural sources especially medicinal plants: a century ago, urgency to expeditiously utilize and scientifically vali-
most of the effective drugs were plant based [21]. Drug date more medicinally useful plants globally, which
 1.4 GLOBALIZATION OF TM 5
needs globalizing local knowledge and localizing global of TM-related information. These strategies based on in-
technologies, through international collaboration and formation, botany, chemistry, and biology of medicinal
cooperation. The major limits for the globalization of plant validation and quality control are essential [30].
TMs are due to having different standards of TM prod- In the era of modern research, some new drug molecules
ucts and practices, including varied terminology and are emerging with the help of combinatorial chemistry
philosophical approaches. Development of effective and high-throughput screening from herbal resources.
guidelines for safety, efficacy, and quality is regarded A study cannot be considered scientifically valid if the
as a fundamental requirement in order to establish the material tested was not authenticated and characterized
evidence base for TM [27]. The International Union of such that the material can be reproduced. In the case of
Pure and Applied Chemistry (International of Pure botanicals, there may be misidentification of the collected
and Applied Chemistry (IUPAC)) has published a series plant, adulteration with other species, or contamination
of protocols on quality control, safety, efficacy, standard- with extraneous ingredients. From the perspective of a
ization, and documentation of herbal medicine in which regulatory action, these cases may range from simple
various significant aspects and features of phytochem- misleading labeling to frank poisoning due to toxic con-
istry and analytical chemistry have been described. If taminants. It can often be difficult to compare reported ef-
these strategies are fully implemented by the IUPAC, ficacy or toxicity studies even when “standardized”
the World Health Organization (WHO) will explore material has been used. Many studies refer to the use of
TM from its pessimistic view to modern medicine [28]. standardized botanical material, which usually implies
a chemical standardization [31]. Interdisciplinary
approach of work on TM is to be explored for the discov-
1.4.1 Strategies for Globalization of TM ery of novel bioactive compounds. Issues related to the
The term “globalization” means the increased appropriateness of conventional biomedical and clinical
mobility of individuals, information, goods, services, la- models for evaluating the efficacy of TM are sometimes
bor, technology, and capital throughout the world. There very crucial. A holistic approach based on systems
are huge databases of TM, which are used by ancient biology seems much more suited to study the therapeutic
people as folk medicine, and this evidence was found efficacy and pharmacodynamics of TM-based drug
in many written textbooks [29]. There are several strate- development [10]. Approaches for drug development
gies for the expansion of TM such as (1) addition in the based on traditional leads are described in Figure 1.2.
health care system, (2) promotion of secure and valuable Most of the Indian and Chinese herbal formulations
use, (3) increasing its access, (4) increasing communica- contain a mixture of herbs, and several methods are
tion, and (5) cooperation in generation and distribution available to classify them. When two or more herbs

FIGURE 1.2 Drug development based on traditional claims.

6 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

with their bioactive compounds are combined to pre-

pare formulations, they can be observed to have the
following effects [32]:

• Synergistic significance • Mutual repression

• Synergistic improvement • Communal aggression

• Cocounteraction • Communal inappropriateness

The main ingredient is a component that provides the

main therapeutic action; the second ingredient potenti-
ates the therapeutic actions of the other, which is known
as synergistic reaction, and the component is synergism.
The rest serve one of the following functions:
• Treat accompanying symptoms,
• Moderate the ruggedness or toxicity of the primary
ones,
• Target the medicine to the proper organs, FIGURE 1.3 Thrust areas for botanical research.
• Exert a complementary effect.
In some cases, standardized bioactive compounds are
subjected to animal models to correlate the presence of noted that the plant Commiphora mukul Hook was useful
certain phytoconstituents with a pathophysiological in the treatment of obesity and related diseases. In recent
condition of the human body [33]. In the context of mod- years, a confluence of spectacular advances in chemistry,
ern biomedical research, there should also be necessary molecular biology, genomics, and chemical technology
prerequisites for clinical trials. and the cognate fields of spectroscopy, chromatography,
and crystallography may influence several therapeuti-
cally potent leading compounds from TM [11].
1.5 TM INSPIRED DRUG DISCOVERY There are many approaches for the search of new bio-
AND DRUG DEVELOPMENT logically active principles from botanicals. One can sim-
ply look for new chemical constituents and find a
In order to revitalize herbal medicine in line with biologist who will test the substance pharmacologically.
modern medicine, various strategic areas in medicinal This is not considered to be a very valid approach. A
plant research are to be considered of global importance second approach is simply to collect every readily avail-
[34]. Integration of herbal medicine and modern tools able plant, prepare extracts, and test each extract for one
would not only benefit their own development but or more types of pharmacological activity. This testing
will also help to fight against many complex diseases will help in the standardization of extracts and the
through the development of new entities. Such dedi- bioassay-guided isolation of the active constituents. The
cated research would be beneficial only with support phytoconstituents obtained can then be taken further for
from advanced approaches and novel strategies [35]. structureeactivity relationship studies [7]. Once all these
There are various thrust areas that play a very signifi- factors are determined, the constituent/extract obtained
cant role for research and development of natural prod- can be further examined for its toxicity and safety evalu-
ucts as represented in Figure 1.3. ation, followed by clinical trials. This random collection
TM is helpful in all aspects of drug development from and extensive screening method is a reasonable and the
natural resources. A few examples of drugs from natural most effective approach that eventually should produce
products would better explain the history of its own tradi- useful drugs, which can be well produced and formu-
tion. Several approaches on drug discovery and develop- lated in industries. The classic method of pharmacologic
ment from TM had been practiced by scientists for many screening involves sequential testing of herbal extracts or
years. Several therapeutically potential constituents were phytoconstituents from biological materials in isolated
isolated from plants such as artemisinin (antimalaria), organs followed by testing in whole animals, mostly in
vincristine, vinblastine, camptothecin podophyllotoxin, rats and mice. Most of the drugs in use today as therapeu-
etoposide, teniposide, and paclitaxel (anticancer) [36]. tic agents have been found and evaluated with these
The development of drugs from ayurvedic plants is methods [37]. However, for the evaluation of TM, we
ongoing, with pharmaceutical companies engaged in should not follow the reductionist approach, but go
large-scale pharmacologic screening of herbs. “Sushrutae back to the holistic in vivo approach. This can be done
Samhita,” a Sanskrit text on Ayurveda written in 600 BC in two different ways: one is through clinical trials; the
 1.6 ISSUES FOR QUALITY CONTROL AND QUALITY ASSURANCE OF HERBAL MEDICINE 7
other is through animal experiments. Besides the classic pharmaceutical quality. They have an important role
physiologic observations that can be made by in vivo ex- for the reproducibility of the effect of the active ingredi-
periments, for example, blood pressure, analgesic activ- ents from batch-to-batch uniformity. To maintain and
ity, and sedation, nowadays, it is also possible to comply with standard conditions with respect to quality,
measure gene expression, the proteome, and the metabo- safety, and efficacy of herbal medicine, it is required to
lome. These methods open up a completely new world of follow some important steps for the standardization
possibilities with several new technologies now giving a [41]. This includes the (1) proper authentication and
much better insight into the possible changes in the or- taxonomic assignment, such as through DNA finger-
ganism, in a holistic way. It will give us the possibility printing and DNA bar coding; (2) structural elucidation
to better understand the mode of action by comparing of all isolated compounds of the medicinal plant; (3)
the changes in the transcriptome, proteome, and metabo- identification and characterization of the bioactive con-
lomic patterns when compared with those observed with stituents for the pharmacological activity; (4) standardi-
known drugs. Such an approach is now known as the zation of the single extracts through spectroscopic
systems biology approach. The metabonomic approach analyses in the multicompound extracts; (5) interna-
requires the statistical analysis of large data sets by tional harmonization of specific standardization process
methods such as multivariate and principle component under the umbrella of the International Federation of
analysis to extract the information from these data [38]. Pharmaceutical Manufacturers Associations. Therefore,
Moreover, by using the systems biology approach for it is very clear that major requisites for standardization
the organism combined with metabolomic data for the of herbal products comply with international standards.
different extracts of the medicinal plant or fractions There are several variables that can influence the stan-
thereof, it should be feasible to make correlations be- dardization process. Therefore, it is compulsory to opti-
tween the occurrence of certain compounds in the extract mize all aspects of cultivation, harvesting, sample
and the activity. preparation, and sample processing to ensure reproduc-
Evidence-based medicine research should be conduct- ibility and eventually standardization of the herbal
ed with the involvement of patients and funding bodies dugs. There are various new hyphenated technologies
to establish a role of medical practitioners in decision present such as chromatographic and spectroscopic an-
making [39]. A widespread revolution in phytochemistry alyses, which need to be effectively incorporated to
has been observed through strengthening its importance ensure that sufficient quality control measures are
with the application of new technologies to enhance implemented. By using several chromatographic and
the original link between phytochemistry and TM. spectroscopic techniques, it is possible to analyze the
Evidence-based research includes developing policies, full herbal product and thus generate a standardized
regulatory criteria, and technical guidelines that would “metabolic fingerprint” of specific herbal dugs. Meta-
ensure and provide the continued availability of quality, bolic profiling can then be incorporated to identify all
safety, and effective traditional medicinal products, which the constituents [42]. The chemical fingerprints obtained
could support inclusion in health care systems, insurance from chromatographic or spectroscopic techniques
programs, and on essential medicine lists [28]. Evidence- should be similar in different samples. Spectroscopic
based submissions for regulatory authorization and inter- and chromatographic techniques are now being used
linking of various pharmacopoeia and monographs together, which leads to effective chemometric ap-
would make it easy for herbal manufacturers and they proaches. When these approaches are used in combina-
will gain greater access to regulated markets across the tion with chemometrics profiles, more precise data can
world [40]. It is under these circumstances that some of be obtained that will be helpful in the establishment of
the rationalists, scientists, scholars, and protagonists of the integrity of the herbal product and similarities and
alternative medicines dedicated themselves to the devel- differences of the observed data will be produced [30].
opment of these alternative systems for drug develop- Generally, it is believed that the risk associated with
ment from natural resources, which required to be herbal drugs is very less, but reports on serious reactions
harmonized through international coordination. indicate the need for the development of effective
marker systems for isolation and identification of the in-
dividual components [43]. Standardization of herbal
1.6 ISSUES FOR QUALITY CONTROL medicine includes the authentication of genuine drugs,
AND QUALITY ASSURANCE OF harvesting of the best quality raw material, assessment
HERBAL MEDICINE of intermediate and finished product, and detection of
harmful and toxic ingredients [44]. Several markers
Quality control and quality assurance of herbal med- such as taxonomic, chemical, genomic, proteomic
icines are very important to protect the integrity of the markers aid in the identification of herbal drug compo-
herbal extracts/products for the management of nents. Chemical markers help in the identification of
8 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

adulterants, confirmation of collection site, and quality pesticides, and others [47]. From the cultivation to the
evaluation and diagnosis of herbal intoxication. As per final herbal product development of herbal products,
the WHO definition, there are three kinds of herbal med- there are so many significant factors that can influence
icines that are obtained from raw plant material, pro- the quality of herbal products. Some significant issues
cessed plant material, and medicinal herbal products related to the quality control of herbal medicine are
[38]. Herbal medicine products are dietary supplements being described briefly in the subsequent section.
that people take to improve their health and are mar-
keted as tablets, capsules, powders, extracts, and fresh
or dried plants. Herbals are traditionally considered 1.6.1 Contamination
harmless and increasingly being consumed by people, There are so many contaminants mostly found in me-
without any prescription. The evidence for the therapeu- dicinal herbs including pesticides, heavy metals, mi-
tic actions of herbal drugs is documented in Indian, Chi- crobes, and mycotoxins. Contaminations also present
nese, European, and African systems of medicine [23]. serious obstacles for the trade of herbal medicines [48].
There are several important aspects for quality control Heavy metals have been found in herbal medicines
of herbal medicine that are shown in Figure 1.4. with some regularity. Three most commonly detected
The WHO has recognized the importance of the qual- toxic metals are mercury, arsenic, and lead. These
ity control of herbal medicine and developed a series of contaminations may occur due to (1) the accumulation
guidelines to assist several nations to develop their stra- of heavy metals in the environment (e.g., from contami-
tegies for the quality control of herbal medicines and for nated soil or atmosphere); (2) unintentional pollution dur-
conducting research on TMs [45]. The WHO had pub- ing the production process; (3) deliberate addition. In
lished the “Quality Control Methods for Medicinal Plant some of the herbal products, residues of pesticides
Materials,” a collection of recommended test procedures including their metabolites and depredated products
for assessing the identity, purity, and content of medici- remained in plants, and such residues have become a
nal plant materials to assist national laboratories notable source of contamination for herbal medicines [49].
engaged in drug quality control [46]. The WHO pub-
lished the “Guidelines on good agricultural and collec-
tion practices (GACP) for medicinal plants” and in 1.6.2 Adulteration
2007, a new guideline “WHO guidelines for assessing
quality of herbal medicines with reference to contami- Adulteration in herbal medicine increases the impu-
nants and residues” were formulated. The European rity by adding some extraneous, improper, or inferior in-
Union, China, and Japan have developed regional and gredients. Herbal medicines are adulterated with
national guidelines for good agricultural and collection conventional drugs, and plant materials have repeatedly
practices for medicinal plants that ensure that soil and been documented. Adulterations can be done in the
irrigation water used for herbal material cultivation following way including addition of orthodox drugs,
and propagation are within the limits or are free from substitution of fake or inferior plant materials, and addi-
harmful heavy metals, pesticides, herbicides, and toxico- tion of foreign materials [35].
logically hazardous substances. The certification for this
is based on parameters such as identification, water con-
tent, and chemical assay of active ingredients, inorganic
1.6.3 Misidentification
impurities (toxic metals), microbial limits, mycotoxins, Conflicting to adulteration or substitutions, misidenti-
fication of herbal medicine mostly happens unintention-
ally. False identification can occur when an importer or
retailer mistakes one herb for another, due to incorrect la-
beling and similar appearance of the herbal materials.
Confusing nomenclature can be one of the reasons,
because one herb may be known by many names: one
or more common names, a Latin name, local names,
and the brand name. Some different medicinal herbs of
different plant species with different constituents may
have similar names. The problem becomes even more
complex through confusing terminologies and the use
of different languages in different countries [50]. The
common names of herbs usually do not reflect differences
in scientific taxonomy; and the description and micro-
FIGURE 1.4 Important steps for quality control of herbs. scopic identification of n herb cannot identify its
 1.7 MARKER ANALYSIS AND STANDARDIZATION OF BOTANICALS 9
constituents. Thus, a study of ancient documents and the Therefore, more attention should be given for the biolog-
use of modern analysis techniques are often necessary to ical activity relevant to the reputation and claims for
properly authenticate herbal materials. treating particular diseases associated with herbal med-
icines [35].
The Indian Pharmacopoeia 2007 includes pharmaco-
1.6.4 Nonuniform Chemical Constituents poeial specifications with monographs for some medic-
inal plants being most commonly used as therapeutic
The chemical composition of herbal products varies
agents. The specifications include the name of the drug
and depends on the growing conditions and geographic
(along with its common name), its biological source
region. Several environmental factors that include atmo-
(Latin name), the part of the plant under consideration,
spheric humidity, rainfall pattern, soil, altitude, seasonal
its description, macroscopic and microscopic study,
variation, temperature, length of day light, may affect
identification, several quality control parameters,
the concentration of chemical constituents in medicinal
and assays with respect to the phytochemical reference
plants. Some other relevant factors, such as genetic
standards or botanical reference standards [54]. The Ay-
make-up, seeding time, use of pesticides and fertilizers,
urvedic Pharmacopoeia of India is another official com-
planting density, also play a significant role. Various
pendium published by the Ministry of Health and
processing steps of raw materials can also change the
Family Welfare, Government of India. This describes
pharmacological activity of the plant extract. Therefore,
different methods for quality control and standardiza-
batch-to-batch standardization is very essential to main-
tion of medicinal plants and herbal preparations. Several
tain the uniformity of active constituents [51].
specifications for quality evaluation of natural products
as prescribed in the Ayurvedic Pharmacopoeia include
morphological study, determination of quantitative
1.6.5 Pharmacopoeial Standards for Evaluation
data (e.g., extractive values and foreign matter), limit
of Herbal Products tests, and different physical tests (e.g., boiling range,
Safety and efficacy assessment for any pharmaceu- refractive index, and pH) [55].
tical must be taken into account for the quality of the
prepared formulation. Minimum standards for accept-
able quality are generally laid down in pharmacopoeial 1.7 MARKER ANALYSIS AND
monographs, which provide all the details of the accept- STANDARDIZATION OF BOTANICALS
able substance and give the niceties of significant tests to
determine its identity and purity. One type of pharmaco- Chemoprofiling of NHP helps in identifying the ma-
poeial monograph is found in the British or European jor metabolites and is useful to assess biological effects.
pharmacopoeias, which give only details of the tests to The development of marker-based medicines requires
be used to establish quality, with very concise notes a comprehensive understanding of plant systems
about its therapeutic application. Another type of mono- including biological, chemical, genetic, and agronomic
graph is more concerned with the complete information aspects. Chemical consistency at all stages of
about a medicinal plant and consists of all the informa- manufacturing processes is most important to ensure
tion about its chemical constituents, pharmacology, toxi- medicinal efficacy and consumer safety. This includes
cology, clinical studies and usage [52]. all the stages such as extraction, stability, shelf life, and
Pharmacopoeial monographs for the medicinal herbs purity of herbal medicines. Different methods for char-
deal with all types of pharmaceuticals and plant mate- acterization of herbal drugs such as morphological iden-
rials which have been included since the earliest editions tification, anatomical identification, and chemical
with authorization at a national or international level. It analysis, such as thin layer chromatography (TLC),
is interesting to trace the evolution of a monograph for high-performance thin layer chromatography (HPTLC),
one particular medicinal plant because it reflects devel- high-performance liquid chromatography (HPLC),
opments in analytical techniques, the increasing knowl- capillary electrophoresis (CE), Liquid chromatographye
edge of the chemical compounds present, and the mass spectrometry (LCeMS), and protein analysis are
growing body of knowledge that links the compounds extensively used [27].
present to the desired biological or clinical effect [53]. According to the European Medicines Agency
More recent editions of the British Pharmacopoeia and (EMEA), markers may be defined as chemical constitu-
European Pharmacopoeia have included monographs for ents or groups of constituents of a herbal medicinal
many more herbal drugs and more sophisticated chro- product that are very important for quality control pur-
matographic methods, especially liquid chromatog- poses regardless of whether they possess any therapeu-
raphy (LC), have been introduced for both identity tic effect. Chemical markers are basically categorized
tests, impurities tests, and for assay procedures. into the analytical markers and active markers.
10 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

Analytical markers are the constituents or groups of listed in Table 1.1. Marker selection may be based
constituents that solely serve for analytical purposes, upon a variety of different factors including stability,
whereas active markers are the constituents or groups ease of analysis, time and cost of analysis, relevance to
of constituents that contribute to therapeutic activities therapeutic effect, indicator of product quality, or stabil-
[56]. Secondary metabolites as markers have been ity or previous use by other manufacturers or re-
widely used in quality control and standardization of searchers [59].
herbal medicines. Herbal products derived from botan- A list of several therapeutically potent phytomarkers
icals are mostly obtained from wild sources and have from plant species is shown in Table 1.1. Development of
the greatest challenge for ensuring consistent product lead compounds from these medicinal plants and their
quality. These are used for making medicines where evaluation may help to promote natural products based
the standardization and quality control with proper on their quality, efficacy, and safety. Marker analysis of
integration of scientific techniques and traditional several herbal drugs including polyherbal formulations
knowledge are vital requirements [57]. Marker com- from the Indian system of medicine has been performed.
pound selection is generally based upon a variety of The fingerprint profiles of Emodin from Aloe vera, Gallic
different factors including stability, ease of analysis, acid from Terminalia chebula, Boswellic acids from Bos-
time and cost of analysis, relevance to therapeutic ef- wellia serrata, Capsaicin from Capsicum annum, Glycyr-
fect, and indicator of product quality or stability. Chem- rhizin from Glycyrrhiza glabra, epicatechin from
ical markers are frequently used for assuring quality Camellia sinensis, Eugenol from Eugenia caryophyllata,
consistency of natural products derived from botanical Ferulic acid from Coffea Arabica, Garlicin from Allium sat-
sources [58]. An ideal chemical marker for a natural ivum, Genistein from Glycine max, Ellagic acid from
product should not only be a characteristic constituent
but also a therapeutic constituent. Marker compounds TABLE 1.1 Some Medicinally Important Plants and Their
are not necessarily pharmacologically active all the Known Phytomarkers
time, but their presence is well established in products
with characteristic chemical features. Marker compo- Marker
Scientific name Family Parts used compound
nents may be classified as active principles, active
markers, and analytical makers, while biomarkers Aloe vera Liliaceae Leaves Emodin
may be defined as pharmacologically active [59]. Herbal Terminalia chebula Combretaceae Fruit Gallic acid
manufacturers and researchers need to address these
Boswellia serrata Burseraceae Resin Boswellic acids
critical questions to aid in the harmonization of specifi-
cations and analytical methodologies for natural prod- Capsicum annum Solanaceae Fruits Capsaicin
ucts. Usually, determination of single or several Glycyrrhiza glabra Leguminaceae Root Glycyrrhizin
marker compounds by a developed method is required
Camellia sinensis Theaceae Leaves Epicatechin
for quality control purpose [60]. Standardization
methods through chemical fingerprinting should take Eugenia caryophyllata Myrtaceae Flower bud Eugenol
into account all the aspects that contribute to the quality Coffea arabica Rubiaceae Seed Ferulic acid
of the herbal medicine, including correct identification
Allium sativum Amaryllidaceae Bulb Garlicin
of sample, pharmacognostic evaluation, organoleptic
evaluation, volatile matter evaluation, quantitative Glycine max Fabaceae Seed Genistein
evaluation (ash values, extractive value, foreign mat- Punica granatum Punicaceae Fruit Ellagic acid
ter), phytochemical evaluation, xenobiotic testing,
Piper betel Piperaceae Leaves Piperine
toxicity testing, microbial load testing, and biological
activity determination [39]. Medicinal plants contain Tagetes erecta Asteraceae Leaves Syringic acid
several phytoconstituents in certain ratios and in stan- Paullinia cupana Sapindaceae Seed Anthocyanidin
dardized extracts. The ratio of these chemical constitu-
Matricaria recutita Asteraceae Flowering Apigenin
ents must be constant within narrow limits from one
head
batch to another [61]. Chemical fingerprints obtained
by chromatographic, spectroscopic, thermogravimetric Citrus sinensis Rutaceae Fruit Ascorbic acid
analyses; CE; and polarography techniques have Berberis aristata Berberidaceae Berries Berberine
become the most important tools for the quality control
Curcuma longa Zingiberacease Rhizome Curcumin
and standardization of herbal medicines [12].
For ensuring consistent quality, the use of markers, Zingiber officinale Zingiberacease Rhizome Gingerol
standardization, chemical and DNA fingerprinting, bio- Citrus lemon Rutaceae Fruit Naringenin
assays, and the emerging field of phytomics are very
Vitis vinifera Vitaceae Fruit Resveratrol
important [62]. Some medicinally important plants are
 1.8 PHARMACOVIGILANCE OF HERBAL MEDICINE 11
Punica granatum, and Piperine from Piper betel, Syringic
acid from Tagetes erecta, Anthocyanidin from Paullinia
cupana, Apigenin from Matricaria recutita, Ascorbic acid
from Citrus sinensis, Berberine from Berberis aristata, Cur-
cumin from Curcuma longa, Gingerol from Z. officinale,
Naringenin from Citrus lemon, Resveratrol from Vitis
vinifera, and their pharmacological activities have been
reported. Marker analysis of Glycyrrhizin from G. glabra
has been reported through HPTLC densitometry. This is
a validated method as per the International Conference
on Harmonization guideline where the amount of gly-
cyrrhizin was determined in the extract of G. glabra
through HPTLC. The method was validated in terms
of specificity, linearity, precision, detection limit, and
quantification limit [63]. FIGURE 1.5 Application of marker analysis.

1.7.1 Applications of Marker Profiling emerging field of phytomics can provide mechanisms
for ensuring consistent quality. Marker profiling plays
Identification, authentication, and quality evaluation
a vital role in several ways to evaluate quality control
of medicinal plants are fundamental requirements of in-
parameters so as to ensure the efficacy and safety of
dustries and other organizations dealing with herbal
the herbal products Figure 1.5 [23]. There are numerous
health products. The fact must be taken into account
challenges in the isolation and identification of marker
that the plant material to be examined has a complex
components of medicinal plants. Herbal manufacturers
and inconsistent composition based on its content of sec-
and researchers need to address these critical questions
ondary breakdown products or metabolites [18]. It is an
to aid in the harmonization of specifications and analyt-
accepted fact that the qualitative and quantitative anal-
ical methodologies for the development of natural
ysis of major bioactive marker components of plant ma-
products [65].
terial is an important and reliable part of a quality
control protocol because any change in the quality of
the plant material directly affects the constituents. Me-
dicinal plant materials that qualified within the require- 1.8 PHARMACOVIGILANCE
ments of the WHO guidelines and other regulatory OF HERBAL MEDICINE
affairs can be used to develop reference fingerprints of
phytoconstituents. The presence of marker compounds Pharmacovigilance is the process of monitoring, evalu-
may be detected by the densitometric scanning of the ating, and communicating drug safety with profound im-
chromatograms [64]. The presence of these marker com- plications that depend on the integrity and collective
pounds in plant materials may be useful for quantifying responsibility of all parties such as consumers, health pro-
the plant materials in formulations or herbal medicinal fessionals, researchers, academic, media, pharmaceutical
products and will be helpful for the quality control of industry, drug regulators, governments, and international
single and polyherbal formulations. The marker organizations. The main objective of pharmacovigilance is
profiling system is helpful as a tool in the quality control to extend the safety monitoring and detect any adverse
and standardization of the raw plant materials and drug reactions that have previously been unrecognized
finished herbal formulations. Marker analysis of phyto- in the evaluation of clinical trials. There is an ongoing
constituents may also be helpful in phytoequivalence problem with unexpected toxicity of herbal products
studies, including issues such as pharmacokinetics and due to quality issues, including the use of poor quality
other related parameters that can be recognized by herbal materials, incorrect or misidentified herbs, incor-
studying the absorption, distribution, and metabolism rect processing methods, and supply of adulterated or
of herbal drugs [16]. contaminated herbs or products. The Medicines and
Further, medicinal plants do not have a constant Health care Products Regulatory Agency define some sig-
chemical composition to their different parts such as nificant problems in the regulation of herbal medicines in
roots, leaves, stems, flowers, and fruits. Therefore, each the United Kingdom [66]. These include (1) Lack of
part needs individual chemoprofiling based on their knowledge about the products being used, (2) Limited
different phytoconstituents. The use of marker profiling, use of yellow card adverse drug reporting-scheme, which
standardization, DNA fingerprinting, bioassays, and represents underreporting rather than indicating an
related metabolomics studies, which is the new absence of adverse reactions, (3) Uniform manufacturing
12 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

standards mostly of unlicensed products, and (4) Herbe scarcity of local data and lack of rigorous investigations
drug interactions of herbal medicines. on herbal traditional remedies, the promotion of the use
These quality issues can be addressed to some degree of such products focused on claims of the beneficial ef-
by improved regulation requiring good manufacturing fects and ignored the possible adverse effects. Therefore,
practice (GMP) standards for manufacturing. Pharmaco- there is now a need to revise the registration procedure
vigilance is a very essential tool for developing reliable in- for herbal products. The WHO has recognized the
formation on the safety of herbal medicines. The existing importance of the use of herbal medicines and devel-
systems were developed for synthetic medicines and oped some guidelines for monitoring herbal safety
require some modification to address the specific differ- within the existing pharmacovigilance framework [68].
ences of herbal medicine. Systematic pharmacovigilance Herbal medicines are promoted in the market as natural
is essential to build up reliable information on the safety and therefore as being safe and harmless. However, there
of herbal medicines for the development of appropriate is very less regulation control in the manufacturing of such
guidelines for their safe and effective use [67]. products; consequently, quality control issues such as
misidentification of herbs, mislabeling, contamination,
standardization of dose, method of processing, product
1.8.1 Why Pharmacovigilance for Herbal
uniformity, batch-to-batch variation, and toxicity are the
Drugs? major problems in herbal drugs [70]. Manufacturing bo-
The importance and significance of the pharmacovi- tanicals to meet analytical standards for marker com-
gilance of herbal medicines are increasing day by day. pounds does not necessarily ensure product efficacy or
Presently, accessible surveillance system for herbal generic equivalence with the products that have shown
drug monitoring is not enough, and various cases of efficacy. Herbal medicines are complex mixtures of
herbal toxicity are likely to be significantly underre- more than one active ingredient. Many times, it is unclear
ported. There is no discrimination between chemically as to which or how many constituents are responsible
defined drugs and herbal drugs in the filing of proce- for pharmacologically activity. This multitude of active
dure of adverse events. The recent information on “phy- ingredients increases the possibilities of interactions be-
tovigilance” (the term used for pharmacovigilance of tween conventional medicines and herbeherb interac-
herbal drugs) raises the suspicion that there is a ten- tions. The interaction of drugs with herbal medicine is a
dency to unequal treatment of herbal medicine [68]. Bo- significant safety concern, especially for drugs with a nar-
tanicals are complex mixtures of multiple components row therapeutic index such as warfarin and digoxin [71].
or unknown active ingredients. This can change phar- In view of establishing the safety of herbs, initiating the
macokinetic characteristics through various mecha- pharmacovigilance program will assist in the under-
nisms of action. Because the process defines the standing and prevention of adverse effects or any other
product, extrapolation of scientific data across products possible drug-related problems. The strength and po-
from different manufacturers or sources is not possible. tency of herbal products are not easily quantified, and im-
Defining the herbedrug interaction lies in the proper purity and stability are habitually not easy to examine.
identification of plants, which includes Latin binominal Therefore, botanicals should be regulated as in western
and authority, identification of the plant and part(s) used countries, and the necessities include labeling, GMP,
in the preparation of herbal products, and the processes packaging, marketing, and adverse effects reporting re-
used to extract and isolate the desired active from plant quirements, etc. [67]. Researchers, manufacturers, and
resources [67]. regulatory agencies must apply precise systematic meth-
Concomitant administration of herbal medicine with odologies and clinical trials to ensure the quality, safety,
approved conventional medications can result in thera- and consistency of the herbal products, to gain public
peutic failures or in adverse effects. Several research re- faith and confidence and to bring herbal products into
ports have suggested that St John’s Wort decreases the mainstream of health care systems.
plasma levels of various other drugs [69]. There are
no strict regulatory guidelines, and there are gaps in
1.8.2 Steps to Initiate Herbal
the inefficient regulatory processes that have allowed
Pharmacovigilance
entry of unsafe products in the market. With prescrip-
tion medicines, self-medication is a long-time practice Due to an increasing awareness at several levels of
that is unsafe and yet difficult to control, due to public herbal medicine, it is compulsory to develop pharmaco-
assumptions that herbs are generally safe because of vigilance practices. Several models of pharmacovigi-
the long tradition of their usage and the concept of being lance and its associated tools have been developed in
natural. It is surprising that these are not recognized and relation to synthetic drugs, and applying these methods
if ever observed are attributed to the remedy’s beneficial to monitoring the safety of herbal medicines presents
healing effect rather than harmful effects. Because of the unique challenges in addition to those described for
 1.9 SAFETY ISSUES ON HERBAL MEDICINE-CYTOCHROME P450 STUDY 13
Several phytoconstituents have been identified as inhib-
itors or inducers of cytochrome resulting in herbe
drug interaction. Interaction of active compounds
including allicin, quercetin, silymarin compounds, etc.,
has also been reported. Conventional pharmacokinetic
literature generally deals with drugedrug interactions,
but recently, such interactions between phytoconstitu-
ents and prescription drugs have drawn attention,
because of increasing physician awareness of the wide-
spread adverse effects of undisclosed herbal use by the
patients [11].
Cytochrome P450 (CYP450) enzymes are the major
drug-metabolizing enzymes and responsible for the
metabolism of a variety of xenobiotics including thera-
peutic drugs and some important endogenous sub-
stances, and most of the herbedrug interactions occur
FIGURE 1.6 Steps to initiate herbal pharmacovigilance. due to either induction or inhibition of these enzymes.
CYP450 enzymes are necessary for the production of
cholesterol, steroids, prostacyclins, and thromboxane
conventional medicines [72]. Several steps to initiate [76]. These enzymes also play an important role in the
herbal pharmacovigilance to monitor the safety profiles detoxification of foreign chemicals and the metabolism
of herbal medicines are shown in Figure 1.6. of drugs. There are more than 50 CYP450 enzymes, but
the CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4,
and CYP3A5 enzymes metabolize about 90% of drugs.
These isozymes are predominantly found in the liver,
1.9 SAFETY ISSUES ON HERBAL and are also present in the small intestine, placenta,
MEDICINE-CYTOCHROME P450 STUDY lungs, and kidneys [77]. An abundance of several cyto-
chrome isoforms has been determined as 30%
It is a common belief that modern drugs are CYP3A4, 20% CYP2C, 13% CYP1A2, 7% CYP2E1, 4%
dangerous and produce side effects, while herbal medi- CYP2A6, 2% CYP2D6, and 1% CYP2B6, which appears
cines are natural and very safe. In fact, some herbs can to be most relevant for the metabolism of drugs [78].
also be unsafe and even cause serious adverse effects Repeated administration of drugs can induce CYP en-
leading to death, if used inappropriately. The complexity zymes by enhancing the rate of enzyme synthesis. In-
of herbal medicine preparations and the interpretation of duction of enzymes leads to an increase in the rate of
bibliographic data on safety and efficacy reflecting the metabolite production and hepatic biotransformation
experience gathered during long-term use are best and decreases in serum drug half-life, response and
addressed by involving specific expertise and experi- can also lead to alteration of the pharmacokinetic
ence. Without the knowledge of the prescriber, the con- profile of the drug. CYP enzyme inhibition can be
sumer tends to consume the herbal products along classified into reversible inhibition and irreversible
with prescription medicine, which may lead to herbe inhibition based on the enzymatic mechanism. Revers-
drug interaction [73]. The use of complementary and ible inhibition occurs as a result of direct competition
substitute medical therapies has now become a very for the binding site on a CYP enzyme between a sub-
common trend, and their use has been documented strate and an inhibitor, whereas irreversible inhibition
widely during 2004e2014. The increased usage of herbs is caused by reactive metabolites generated from
as dietary supplements and over-the-counter products CYP-catalyzed reactions. Thus, modulation of CYP450
suggested the need for the development of clinical and enzyme metabolism is the key to change systemic
scientific data for quality and safety evaluation [74]. drug concentrations.
Toxicity may be due to the interaction of the herbal With the increasing consumption of herbal extracts
material with conventional drugs. Besides there are along with prescription medicine, issues related to the
large number of clinical drugs reported to have potential safety of herbs have become a key concern. The medical
hepato, renal, cardio toxicity, etc. during epidemiolog- and scientific literature is abundant in in vitro and in vivo
ical and other prospective studies. Other agents, reports; this suggests that the concomitant oral adminis-
such as excipients present in formulations and herbal tration of natural products and prescription drugs or
medicines that are consumed and often not disclosed over-the-counter products may affect human drug meta-
should also be considered for safety evaluation [75]. bolism and significantly increase the risk of serious
14 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

(clinical) adverse reactions. When conventional drug some medicinal plants are being described in the
substances are co-administered along with dietary sub- following section.
stances or herbal components, herbedrug interaction
may occur by CYP450 isozymes in several ways [76].
• A herbal constituent can be a substrate of one or 1.9.1 Cimicifuga racemosa (Family:
several isoforms of CYP enzymes. Therefore, one Ranunculaceae)
substrate can compete for another substrate for
metabolism by the same CYP isoenzyme, resulting in A commercially available dietary supplement made
higher plasma concentrations of the drug due to from black cohosh was identified as CYP3A4 inhibitor
competitive inhibition. [90]. The polar fraction from the extract showed 44%
• A herbal constituent can also be an inducer of one inhibition at 5 mg/ml, which was as potent as the inhibi-
or several CYP isoforms, and thereby lower tion produced by ketoconazole 58% at 5 mg/ml. The IC50
plasma concentrations of the drug due to a higher values of these six compounds were in the range of
rate of metabolism. Such interactions may produce 0.10e7.78 mM [76].
subtherapeutic plasma drug concentrations.
• A compound can also be an inhibitor of CYP450
enzymes and result in reduced activity of one or 1.9.2 Boswellia serrata (Family: Burseraceae)
several isoforms of CYP450.
The main constituents of salai guggal are volatile
Several attempts have been made to evaluate the oils, polysaccharides, triterpene acids such as a, b-
inhibitory effects of medicinal plants on CYP enzymes. boswellic acids (1-2). Boswellia carteri, Boswellia frereana,
The potential for herbal remedies to induce or inhibit Boswellia sara, and Boswellia serrata (Family: Bursera-
CYP levels has been examined. By using in vitro and ceae) herbal extracts were studied for their potential
in vivo methods, several herbs have been studied for inhibitory activity on CYP1A2, CYP2C8, CYP2C9,
their potential inhibitory effects on human liver micro- CYP2C19, CYP2D6, and 3A4 [91]. The aqueous metha-
somes, rat liver microsomes, and cDNA expressed nolic (MeoH 80% v/v) extracts of Boswellia species
human liver microsomes [79]. Studies on drug- showed potential inhibitory activities. CYP1A2 and
metabolizing enzymes enhance our understanding of CYP2D6 were inhibited at a concentration of 10 mg/
the possibilities for herbedrug interactions. Several ml. CYP2C8/9/19 and 3A4 were strongly inhibited at
existing reports on the role of drug metabolism enzymes, this concentration (IC50 values between 1 and 10 mg/
mainly CYP450, in herbedrug interactions are summa- ml). In order to evaluate the contribution of boswellic
rized in Table 1.2 with reference to individual herbs. acids, selected boswellic acids were evaluated for their
Several therapeutically active plant extracts have inhibition activity. From the study, boswellic acids were
been reported to interact with drugs leading to clinically identified as moderate to potent inhibitors of the CYP
relevant adverse drug reactions. Interaction potentials of enzymes (IC50 values between 5 and 120 mM) [76].

H
H

HO HO
H H
O O

OH OH
a-Boswellic acid (1) b-Boswellic acid (2)
 1.9 SAFETY ISSUES ON HERBAL MEDICINE-CYTOCHROME P450 STUDY 15
TABLE 1.2 Effects of Herbal Extracts on CYP450 Enzymes [76]

IC50 value/
Name of the Part used Type of Study CYP isoforms Effects on percentage
plant in the study extract method used CYP450 inhibition Reference

Acorus calamus Root Hydroalcohol Fluorimetry Human Inhibition [80]

CYP3A4 and 46.84 m/ml and
CYP2D6 36.81 mg/ml
Capsicum Fruit Methanolic Fluorimetry Human Inhibition [81]
annuum CYP3A4 99.69 mg/ml
CYP2D6 68.25 mg/ml
CYP2C9 and 88.03 mg/ml and
CYP2C19 84.16 mg/ml
Centella asiatica Whole plant Aqueous HPLC Human Inhibition [82]
CYP2C9 599.0 mg/ml
CYP2D6 413.1 mg/ml
CYP3A4 229.5 mg/ml
Centella asiatica Whole plant Ethanolic HPLC Inhibition [82]
CYP2C9 28.3 mg/ml
CYP2D6 and 418.9 mg/ml and
CYP3A4 465.8 mg/ml

Curcuma longa Rhizome Aqueous HPLC Human Inhibition [83]

CYP2C9 82.3%
CYP2C19 92.7%
CYP2D6 48.6%
CYP3A4 92.8%
Cymbopogon Aerial part Methanolic Radiometry Rat Inhibition [84]
nardus CYP3A4 370 mg/ml

Emblica officinale Fruit Hydroalcohol Fluorimetry Human Inhibition [85]

CYP3A4 and 152.11 mg/ml and
CYP2D6 109.96 mg/ml
Origanum Leaves Aqueous HPLC Human Inhibition [83]
vulgare CYP2C9 35.4%
CYP2C19 80.2%
CYP2D6 94.6%
CYP3A4 98.6%
Piper longum Fruit Methanolic Fluorimetry Human Inhibition [86]
CYP3A4 and 164.81 mg/ml and
CYP2D6 210.60 mg/ml
Piper nigrum Fruit Methanolic Fluorimetry Human Inhibition [86]
CYP3A4 and 178.34 mg/ml and
CYP2D6 234.90 mg/ml

Rheum palmatum Root Methanolic Radiometry Rat Inhibition [84]

CYP3A4 and 467 mg/ml and
CYP2D6 385 mg/ml
Rhodiola rosea Root Ethanolic Spectrofluorimetry CYP2D6 and Inhibition 32 mg/ml and [87]
CYP3A4 67%
Rhododendron Leaf Ethanolic Spectrofluorimetry Human Inhibition [87]
groenlandicum CYP3A4 48%
Salvia officinalis Leaves Aqueous HPLC Human Inhibition [83]
CYP2C9 97.2%
CYP2C19 99.9%
CYP2D6 99.8%
CYP3A4 97.0%
Continued
16 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

TABLE 1.2 Effects of Herbal Extracts on CYP450 Enzymes [76]dcont’d

IC50 value/
Name of the Part used Type of Study CYP isoforms Effects on percentage
plant in the study extract method used CYP450 inhibition Reference

Sanatalum album Wood Methanolic Radiometry CYP3A4 and Inhibition 337 mg/ml and [84]
CYP2D6 886 mg/ml
Strychnos Wood Methanolic Radiometry Rat CYP2D6 Inhibition 637 mg/ml [84]
ligustriana
Strychnos Leaf Methanolic Radiometry Rat CYP2D6 Inhibition 302 mg/ml [84]
ligustrina
Syzygium Flower Methanolic Radiometry Rat Inhibition [84]
aromaticum CYP3A4 and 219 mg/ml and
CYP2D6 249 mg/ml
Tanacetum Leaves Hydroalcohol HPLC Human Inhibition [83]
parthenium CYP2C9 51.5%
CYP2C1 46.2%
CYP2D6 and 54.1% and
CYP3A4 64.7%
Terminalia Fruit Hydroalcohol Fluorimetry Human Inhibition [85]
bellerica CYP3A4 and 77.94 mg/ml and
CYP2D6 90.20 mg/ml
Terminalia Fruit Hydroalcohol Fluorimetry Human Inhibition [88]
chebula CYP3A4 and 95.52 mg/ml and
CYP2D6 102.35 mg/ml
Thonningia Root Aqueous Spectrophotometry Rat Inhibition [89]
sanguinea CYP1A1 19%
CYP2B1 18%
CYP2B2 18%
CY1A2 40%
Tinospora crispa Methanolic Radiometry CYP3A4 and Inhibition 428 mg/ml [84]
CYP2D6 488 mg/ml
Zingiber Rhizome Ethanolic Radiometry Rat Inhibition [83]
aromaticum CYP3A4 and 102 mg/ml and
CYP2D6 693 mg/ml
Zingiber officinale Fruit Methanolic Fluorimetry Human Inhibition [86]
CYP3A4 and 286.49 mg/ml and
CYP2D6 249.52 mg/ml

1.9.3 Gardenia jasminoids (Family: Rubiaceae) 1.9.4 Matricaria recutita (Family: Asteraceae)
The effects of Gardenia jasminoids extract on liver The inhibitory effect of Matricaria recutita essential oil
CYP-450-dependent monooxygenases, glutathione, and and its major constituents such as chamazulen (3) and
glutathione-S-transferase were investigated using rats a-bisabolol (4) on four selected human CYP450 enzymes
treated orally with the iridoid glycoside (0.1 g/kg body (CYP1A2, CYP2C9, CYP2D6, and CYP3A4) demon-
weight/day) or the crude extract of its fruits (2 g/kg/ strated the inhibition of these enzymes, with CYP1A2
day) for 4 days [92]. Gardenia jasminoides decreased the being more sensitive than the other isoforms. Chamazu-
CYP450 content in liver microsomes and demonstrate lene (IC50 ¼ 4.41 mM), cis-spiroether (IC50 ¼ 2.01 mM),
that geniposide from G. jasminoides has the ability to and trans-spiroether (IC50 ¼ 0.47 mM) were shown to
inhibit a CYP3A4 monooxygenase and increase gluta- be potent inhibitors of this enzyme and also active
thione content in rat liver. Further, immunochemical toward CYP3A4, CYP2C9, and CYP2D6. Chamazulene
data using immunoblotting studies using antibodies (IC50 ¼ 1.06 mM) and a-bisabolol (IC50 ¼ 2.18 mM)
revealed that geniposide treatment markedly decreased caused a significant inhibition of CYP2D6. As indicated
the protein immunorelated to CYP3A in rat liver [76]. by these in vitro data, chamomile preparations contain
 1.9 SAFETY ISSUES ON HERBAL MEDICINE-CYTOCHROME P450 STUDY 17
constituents inhibiting the activities of major human of Ginkgo biloba are potent in vitro inhibitors of human
drug-metabolizing enzymes [93]. CYP2C9. Another study was undertaken to clarify the in-
fluence of repeated oral administration of ginkgo extract
on CYP2C9 and CYP3A4. A combination of G. biloba and
tolbutamide is to be administered cautiously in terms of
O potential interactions, especially in elderly patients or
patients treated with drugs exerting relatively narrow
N therapeutic windows. Greenblatt et al. (2006) investi-
gated the effect of G. biloba on the activity of CYP2C9
when administered with warfarin in humans [96].
Chamazulene (3)
N N
1.9.7 Evodia rutaecarpa (Family: Rutaceae)
Rutaecarpine (5), a quinazolinocarboline alkaloid that
a-Bisabolol (4) is a major constituent of Evodia fruit, caused the most
dramatic decrease in residual CYP3A4 activity. It was
further identified as a mechanism-based inhibitor of
CYP3A4. Rutaecarpin also showed potent inhibition
1.9.5 Echinacea purpurea (Family: Asteraceae)
to CYP1A1 and CYP1A2 (IC50 0.90 and 0.06 mM). An
In vitro CYP3A4 inhibition profiles of Echinacea pur- analysis showed that methanol extract increased the
purea and ketoconazole were evaluated by different sub- levels of CYP1A1, CYP1A2, CYP2B, and glutathione-
strates and showed a much lower CYP3A4 inhibition S-transferase Yb immunoreactive proteins and aqueous
by E. purpurea (IC50 ¼ 5394 mg/ml) compared to that extract increased CYP1A2 protein level. Three major
by fluorescent substrates (IC50 354 and 452 mg/ml, bioactive alkaloids, that is, rutaecarpine, evodiamine
respectively). From the study, it was suggested that the (6), and dehydroevodiamine (7) at 25 mg/kg increased
substrate/assay-dependent effects may occur due to hepatic ethoxyresorufin-O-deethylase (EROD) activity.
the complex nature of E. purpurea constituents, involving These results demonstrated that Evodia rutaecarpa meth-
different CYP3A4 substrate binding sites. A weak inhibi- anol and aqueous extracts could affect drug-
tion potential of E. purpurea toward CYP3A4-mediated metabolizing enzyme activities [76].

O O O

N N
N

N
N N
N
Me
N NH

H Dehydroevodiamine (7)
Evodiamine (6)
Rutaecarpine (5)

metabolism in vitro was confirmed by the use of three 1.9.8 Hydrastis Canadensis (Family:
different substrates [94]. Ranunculaceae)
This contains the alkaloids berberine (8) and
hydrastine (9), hydrastinine (10), and canadine
1.9.6 Ginkgo biloba (Family: Ginkgoaceae) (11). Its extract contains approximately equal concen-
Ginkgo is believed to improve cerebral and peripheral trations (w17 mM) of two ethylenedioxyphenyl alka-
blood flow through nitric oxide-induced vasodilation and loids, berberine and hydrastine, inhibited with
possesses antioxidant activity [95]. Certain components increasing potency the isoform CYP2C9 (diclofenac
18 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

4b-hydroxylation), CYP2D6 (bufuralol 1b-hydroxyl- inhibitors of CYP1A2, CYP2C9, CYP2C19, CYP2D6,

ation), and CYP3A4 (testosterone 6b-hydroxylation) CYP3A4, and CYP4A9/11 and studied the effect of
activities in human hepatic microsomes with interpo- kava rhizome extracts and kava alkaloids, pipermethys-
lated IC50 values of 0.98%, 0.66%, and 0.18%, respec- tine (12) in rat liver microsomes. Pipermethystine alone
tively [76]. demonstrated a nonsignificant increase in CYP1A2,

O O

O O
N+

O
N

O
O
Hydrastine (9)
O
Berberine (8)

O
O
O
N N

O
Hydrastinine (10)
O

Canadine (11)

while kava rhizome extracts alone and in combination

1.9.9 Piper methysticum (Family: Piperaceae)
with pipermethystine increased hepatic CYP1A2 protein
Whole kava extract (normalized to 100 1 M total kava- levels by 98%. From the study, it is understood that kava
lactones) caused concentration-dependent decreases in kava may have a potential to produce drug interactions.
CYP450 activities, with significant inhibition of the ac- Zou et al. (2002) [98] investigated the influence of iso-
tivities of CYP1A2 (56% inhibition), CYP2C9 (92%), lated kavalactones kavain (13), dihydrokavain (14),
CYP2C19 (86%), CYP2D6 (73%), CYP3A4 (78%), and methysticin (15), yangonin (16), and desmethoxyyango-
CYP4A9/11 (65%) following preincubation for 15 min. nin (17) on recombinant human CYP isoforms such as
These data indicated that kava has a high potential for CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4.
causing drug interactions through inhibition of They calculated IC50 values between 0.43 and
CYP450 enzymes. Jennifer and Ramzan and Jennifer 153.20 mM from the mean of four determinations for
(2004) [97] reported that several kavalactones are potent the potent inhibitory active compounds.
 1.9 SAFETY ISSUES ON HERBAL MEDICINE-CYTOCHROME P450 STUDY 19

O
O O

O N
O O
O O

Dihydrokavain (14)

Kavain (13)

Pipermethystine (12)

O
O O

O O O

O
O O
O O
O Yanogonin (16) Desmethoxyyanogonin (17)
Methysticin (15)

1.9.10 Radix pueraria (Family: Fabaceae)

phosphate-(CYP)-c-reductase activity were significantly
This possesses a high content of flavonoids, couma- increased, a complex pattern of CYP modulation was
rins, and isoflavones such as daidzein (18) and puerarin observed, including both induction (puerarin:
(19). Crude extracts containing puerarin inhibited in a CYP2A1, CYP1A1/2, CYP3A1, CYP2C11; Ge-gen:
dose-dependent fashion. Although both CYP content CYP1A2, CYP3A1, CYP2B1) and inactivation (Ge-gen
and reduced nicotinamide adenine dinucleotide and puerarin: CYP3A, CYP2E1, CYP2B1) [99].

OH
OH
O
O OH
OH

HO O O
OH
O
Daidzein (18)
HO OH

Puerarin (19)
20 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

1.9.11 Phyllanthus amarus (Family: extracts showed higher IC50 values (<0.1 mg/ml)
Euphorbiaceae) compared to those of gallic acid (0.09 mg/ml) against
CYP2D6. On the other hand, quinidine and ketocona-
All the CYP450 enzymes were significantly inhibited zole showed relatively strong inhibition with an IC50
by the Phyllanthus amarus extract in a concentration- value of <0.007 mg/ml. The results indicated that the
dependent manner. The concentrations needed for 50% test substances have much less interaction potential
inhibition of CYP1A1, CYP1A2, and CYP2B1/2 were with coadministered drugs than do the known inhibi-
4.6 mg/ml, 7.725 mg/ml, and 4.18 mg/ml, respectively. tors. Triphala formulation has less interaction potential
Oral administration of P. amarus (250 mg/kg) was found when compared with individual plant extracts
to reduce the activity of these enzymes and an increased and bioactive molecules. Triphala formulation and its
concentration of the extract (750 mg/kg) [100]. individual ingredients may likely interact with drug-
metabolizing enzymes, but are less likely to produce
significant drug interactions [85].
1.9.12 Valeriana officinalis (Family:
Valerianaceae) OH
Its essential oil contains terpenoid-like hydroxylva-
HO
lerenic acid (20), and valerenic acid (21). CYP3A4-
mediated metabolism tended to be slightly lower in OH
aqueous extracts and slightly higher in those extracted
with acetonitrile. In most cases, the ethanolic extracts O
were slightly less active than the corresponding aqueous
or acetonitrile extracts. From the study, it was concluded OH
that valerian extracts exhibited a marked inhibition of Gallic acid (22)
CYP3A4-mediated metabolism [76].

O
H
HO
OH OH
H

O
Hydroxyvalerenic acid (20)
Valerenic acid (21)

1.9.13 Triphala
1.9.14 Trikatu
“Triphala” is a well-known polyherbal formulation
from Indian Ayurveda called “Rasayana” in Sanskrit. It Trikatu is a very well-known “Rasayana” in Ayur-
consists of dried fruits of Emblica officinalis, Terminalia beler- veda and widely used as a polyherbal ayurvedic formu-
ica, and Terminalia chebula in equal proportions (1:1:1). lation in India. Trikatu means the mixture of three acids.
Traditionally, this formulation has been prescribed for It consists of three well-known plants, namely, Piper
several ailments and used as a laxative, detoxifying longum, Piper nigrum, and Zingiber officinale, in an equal
agent, digestive agent, and rejuvenator [101]. It has ratio. Trikatu has been prescribed for cough, cold, fever,
been reported that E. officinale extract showed the high- asthma, respiratory problems, and improvement of
est IC50 value (152.11  2.18 mg/ml) and Terminalia beller- digestive disorders. It is reported that the trikatu and
ica showed the lowest IC50 value (69.89  2.50 mg/ml) its biomarkers have very less inhibitory effect on
against CYP3A4. The inhibitory activity against CYP450 enzymes. Different concentrations of the trikatu
CYP3A4 with IC50 values <0.1 mg/ml were found formulation and its individual components showed
with the three fruit extracts and the formulation, while significantly (P < 0.001) less inhibitory activity on indi-
gallic acid (22) was found to produce inhibitory activity vidual isoenzymes as compared to that shown by the
with approximately 0.09 mg/ml. Likewise, all the fruit positive control [86].
 1.10 HERB-DRUG INTERACTIONS 21
1.9.15 Murraya koenigii (Family: Rutaceae) surveillance programs. This information may be
used as a basis for a simplified registration procedure
Murraya koenigii extract has a higher IC50 value and may serve as a substitute for animal experiments
(160.47  5.45, 206.63  1.99, and 156.56  3.77 mg/ml and reduce the number of clinical trials in humans
of CYP3A4, 2D6, and 2C9 isozymes, respectively) than [105]. Prescribers and consumers of herbal products
do the standard biomarkers. M. koenigii extract and its will be able to recognize and report on major acute
bioactive compounds have an inhibitory effect on drug adverse events, such as dermatological reactions,
metabolism enzymes when consumed along with con- nausea, and disturbances of the gastrointestinal tract.
ventional medicine. The IC50 values were higher than Consequently, data on traditional use are unlikely to
those of the positive control and indicated that the test provide information on chronic toxicity and carcino-
extracts and constituents have moderate interaction in genic, mutagenic, and teratogenic effects [76]. Several
drug metabolism [81]. regulatory bodies have acknowledged this problem
and have given the right to national authorities to de-
mand such supplementary safety testing when biblio-
1.9.16 Glycyrrhiza glabra (Family: Fabaceae)
graphic evidence is deemed to be insufficient to prove
The major bioactive constituents of Glycyrrhiza glabra safety before marketing authorization of the herbal
are glycyrrhizin, glabranin A and B, glycyrrhetol, glabri- products [78].
din, formononetin, glabrone, etc. [62]. A research report Consumption of herbal products that are capable of
on the CYP450 interaction profiles of G. glabra and its modulating CYP metabolism can cause clinically rele-
bioactive compound glycyrrhizin showed that the vant herbedrug interactions and can alter drug bioavail-
extract and glycyrrhizin have a higher interaction poten- ability. Any inhibitory effect of herbal extracts on CYP
tial with CYP2D6 compared with CYP3A4. The licorice may result in enhanced plasma and tissue concentra-
extract showed a comparably higher IC50 value with tions leading to toxicity, while any inductive effect
both the isozymes, which may be related to the synergis- may cause reduced drug concentrations leading to
tic effects for the presence of other constituents in the decreased drug efficacy and treatment failure [104].
extract. The IC50 values that are higher than those of The complexity of herbal medicine preparations and
the positive inhibitors indicated that the test samples the interpretation of bibliographic data on safety and
have only a weak interaction potential in drug meta- efficacy reflecting the experience gathered during
bolism. An IC50 value of the extract that is lower than long-term use are best addressed by involving specific
that of the pure compound indicates that care should expertise and experience.
be taken when administering the extract with other
CYP450-interacting compounds, particularly those
with a low therapeutic index [102]. 1.10.1 Synergistic Effects of Herbs
The mechanisms of synergistic actions of herbal in-
gredients can be explored for designing new multitarget
1.10 HERB-DRUG INTERACTIONS drugs and drug combinations and for discovering
potent drug combinations that are individually subther-
The plant material as an active ingredient in herbal apeutic but efficacious in combination. Synergistic ac-
products may also be related to health risks, because it tions involve interactions with multiple sites, targets,
also contains some toxic constituents or constituents and pathways that are sensitively influenced by genetic,
that are known to affect the bioavailability and pharma- environmental, behavioral, and scheduling profiles
cokinetic or pharmacodynamic interaction of other com- [106]. It is claimed that combinations of herbs have
pounds or drugs [103]. Another problem associated with synergistic effects. There is much in vitro and/or in
the use of herbal products is underreporting of observed vivo evidence to support the occurrence of synergism
adverse reactions and herbedrug interactions. In a between constituents in certain herbal extracts. Synergy
study, it was found that 58% of users do not inform their is also taken to mean an attenuation of undesirable
physician when they buy any herbal medicinal prod- effects, another key theory of herbalism being that the
ucts. It has been reported that 69% of Britishers and toxicity of plant extracts is less than that of a single
61.7% of Italians that use herbal product do not consult isolated constituent.
their physicians [104]. Synergism has a major role in therapeutic efficacy of
Many medicinal herbs have a long history of use, and medicinal plants and plant-derived formulation. This
this may have generated a significant amount of pub- cannot be easily distinguished from additive effects
lished toxicological information including scientifically and usually relies on high margins of variation. In
accepted monographs, clinical experience, and epidemi- fact, the mechanism of action of many phytomedicines
ological studies, as well as data from postmarketing is still unknown, and there are several instances of a
22 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

total herb extract showing a better effect than an equiv- 1.11 SYSTEM BIOLOGY AND
alent dose of an isolated compound [107]. Generally, METABOLOMICS
synergistic effects are considered to be positive, with
the low doses used perceived as a benefit, although it System biology intends to recognize the biological
is obvious that there may also be negative aspects. complexity of different measurements without any
Pepper contains the alkaloid piperine, which is known hypothesis. The major focus of systems biology is to
to increase the bioavailability of a number of clinically enquire the dynamics of all genetic, regulatory, and
used drugs. Unwanted interactions, for example, would metabolic processes in a cell and to understand the
be the presence of tannins in herbal drugs, which may complexity of cellular networks [108]. Adoption of the
hinder the absorption of proteins and alkaloids, or the systems biology approach would be more helpful for
induction of enzymes, such as CYP450, which may exploring the scientific implication of herbal medicine
accelerate drug metabolism resulting in blood levels of and the modernization of TM.
actives too low for a therapeutic effect [101]. The issues There are several technological platforms of system
concerning the safety of herbs with increasing consump- biology, such as genomics, proteomics, and metabolo-
tion of herbal extracts along with prescription medicine mics, that provide powerful tools for the study on the
have become a major concern. This leads to several essence and function of herbal drugs (Figure 1.7). Scien-
studies on evaluation of their effects on drug- tifically and technologically validated herbal products
metabolizing enzymes. These studies will help to under- can be explored on a fast track using various innovative
stand and ensure the possibilities for herbedrug approaches such as reverse pharmacology and systems
interactions. biology, which are based on a knowledge of TM. TM

FIGURE 1.7 System biology in drug discovery.

 1.11 SYSTEM BIOLOGY AND METABOLOMICS 23
comprises of evolutionary process as communities to reported. Besides a recognizable pattern, the quantita-
discover practice transforming techniques. The methods tive analysis of ginkgolides and bilobalide could be
for carrying out metabolic modeling by means of collect- done with a 5-min acquisition time of the spectrum,
ing, storing, and analyzing metabolomic data are without the need for any elaborate sample preparation.
considerably different, which will generally be per- Also, for other preparations, it was found that this
formed by individuals or in laboratories with different method is very suitable; among other preparations
skill sets, and yet necessarily will deal with the same studied were strychnos, ephedra, and cannabis. Such
molecules [109]. It is therefore very essential to timely studies are the first steps in the long way to a better
bring together the known or conditional metabolic understanding of the activity of medicinal plants [114].
maps of suitable organisms with measurements of their Metabolomic study reveals the quantitative and
metabolomes to provide a system level understanding qualitative estimation of “whole” metabolite found in
of the metabolic fluxes and metabolite concentration in a cellular or organism system. It can be defined as the
these organisms, and their way of changing under systemic study of the individual chemical fingerprints
different conditions [110]. that a definite cellular process leaves behind and even
LCeMS is an analytical technique that combines the more particularly the technique of the metabolite profile
physical separation capabilities of LC with the mass of the “whole” small molecules in an organism. The
analysis capabilities of MS. LCeMS is a powerful tech- combined data of all the metabolites in a biological sys-
nique used for many applications and has a very high tem, which are the final products of its gene expression,
sensitivity and selectivity. LCeMS consists of an ultra- are known as the metabolome. These approaches deal
high LC combined with a mass spectrometer that con- with the study of genomics, transcriptomics, and prote-
tributes to achieve high-throughput studies in omics of biological systems. It involves four major steps
metabolomic analysis. LCeMS is frequently used in of analysis of the unknown compound present in the
drug development at many different stages including herbal medicine, which includes the following:
natural product dereplication, metabolomic stability
• Targeted investigation of the compound: This deals
screening, metabolomic identification, impurity identifi-
with quantitative estimation of definite metabolites.
cation, quantitative analysis, and quality control. Gener-
• Metabolic documentation: Quantitative and
ally, its application is oriented toward the specific
qualitative data for the estimation of the unknown
detection and potential identification of phytoconstitu-
compound or of definite metabolic pathways.
ents in a plant extract. LCeMS becomes a powerful
• Metabolomic fingerprinting: This is the process of
approach for the rapid identification of phytochemical
sample classification by rapid global investigation.
constituents in botanical extracts, and it can avoid the
• Metabolomic examination: This pertains to the
time-consuming isolation of all compounds to be identi-
quantitative and qualitative analysis of “whole”
fied. LCeMS is most commonly used for metabolomic
metabolites.
analysis of plant extract where secondary metabolite
masses may overlap even with a high-resolution mass Metabolomic analyses use a particular set of analyt-
spectrometer. Profiling of secondary metabolites in ical techniques such as Fourier transformed infrared
plants or food, such as phenolics, can be achieved with spectroscopy, gas chromatographyemass spectrometry,
LCeMS [111]. LCeMS, NMR, CEeMS, and TLC. Recent advances
Metabolomics aims at qualitatively and quantita- made in analytical chemistry for small mass compound
tively determining as many compounds as possible. detection and characterization, such as MS and high-
This can not only be in extracts of tissues but also in field NMR, coupled with modern multivariate statistics,
body fluids such as serum or urine in the case of have led to a highly efficient system for the comprehen-
humans. Chromatographic methods in combination sive analysis of the metabolite data matrices generated
with MS, MS with nuclear magnetic resonance (NMR) by metabolomic experiments [9]. In the past decades,
spectrometry, are used for such analyses [112]. By several attempts have been made to solve these prob-
combining the result of such analyses with other param- lems using metabolomics. Metabolomics is a relatively
eters, novel correlations can be found, for example, a new field of “omics” research concerned with the
relationship between the occurrence of certain com- high-throughput identification and quantification of
pounds in extracts and a biological activity. Analysis of small-molecule metabolites in the metabolome. Analysis
metabolites in urine by means of 1H-NMR is already of a large number of samples might facilitate the identi-
extensively applied for studying the toxicity of drugs fication of patterns or metabolite markers that are char-
[113]. The metabolomics approach is also a very prom- acteristic of a species, a cultivar, a certain stage of
ising tool for the quality control of botanicals. A study development or conditions, such as disease state, stress,
on the metabolic profiling of G. biloba L. in pharmaceu- or daily and seasonal changes. Therefore, the high-
tical preparation by means of 1H-NMR has been throughput global analysis of a metabolome through
24 1. QUALITY RELATED SAFETY ISSUE-EVIDENCE-BASED VALIDATION OF HERBAL MEDICINE FARM TO PHARMA

hyphenated technologies is a key factor in this field but as yet there is no consensus as to how these should
[115]. Thus, metabolomics is now emerging as a power- be adopted. There are several publications: US Pharma-
ful tool for the characterization of complex phenotypes copoeia, British Herbal Compendium, British Herbal
affected by both genetic and environmental factors. Pharmacopoeia, Chinese Pharmacopoeia, and Physi-
Nevertheless, metabolomic fingerprinting often lacks cian’s Desk Reference for herbal medicines, Ayurvedic
robustness, so targeted or profiling approaches may be Pharmacopoeia of India have monographs for herbal
useful techniques for the validation of herbal medicine raw materials [24]. For a single plant, the monograph
with the necessary specificity, precision, accuracy, line- may vary in different publications, different country
arity, sensitivity, recovery, and stability in the presence standards with respect to a single formulation, which
of potentially interfering compounds [116]. creates difficulties for manufacturers in herbal drug
trade. Thus, the need to establish global regulatory
mechanisms for regulating herbal drugs seems obvious.
1.12 INTERNATIONAL An improvement in the processes of regulation and
HARMONIZATION global harmonization is desirable and necessary, which
combines scientific data and traditional knowledge [34].
To ensure homogeny of quality, safety, and efficacy of Several challenges and issues on promotion and
the herbal medicines across countries, harmonizing ef- development of herbal drug have been identified, which
forts have been initiated on pharmacopoeial specifica- should be solved through international coordination
tions, standardization, and classification of herbal and collaboration. Considering the widespread use
drugs. Different specifications have been found in and popularity of herbal products, there is a need for
different pharmacopoeias of Korea, Japan, and China adequate evidence in the quality, safety, and efficacy of
for a single herbal medicine. The same crude plant ma- herbal products, which is mandatory. Therefore, there
terial may be described, but the family or species of is a need for coordination and harmonization of research
the original plant may be different [117]. The Western and development of medicinal plants as both pharma-
Pacific Regional Forum for the Harmonization of herbal ceuticals and food supplements.
medicine tried to harmonize the crude drug mono-
graphs in the pharmacopoeias of six Asian countries
(Japan, China, Korea, Singapore, Vietnam, and Hong 1.13 CONCLUSION
Kong) in order to help in promoting commercialization
of safe and effective herbal drugs across countries. The existing knowledge of herbal medicines needs to
Harmonization process and herbal product registration be validated and documented through regulatory
were initiated in 2000 among different countries. Inter- guidelines of quality control, standardization, and
national harmonization would help in developing manufacturing process. Chemical consistency at all
evidence-based clinical practice guidelines on TM. India stages of manufacturing processes such as extraction,
has nearly 8000 herbal drug companies, among which standardization, quality control, quality assurance, sta-
about five thousand companies have GMP-compliant bility, shelf life, and purity is of utmost important to
manufacturing units, and most of them are of small ensure medicinal efficacy and safety. Considering the
and medium size. Seventy percent of the Indian exports widespread use and popularity of herbal medicines,
from the herbal sector consist of only raw materials, and proper standardization and validation methods have
30 percent consist of finished products including herbal been developed for the promotion of natural products.
extracts. There are 55 major herbal drugs exporting com- Medicinal plants are most commonly used in TMs and
panies in India [28]. As discussed in the monographs of can potentially influence the bioavailability and phar-
the American Herbal Pharmacopoeia, the use of single macokinetics of some pharmaceuticals by affecting
or multiple chemical markers was important for quality CYP450 drug metabolism. Evidence-based submissions
control. Protocols and guidance documents on safety for regulatory approval and interlinking of various
and toxicity testing of TMs have been issued by the In- pharmacopoeial monographs would be helpful for herb-
ternational Life Sciences Institute, the Institute of Medi- al manufacturers to regulate markets across the world.
cine, National Research Council (2004), the Union of Research through collaboration and cooperation can
Pure and Applied Chemistry, the EMEA (e.g., EMEA, help to a large extent in the promotion and development
2009) [118], and by the European Food Safety Authority of TM for the betterment of health care globally. This
(EFSA, 2009) [119] These regulation documents tell us may bring scientists and other stakeholders together to
about the assessment of the safety, efficacy, and quality discuss global issues and implications of new strategic
of herbs for food and medicine purposes. Standards terms, with a new set of goals, a new agenda, but most
for medicinal plants are being developed worldwide, importantly, a new vigor is vital for global development.
 REFERENCES 25
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[117] Jordan SA, Cunningham DG, Marles RJ. Assessment of herbal LIST OF ABBREVIATIONS
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 C H A P T E R

2
Value Chains of Herbal
MedicinesdEthnopharmacological and
Analytical Challenges in a Globalizing World
Anthony Booker1, Deborah Johnston2, Michael Heinrich1
1
Centre for Pharmacognosy and Phytotherapy/Research Cluster Biodiversity and Medicines, UCL School of Pharmacy,
London, UK; 2Department of Economics, School of Oriental and African Studies, University of London, London, UK

O U T L I N E

2.1 Introduction 29 2.7 The Tea Value Chain 35

2.2 The Concept of Value Chains 30 2.8 The Ginseng Value Chain 36
2.3 The Medicinal Plant Value ChainsdResearch 2.9 Plant Metabolomics and Analytical Challenges 37
Needs 30
2.10 Discussion 39
2.4 Medicinal Plant Value Chains in Asia 31
Acknowledgments 42
2.5 Medicinal Plant Production in China and India 32
References 42
2.5.1 China 32
2.5.2 India 32 List of Abbreviations 44
2.6 Supply, Demand, and Sustainability 34

2.1 INTRODUCTION high-quality herbal medicinal products (HMPs). First,

with diverse national initiatives and with the develop-
According to the World Health Organization, in ment of quality standards, for example, in the European
2008, world trade in herbal medicine was estimated Pharmacopoeia and, in 2004, with the introduction of
at US$83 billion [1]. Much of the trade in medicinal the EU-wide, Traditional Herbal Medicinal Products
plants has been based within national market systems. Directive (THMPD), which requires well-defined stan-
While global trade in high-value products, such as dards of quality and safety to be ensured before a prod-
spices and medicines, has a long history (e.g., the spice uct can be released onto the market. Until December
route [2]), during 1984e2014, international trade has 2012, there has been >1000 traditional use registration
flourished, and a main thoroughfare of this trade exists licenses granted in Europe [3].
between Asia and Europe, the United States and The effect of these new regulations on products orig-
Australia. inating outside of the EU and particularly in Asia has
From a global perspective and particularly with been substantial, with protests being voiced from trade
regards to products sold with a specific medical claim, associations and government officials. This has been
Europe has been leading the way in terms of supplying particularly noticeable in China and India, two countries

Evidence-Based Validation of Herbal Medicine

http://dx.doi.org/10.1016/B978-0-12-800874-4.00002-7 29 Copyright © 2015 Elsevier Inc. All rights reserved.
30 2. VALUE CHAINS OF HERBAL MEDICINESdETHNOPHARMACOLOGICAL AND ANALYTICAL CHALLENGES IN A GLOBALIZING WORLD

that have had a long history of trade in HMPs with the Changes in production, distribution, and financial
UK and the rest of Europe [4,5]. systems, in synergy with the globalization of markets
The main problems highlighted and voiced by manu- and the spread of information and communication tech-
facturers are that the more sophisticated inputs involved nologies, suggest that more attention needs to be paid to
in meeting the standards and the costs attached to these both external and internal linkages within a company
registered products are seen as being too high for many structure. The concept of the value chain allows us to
companies that operate within Asia and particularly shift the focus from manufacturing only to the other ac-
within India [6]. tivities involved in the supply of goods and services,
These concerns are similar to those held generally including distribution and marketing.
on the impact of production standards [7]. However,
changes in regulatory requirements or the setting of Value chain research focuses on the nature of the relation-
higher entry bars, although presenting many challenges ships among the various participants involved in the chain,
and on their implications for development. At any point in
to the industry, can also offer opportunities. An increase the chain, some degree of governance or coordination is neces-
in quality standards is often regarded as a positive step sary in order to take decisions on how the chain should be
by consumers, particularly when related to food or managed effectively [10].
medicines, and these improvements in quality and
safety can be regarded as adding value to a product. However, value chain analysis has also been criti-
If companies are able to use these added requirements cized for not giving enough attention to wider social,
to their advantage, it may be possible for new and economic, and political factors. Its strength lies in
more sustainable enterprises to be established, particu- describing the value chain linkages, but it does not
larly if there is cooperation between Asian and Euro- explain how and why these linkages have developed.
pean actors. Therefore, any research investigating the value chain
The creation of vertically integrated value chains should also include a wider exploration of these sur-
(VIVCs) is one way to achieve this cooperation and for rounding issues.
producers in less economically developed countries
(LEDCs) to gain better access to highly regulated markets.
An introduction to value chains, the impact that these
chains have on the quality of finished HMPs, the impact 2.3 THE MEDICINAL PLANT VALUE
on livelihoods, and an outline of the potential effects that CHAINSdRESEARCH NEEDS
overharvesting may have on the industry, forms the
main aims of this chapter. Value chain analysis has been applied to a variety of
consumer goods, but only in the last 10 years has it
been applied to medicinal plant production [11e17].
By examining medicinal plant value chains (MPVCs),
it is possible to gain a better understanding of the role of
2.2 THE CONCEPT OF VALUE CHAINS different actors and inputs in the chain and their influ-
ence over chain management. Understanding the pro-
The literature on global commodity chains has moved cess is a vital step toward suggesting any meaningful
away from the term “commodity chain or supply chain” strategies for improvement. The cultivation of medicinal
in favor of the term “value chain.” The latter is thought plants is a relatively new industry, and wild collection
to better describe a wider variety of products and ser- represents the main route of supply in terms of the num-
vices, while also focusing on the distribution of benefits. ber of species collected [18,19]. On many occasions, wild
As a result, the global commodity chain approach is now medicinal plants are preferred by traditional healers and
known as global value chain analysis [8]. consumers over cultivated ones [20], as there is a general
feeling that wild plant species are more clinically
Value chain analysis describes the activities within and effective.
attached to an organisation, and relates them to an analysis of Worldwide, medicinal plant species are getting
the competitive strength of the organisation. Therefore, in eco-
nomic terms, it evaluates that value each particular activity depleted rapidly due to overcollection from their natural
adds to the organisations’ commodities or services. This idea habitats [21,22]. The collection and marketing of medic-
was founded upon the insight that any company is more than inal plants from the wild is an important source of liveli-
a random assembly of machinery, people and finance and hood for many of the poor in LEDCs. In Nepal, >300,000
only if these things are arranged into definable systems will it households are engaged in the collection of medicinal
become possible to produce something for which customers
are willing to pay a price. Therefore, it is argued that the ability plants [12]. It is claimed that up to 15,000 species of me-
to perform particular activities and to manage the linkages be- dicinal plants are globally threatened [23], and two ways
tween these activities is a source of competitive advantage [9]. are identified to conserve threatened species: first by
 2.4 MEDICINAL PLANT VALUE CHAINS IN ASIA 31
tightening restrictions on collection practices, and sec- Cultivation is seen as a way of breaking free from
ond by cultivation on a large scale [24]. some of these ties, and small-scale projects have
commenced in the area. There are difficulties and risks
attached to cultivation, and publiceprivate collabora-
tions are suggested as a way of minimizing these con-
2.4 MEDICINAL PLANT VALUE CHAINS straints. However, these collaborations have remained
IN ASIA small and further promotion, regulation, and invest-
ment are needed if these ventures are to produce a
Kala et al. [21] assert that the marketing system in meaningful result [12].
India is largely unregulated and inequitable. The medic- The work of Alam and Belt focused on a project in
inal plant cultivators are generally marginal farmers and Uttarakhand, northern India, where 80% of the popula-
laborers. They receive a cash income to meet their basic tion relies on agriculture as their main economic activity,
requirements of food, health, and the education of their and 40% of people live below the poverty line. It was
children. They are often unaware of the real market pri- proposed that the cultivation of a medicinal plant, kutki,
ces of many of the medicinal plant species. It is often Picrorhiza kurroa Royle ex Benth. Plantaginaceae, would
difficult for farmers to sell certain herbs due to their benefit the farmers financially, provide social benefits,
lack of knowledge of the marketing system, and and help preserve wild species. Moreover, the European
conversely, many medicinal plants are destined to be buyer would have a secure supply of the plant from a
traded through illegal channels. Other constraints are fully traceable source.
the slow rate of production, a long gestation period, The Uttarakhand project produced disappointing re-
shortage of cultivation technology, low yields, unscien- sults, and in their paper, the authors highlighted the rea-
tific harvesting, poor processing, lack of quality control, sons for this as being poor quality of planting material,
scarcity of good manufacturers, and poor marketing planting on small, poorly irrigated plots, and the emer-
infrastructure. gence of apples as a profitable cash crop, resulting in
In a review [15] of fieldwork conducted in Uttaranchal, farmers switching over from kutki to apples.
one of India’s poorest states, van de Kop et al. document Alam and Belt concluded that the cultivation of me-
that resource-poor people collect plants from the wild dicinal plants is more difficult than usually suggested
to supplement their low income. They point out that in the scientific literature and government promotional
high risks, transaction costs, and a lack of trust among material and stress the importance of agencies and
chain actors prevent small-holder producers from taking nongovernmental organizations (NGOs) taking these
up the cultivation of medicinal plants and suggest difficulties into account and taking steps to minimize
publiceprivate collaboration as a way of reducing these these. The authors further argue that a thorough tech-
constraints and secure market accessibility for small nical and economic feasibility study of the value chain,
producers. They analyze the opportunities for and the long-term involvement of governmental and NGOs,
constraints on developing medicinal plant chains and and an understanding of the prevalent farming system
aim to identify the role of medicinal plant chains in are necessary to ensure the success of the chain.
poverty reduction. Conversely, a project conducted in Bangladesh [14]
Most of the medicinal plants in Uttaranchal are appears to be more optimistic with respect to the eco-
collected from the wild. Permits are issued to coopera- nomic potential of medicinal plants and suggests verti-
tive groups in the area, which in turn employ contractors cal integration as a vehicle to benefit and empower
to organize the collection. The contractors employ col- producers and processors at the beginning of the value
lectors, usually land owning farmers or landless la- chain. The primary and wholesale secondary markets
borers. The contractor can sell the collected plants were dominated by middlemen, but their study chal-
either to the local cooperatives or directly to indepen- lenged the view that medicinal plant cultivation was
dent traders after paying royalties to the cooperative. only appropriate for relatively well-off people with bet-
The cooperatives sell either to local agents or whole- ter access to land, capital, and information.
salers, traders in large cities, or to drug manufacturers. The authors built on previous work by researchers
The traders supply the domestic market and interna- [15] and argue that some of the mechanisms employed
tional markets. in developing and sustaining institutional relationships
Van de Kop et al. suggest that in the value chain, the may also apply equally well to defining the MPVC and
collectors and local contractors are in a weak position as list contracts, quasivertical integration (an especially close
they cannot sell directly to the largest trading companies and long-term relationship), tapered vertical integration
in the cities, and depend on the local traders for market- (when a company sources inputs externally from inde-
ing. This weak position often results in them receiving a pendent suppliers as well as internally within the
considerably lower price than the true market price. same company), cost plus agreements (where the
32 2. VALUE CHAINS OF HERBAL MEDICINESdETHNOPHARMACOLOGICAL AND ANALYTICAL CHALLENGES IN A GLOBALIZING WORLD

contractor is paid a negotiated amount regardless of China has partly addressed its own difficulties
incurred expenses), joint ventures, and strategic alliances relating to the manufacture and supply of TCM prod-
as examples of these potential relationships. ucts by modernizing its traditional medicines profession
Moreover, they argue that the benefits of an inte- with government-sponsored good agricultural practices
grated value chain are numerous. and good manufacturing practices (GMPs). All manu-
facturers of TCM products must comply with standards
It enables primary producers to become active participants set down by the China Food and Drug Administration to
in the process, it removes market access barriers, and it results
gain GMP certification. Only around 1500 companies
in better commercialization of products and is attractive to com-
panies as they can have greater control over quality and supply.
have achieved this standard.
However, TCM products originating in China are of
In their study, Shahidullah and Haque, contrary to particular concern to many regulatory authorities such
previous views, found that the cultivation or production as in the UK and instances of poor quality and adulter-
of medicinal plants could play an important role in ated material are commonly reported [28e30].
improving the livelihoods of those poor or very poor To satisfy the requirements of an international mar-
people who may own only small pieces of land. ket, some companies have found it necessary to put in
They argue that to sustain growth in medicinal plant place systems that help address the quality concerns of
production, foreign customers. For example, Taiwanese herbal prod-
uct manufacturers, which supply herbs to distributors in
A fair distribution of the gross margin to the primary pro- many parts of the world, had to find an alternative strat-
ducers is necessary. egy for buying herbs at local or provincial herbal mar-
kets to better assure their customers that they can
In the value chain system examined in [14], it was source good quality herbal ingredients that can be traced
found that downstream buyers, especially manufac- back to the areas of cultivation. The belief is that by hav-
turers and consumers, pay most of their money for the ing direct links to the farms it is easier and less costly to
value additive opportunistic pricing of middlemen ensure the quality and traceability of the herbs in com-
due to inherent weaknesses in the chain. parison to using the herbal markets that tend to domi-
A vertically integrated chain, with only producers nate herbal medicinal trade. In China, it is difficult for
and processors as commercial actors and NGO’s as pro- non-Chinese organizations to make direct links with in-
moters, could create a better and more equitable dividual farmers, so herbal sourcing companies have
situation. emerged to provide this missing link in the export mar-
However, there are some studies that suggest that ket supply chain.
vertically integrated chains, particularly ones that are
dominated by a powerful company, can lead to negative
effects on the livelihoods of small producers.
This has been illustrated in the work of van Niekerk
2.5.2 India
and Wynberg [25], in which the authors present evi- Kala et al. [21] assert that of the 17,000 higher plant
dence to suggest that (based on their assessment) the species to be found in India, 7500 are known for medic-
monopolistic behavior of one large European company inal uses, with Ayurvedic medicine claiming to use 2000
threatened the livelihoods of farmers in South Africa of these. Most plants used in Indian systems of medicine
(for a single botanical drug) and that there were major are collected from the wild. More than 60 species are in
inequalities between the bargaining powers of the local great demand, and the “tribal belt” of India is abundant
manufacturers and the primary producers. in these plants, and minority groups mainly depend on
this trade for their livelihoods [31]. The annual turnover
of the Indian herbal medicine industry has been esti-
mated by different authors to be between US$377 mil-
2.5 MEDICINAL PLANT PRODUCTION
lion and US$1 billion per annum [21,32], between 0.5
IN CHINA AND INDIA
and 0.8 per cent of world trade.
The globalization of Ayurvedic practices gained
2.5.1 China
momentum during the 1990s and onward into the
China has a long and relatively well-recorded history 2000s, and Ayurvedic products are commonly used as
of using plants for medicinal purposes [26]. There are food supplements in North America, Australia, New
hundreds of state-owned and increasingly shared Zealand, Europe, and Japan [33]. While Indian exports,
ownership companies producing traditional Chinese valued at US$132 million in 2008, contributed less than
medicines (TCMs), many of which export their products one per cent to the global herbal market, industry
internationally [27]. observers suggest that growth is rapid and that Indian
Exploring the Variety of Random
Documents with Different Content
the eggs being fertilised in the water after their extrusion from the
body of the female, and, consequently, any device which will
facilitate the formation of shoals during the breeding season must be
of great advantage to the species by largely increasing the chances
that the ova will be fertilised, and thus secure the more successful
propagation of the race. Hence it may be concluded that the vocal
organs of Fishes are a means to this end, and that the sounds they
produce are in fact recognition-sounds which enable Fishes of the
same species to congregate together at periods when reproductive
activity is greatest. This view is in harmony with much that is known
of the habits of these Fishes, especially with the fact that particular
sounds are often characteristic of particular species, and that the
sounds are produced most frequently and with greater intensity
during the breeding season than at any other time. While useful to
all Fishes that possess them, vocal organs are, no doubt, specially
serviceable to those Fishes which, from the nature of their habitat,
can make but little use of their eyes; and this fact may perhaps
explain the prevalence of such organs in the Siluridae, which are
frequently bottom- or ground-feeding Fishes, and often live in
muddy waters.

The sounds emitted by Fishes may also, in some instances at least,

be warning sounds. Many of the sound-producing Fishes are
provided with exceptionally strong spines either in connexion with
the median and paired fins, as in many Siluridae, or on the general
surface of the body, as in Diodon hystrix. Such spines are very
effective weapons for offensive or defensive purposes, and are
capable of inflicting very severe wounds. The natural enemies of
these Fishes learn by experience or instinct to associate particular
sounds with the possession of dangerous spines, and warned by the
sounds, they refrain from attacking the owner of the spines, to the
mutual advantage of both.
 Fig. 209.—An Electric Ray (Torpedo) dissected to show its electric
organs. On the left the nerves supplying the organ are dissected
out. The prismatic areas on the surface of the organ indicate the
vertical columns of electric plates, of which there may be 500,000
in each organ. The dorsal surface of the brain is exposed. br,
Gills; f, spiracle; o, eye; o.e, electric organs; t, mucus canals; tr,
tri-geminal nerve; tr′, its electric branch; v, vagus; I, fore-brain;
II, mid-brain; III, cerebellum; IV, electric lobe of the medulla
oblongata. (From Parker and Haswell, after Gegenbaur.)

Electric Organs.—Electric organs capable of generating more or

less powerful electric discharges are present in certain Fishes, both
marine and freshwater. They occur in a few Elasmobranchs (species
of Raia, Torpedo, and Hypnos), in such Teleosts as the African Silurid
Malopterurus the "Electric Eel" (Gymnotus), and in species of
Mormyridae (e.g. Mormyrus). With one exception electric organs are
composed of metamorphosed muscular fibres, and their nerve-
endings or motor end-plates. The species of Raia have two small
electric organs, one on each side of the terminal portion of the tail.
[432] In Gymnotus[433] the organs are much larger, and extend the
whole length of the tail, which is fully four-fifths of the total length
of the Fish. The Mormyridae also have their feeble electric organs in
the caudal region. In all these Fishes the electric organs are modified
portions of the caudal muscles. In the Torpedo, however, these
organs are two large oval masses, one on each side of the head,
between the gills and the cephalic prolongation of the pectoral fin
(Fig. 209). Malopterurus[434] is exceptional in possessing an electric
organ derived from the epidermis and not from the muscular
system. In this Fish the organ envelops nearly the whole body like a
mantle, between the skin and the subjacent muscles of the trunk
and tail. An electric organ is composed of an immense number of
"electric plates" (modified motor end-plates), abundantly supplied
with nerves on one of their surfaces, and disposed in a series of
vertical (Torpedo) or longitudinal (Gymnotus) columns, separated by
septa of connective tissue. In the active state of the organ in the
Torpedo[435] the ventral surfaces of the plates, on which the nerves
are distributed, become negative to the dorsal, and "the effect in all
the plates of a column when summed up is, therefore, such that the
dorsal end of a column becomes positive to the ventral end."[436]
Hence the current in the form of a succession of shocks passes from
the ventral to the dorsal surface of the head. In Gymnotus, where
the columns are longitudinally arranged, it is the anterior and
posterior surfaces which become oppositely electrified, and the
current passes from the tail to the head. The shock imparted by an
electric discharge is most powerful in Gymnotus,[437] Malopterurus,
and Torpedo, in the order named, and relatively weak in the
remaining genera. The strength of the shock increases with the
number of electric plates included in the circuit. Thus in Gymnotus
the maximum shock is given when the body of the Fish is so curved
that the head and the tail are in contact with different points on the
surface of some other Fish. The discharge may be reflex or
voluntary. Repeated discharges induce fatigue and weaken the
shocks. Electric organs are powerful offensive or defensive
structures, enabling the Fish to repel the attacks of enemies, or to
stun or kill their prey.

CHAPTER XIV
 NERVOUS SYSTEM AND ORGANS OF SPECIAL SENSE

The nervous system consists of the brain and the spinal cord, and of
the cranial and spinal nerves. The rudiment of the future brain and
spinal cord first appears in the embryos of some Cyclostomes (e.g.
Bdellostoma), of Elasmobranchs, and of Chondrostei (e.g.
Acipenser), and of Neoceratodus among the Dipnoi, in the form of a
tubular medullary canal pinched off from the epiblast of the dorsal
surface of the body. By a somewhat different method, but with the
same final result, a medullary canal is formed in other Cyclostomes
(e.g. Petromyzon), in the Holostei and Teleostei, and in Lepidosiren,
[438] from a solid ingrowing keel of epiblast which subsequently
becomes tubular. Later, the medullary canal in the head enlarges,
and becomes divided by two transverse constrictions into three
vesicles, the primary fore-, mid-, and hind-brain, leaving the rest of
the canal to form the spinal cord.

The Spinal Cord.—This portion of the medullary canal retains a

simpler and more uniform cylindrical structure. Its walls thicken and
their component cells become converted into nerve cells and nerve
fibres, but a remnant of the original cavity remains in the adult as a
minute axial canal, with a ciliated epithelial lining, the central canal
of the spinal cord or myelocoele. In most Fishes the spinal cord
extends the whole length of the body, but in some Teleosts,
especially in certain Plectognathi, it is remarkably short. In a Sun-
Fish (Orthagoriscus), 2½ metres long, and weighing about a ton and
a half, the cord was only 15 mm. in length, or shorter than the brain.

The Brain.—At an early stage in its embryonic history the brain

consists of three simple vesicles, the fore-, the mid-, and the hind-
brain, the first of which lies in front of the anterior end of the
notochord and is therefore pre-chordal in position. As development
proceeds the walls of the vesicles undergo local thickenings, or they
give rise to hollow paired or median outgrowths, and by one or other
of these methods the different parts of the complex adult brain are
evolved, while the original cavities of the vesicles or of their
outgrowths persist as a continuous system of epithelium-lined
spaces or "ventricles."[439] The fore-brain is remarkable for the
number and importance of the parts to which it gives rise. First, it
bulges out in front into a hollow vesicle, the prosencephalon, leaving
the rest of the fore-brain as the thalamencephalon or diencephalon
(Fig. 210). The cavity of the prosencephalon is the prosocoele, and a
pair of thickenings in its floor form two basal ganglia or corpora
striata. In many Fishes the prosencephalon retains this simple
vesicular condition, in which case the roof or pallium is usually
epithelial and non-nervous; but in others two hollow lobes grow out
from it in front and give rise to two cerebral hemispheres or
parencephala.[440] Both contain extensions of the prosocoele, the
paracoeles or lateral ventricles, from the floor of which the corpora
striata now project. The prolongation of the pallium forming the roof
of the lateral ventricles either remains partially epithelial, or it may
acquire a wholly nervous structure and thicken to an extent which
differs greatly in different Fishes. With the formation of the
hemispheres the prosencephalon and its prosocoele become of
secondary importance, and may cease to be recognisable as distinct
from the thalamencephalon and its ventricle. The lateral ventricles
then appear to communicate directly with the third ventricle by two
apertures, the foramina of Munro. The forward growth of the brain is
completed by the development of two hollow lobes, the olfactory
lobes or rhinencephala, each of which contains a ventricle or
rhinocoele communicating behind with the prosocoele, or, if
hemispheres are present, with the corresponding lateral ventricle.
 Fig. 210.—Diagram of the general structure of the brain in Craniates. A,
vertical longitudinal section; B, dorsal view showing the brain
cavities on the right side. c, Cerebellum; c.c, central canal of the
spinal cord; c.h, cerebral hemispheres; c.s, corpus striatum; F.B,
fore-brain; f.m, foramen of Munro; H.B, hind-brain; in,
infundibulum; l.v, lateral ventricle; m, mesocoele; M.B, mid-brain;
m.o, medulla oblongata; o.l, olfactory lobe; op.l, optic lobe; op.t,
optic thalamus; p, paraphysis; pc, prosocoele; pn.o, pineal organ;
p.o, parietal organ; pr, prosencephalon; pt, pituitary body; rh,
rhinocoele; sp.c, spinal cord; s.v, saccus vasculosus; th,
thalamencephalon; iii, iv, third and fourth ventricles. (After Parker
and Haswell.)

Scarcely less complicated, and perhaps even more interesting from a

morphological standpoint, are the structures arising out of the
thalamencephalon. By thickenings of its lateral walls two large
ganglia, the optic thalami, are formed, and on the inner or dorsal
aspect of each of these a ganglion habenulae is developed. From the
sides of the thalamencephalon the primary optic vesicles are
derived, which later become transformed into the retinal parts of the
paired eyes and the optic nerves. Besides the optic vesicles there is
a second pair of embryonic outgrowths which arise from the roof of
the thalamencephalon. These outgrowths form stalked vesicles and
represent a pair of degenerate visual organs. Usually they become
so displaced that the left one lies in front of the right, and they
appear as if median. The subsequent fate of the vesicles differs
greatly in different Craniates. Both persist in the Lamprey, the right
vesicle to some extent retaining its primitive visual function as a
parietal eye and directly overlying the left or pineal vesicle. In
Elasmobranchs the two unite to form a glandular organ, the so-
called pineal body of the adult, and in Teleosts the left vesicle
disappears, leaving the right as a pineal body.[441] There is also an
embryonic median outgrowth from the roof of the prosencephalon,
the paraphysis, which soon disappears and whose significance is not
known. A median hollow downgrowth from the floor of the
thalamencephalon forms the infundibulum, which becomes attached
to a caecal diverticulum from the roof of the mouth. With rare
exceptions the diverticulum loses all connexion with the mouth, and,
as the pituitary body or hypophysis, it appears as an appendage to
the extremity of the infundibulum. In the Crossopterygii the
connexion is retained even in the adult by means of a slender canal
extending from the pituitary body and opening into the oral cavity.
Laterally, the base of the infundibulum grows out into a pair of
rounded lobes, the lobi inferiores, and distally into a thin-walled
glandular sac, the saccus vasculosus, which lies just behind the
pituitary body. The cavity of the thalamencephalon persists as the
third ventricle or diacoele. The parts of the brain developed from the
mid-brain and the hind-brain are much less complicated, and, except
for variations in size, they present a fairly uniform character in most
Fishes.

In the mid-brain the roof bulges out into a pair of optic lobes, and by
the growth of lateral thickenings in its floor two thick strands of
longitudinally disposed nerve fibres, the crura cerebri, are formed.
The cavity of the mid-brain remains as the mesocoele, and from it
an extension may be prolonged into each optic lobe.

From the hind-brain are formed the cerebellum or epencephalon and

the medulla oblongata or metencephalon, the former as a dorsal
bulging, the latter as a ventral thickening. Except where the
cerebellum is developed the dorsal wall remains epithelial, and forms
the roof of the persistent cavity of the hind-brain, the fourth
ventricle or metacoele, which retains its primitive continuity with the
central canal of the spinal cord. Lateral lobe-like outgrowths from
the dorsal columns of the medulla are conspicuous structures in
some Fishes, and are known as corpora restiformia. The paired
portions of the brain are connected across the middle line by a series
of transverse commissures. The more important modifications of the
brain in Cyclostomes and Fishes will now be briefly dealt with.

Fig. 211.—Dorsal (A) and ventral (B) views of the brain of Petromyzon
marinus. ch.pl.1, Anterior choroid plexus forming the roof of the
prosencephalon and thalamencephalon; ch.pl.2, aperture in the
roof of the mid-brain exposed by the removal of the middle
choroid plexus; ch.pl.3, the fourth ventricle exposed by the
removal of the posterior plexus; cr.crb, crura cerebri; crb,
cerebellum; crb.h, cerebral hemispheres; dien, thalamencephalon;
inf, infundibulum; l.gn.hb, left ganglion habenulae; med.obl,
medulla oblongata; nv.1, olfactory; nv.2, optic; nv.3, oculomotor;
nv.5, trigeminal; and nv.8, auditory nerves; olf.l, olfactory lobes;
opt.l, optic lobes; pn, pineal organ; r.gn.hb, right ganglion
habenulae. (From Parker and Haswell, after Ahlborn.)
 Fig. 212.—Dorsal view of the brain of Myxine. c.r, Corpora restiformia;
m.o, medulla oblongata; n.p, naso-pituitary canal; ol.o, olfactory
organ enclosed in its fenestrated cartilaginous capsule; op.l, optic
lobes; pr, prosencephalon; s, s, dorsal roots of spinal nerves;
sp.c, spinal cord; th, thalamencephalon. (From Wiedersheim, after
Retzius.)

In the Cyclostome Petromyzon there is a small prosencephalon with

an undivided prosocoele, and on each side of it a small cerebral
hemisphere which appears as a mere appendage to the much larger
olfactory lobe (Fig. 211). The prosocoele divides in front into two
outwardly directed branches, and of the two diverticula into which
each branch divides one extends as a lateral ventricle into the
hemisphere of its side, and the other as a rhinocoele into the
corresponding olfactory lobe. The ganglia habenulae are unusually
large, the right one being larger than the left. The optic lobes are
large, but not obviously double. So small is the cerebellum that it
seems to be little more than a narrow transverse band crossing the
fore-part of the fourth ventricle. The roof of the brain is largely
epithelial, especially in the prosencephalon, the thalamencephalon,
and the hind-brain. Over these epithelial areas the pia mater is
unusually vascular and forms a series of "choroid plexuses." The
ventricular system is complete and continuous. By contrast with the
Lamprey the brain of Myxine[442] is very primitive, more so perhaps
than in any other Craniate (Fig. 212). In a dorsal view the brain is
divided into four pairs of laterally expanded and longitudinally
compressed lobes by a median longitudinal fissure and three
transverse fissures. The two anterior lobes are little more than the
thickened anterior wall of the thalamencephalon, although, judging
from their histological structure, they represent a very imperfectly
differentiated prosencephalon and olfactory lobes. The second and
largest pair constitute the thalamencephalon. The last two pairs of
lobes represent a transversely divided pair of optic lobes, or "corpora
quadrigemina." There is a large medulla oblongata with a pair of
corpora restiformia, but the cerebellum is entirely absent. The
ventricles are subject to some individual variation. Third and fourth
ventricles are generally recognisable, either as isolated cavities or
connected by a remnant of the mesocoele. In the feeble
development of the prosencephalon, in the striking preponderance
of the mid-brain over the rest of the brain, and in the absence of a
cerebellum, Myxine is unique amongst Craniates.

Fig. 213.—The brain of a Dog-Fish (Scyllium canicula). A, dorsal view;

B, ventral view. The choroid plexuses covering the roof of the
third and fourth ventricles have been removed. b.o, Olfactory
lobe; ep, origin of the stalk of the pineal body; f.b (in A),
prosencephalon; f.b (in B), cerebral hemispheres; fr, fourth
ventricle; h.b, cerebellum; h.p, pituitary body; i.f, lobi inferiores;
m.b, optic lobes; m.d, medulla oblongata; sc, saccus vasculosus;
th, thalamencephalon; t.o (i) olfactory peduncle; i.-x. cranial
nerves. (From Wiedersheim.)

In Elasmobranchs among Fishes the brain attains a much higher

grade of structure. In Scyllium (Fig. 213) there is a large
prosencephalon, and directly in front of it a pair of imperfectly
differentiated cerebral hemispheres, while from its antero-lateral
regions the large olfactory lobes arise. The prosocoele divides in
front into four diverticula, of which the two inner ones extend into
the hemispheres as lateral ventricles, and the two outer as
rhinocoeles into the olfactory lobes (Fig. 214). In connexion with the
infundibulum there is a pair of sacci vasculosi, consisting mainly of
gland-tubules, opening into the infundibular cavity.[443] The
cerebellum is exceptionally large, but it does not form a "valvula
cerebelli." Large ear-like corpora restiformia are present. The third
and fourth ventricles alone retain an epithelial roof in relation with
choroid plexuses.

In all essentials the brain of the Holocephali is a repetition of the

Elasmobranch type, more especially of the elongated form seen in
Notidanus. Indications of a higher grade of structure are, however,
to be seen in the reduction of the prosencephalon which, with its
prosocoele, is now scarcely distinguishable from the
thalamencephalon and its ventricle; and in the more complete
differentiation of the cerebral hemispheres from one another and
from the rest of the brain. Large frilled corpora restiformia are
conspicuous structures on each side of the medulla oblongata.
Besides the usual intra-cranial pituitary body, there is also a separate
extra-cranial portion lodged in a pit on the ventral surface of the
basis cranii: in the embryo the two are continuous.

Fig. 214.—Horizontal longitudinal section of brain of Chiloscyllium, to

show the ventricles; semi-diagrammatic. cer, Origin of cerebellar
ventricle or epicoele; dia, third ventricle; iter, mesocoele; meta,
fourth ventricle; opt, optocoele, or cavity of an optic lobe; para,
lateral ventricles; pros, prosocoele; rh, rhinocoele. (From Parker
and Haswell.)

In the Teleostomi the brain is distinctly of a more primitive type than

in any other Fishes (Fig. 215).[444] The most striking feature is the
absence of cerebral hemispheres, the evolution of the primary fore-
brain proceeding no farther than the formation of an undivided
prosencephalon with a non-nervous roof, and a prosocoele which
forms a continuous cavity with the third ventricle, or at the most is
only separated from it by an infolding of the epithelial roof or velum
transversum.

Fig. 215.—A, dorsal view of the brain of a Trout (Salmo fario); B, a

vertical longitudinal section. c.il, Commissura interlobularis; g.h,
ganglion habenulae; h.c, habenular commissure; i.c, inferior
commissure; l.i, lobus inferior; myc, myelocoele; p.c, posterior
commissure; v.o, valvula cerebelli; v.t, velum transversum; ii.,
optic nerve; v.iii., v.iv., third and fourth ventricles; v, vii, viii, ix, x,
fifth, seventh, eighth, ninth, and tenth cranial nerves; remaining
reference letters as in Fig. 210. (A, From Wiedersheim; B, after
Haller.)

Amongst other diagnostic characters may be mentioned the

predominance of the mid-brain over the other divisions, the anterior
extension of the large cerebellum into the mesocoele as a "valvula
cerebelli," and the absence of corpora restiformia. This type of brain
is most strongly marked in the Teleostei, but in other Teleostomes
some, like Acipenser,[445] are typically Teleostean in this respect
(Fig. 216), while others, such as Lepidosteus, have small cerebral
hemispheres with lateral ventricles as well as a prosencephalon.
 Fig. 216.—Vertical longitudinal section of the brain of a Sturgeon
(Acipenser ruthenus.) c.p, Posterior commissure; c.r, cranial roof;
mc, mesocoele; op.ch, optic chiasma; p.ch.p, posterior choroid
plexus; v.c, valvula cerebelli; v.t, velum transversum; v.iii, v.iv.,
the third and fourth ventricles; other lettering as in Fig. 210.
(From Goronowitsch.)

The most obvious feature in the brain of the Dipnoi is the great
development of the cerebral hemispheres. In this respect these
Fishes approach the Amphibia, but in other features of brain-
structure they present points of agreement with most other groups
of Fishes without being closely related to any one of them. In
Protopterus[446] (Fig. 217) the hemispheres are quite distinct except
behind, and the walls of their spacious lateral ventricles are entirely
nervous. Olfactory lobes are sessile on their anterior extremities, and
behind and below they enlarge into ventral lobes which probably
represent the hippocampal lobes of the higher Vertebrates. A
vesicular pineal body at the end of a slender stalk overlies a singular
conical projection from the roof of the thalamencephalon or "pineal
pillow."
 Fig. 217.—Dorsal (A), ventral (B), and lateral (C) views of the brain of
Protopterus annectens. C, Cerebellum; C.H, cerebral hemisphere;
D.S.E, branches of the sinus endolymphaticus; In, infundibulum;
L.I,, lobi inferiores; M.O, medulla oblongata; O.L, olfactory lobe;
Op.L, optic lobe; P, pituitary body; P.B, "pineal pillow"; S.E, sinus
endolymphaticus; Sp.c, spinal cord; Sp.n, spinal nerve; Vel, velum
transversum; Z, pineal body; IV.V., fourth ventricle; ii., iii., iv., v.,
vi., vii., viii.1, viii.2, viii.3.4, ix., and x., roots of the cranial nerves.
(From Burckhardt.)

The optic lobes form a single oval body, and, as in Petromyzon and
the Amphibia, the cerebellum is very small. A posterior choroid
plexus covers the roof of the fourth ventricle, and an anterior plexus
in connexion with the roof of the thalamencephalon projects
downwards into the third ventricle, and is also prolonged forwards
into each lateral ventricle. In Neoceratodus[447] the brain is certainly
more primitive and distinctly less Amphibian. As compared with
Protopterus the olfactory lobes and the cerebellum are larger, and
the optic lobes are paired. The smaller hemispheres are non-nervous
dorsally and medianly, the roof and inner wall of each being formed
by an extension of the thick, glandular choroid plexus which forms
the roof of the thalamencephalon.

The Spinal Nerves.—The spinal nerves of Cyclostomes (e.g.

Petromyzon) consist of a series of dorsal nerves arising on each side
from the dorsal surface of the spinal cord, and of a similar double
series arising from the ventral surface, the dorsal nerves regularly
alternating with the ventral nerves. Each myotome is supplied by a
dorsal and a ventral nerve which pass separately to their peripheral
distribution in the skin and muscles. In Fishes, as in the higher
Vertebrates, each dorsal nerve, now termed a dorsal root, enlarges
into a ganglion and then unites, either before or directly after issuing
from the neural canal, with the next ventral nerve or ventral root in
front to form a main spinal nerve. At the same time the spinal
nerves of opposite sides tend to form pairs in the same transverse
plane. After the union of the two roots the spinal nerve divides into
three typical branches: a dorsal nerve (ramus dorsalis), and a ventral
nerve (ramus ventralis), both of which include somatic sensory or
afferent fibres, and somatic motor or efferent fibres, for the
innervation of the skin and muscles of the dorsal and lateral portions
of a myotome; and a visceral branch (ramus visceralis), composed of
afferent and efferent visceral fibres, which supplies the adjacent
viscera (alimentary canal and its glands and blood-vessels), and
helps to form the sympathetic nervous system.[448] The somatic
afferent and the visceral afferent fibres enter the spinal cord by the
dorsal roots, the somatic efferent leaving the cord through the
ventral roots, although the visceral efferent fibres traverse both
roots. In the region of the paired fins more or fewer of the rami
ventrales unite to form a plexus, the brachial or the pelvic plexus,
from which the nerves to the fins take their origin.

The Cranial Nerves.—It is usual to describe the cranial nerves of

Cyclostomes and Fishes as consisting of ten serially disposed pairs,
viz.: the olfactory (i.), optic (ii.), oculomotor (iii.), trochlear (iv.),
trigeminal (v.), abducens (vi.), facial (vii.), auditory (viii.),
glossopharyngeal (ix.), and the vagus (x.) Like the spinal nerves, the
cranial nerves collectively include somatic sensory (general
cutaneous) and motor fibres, and also visceral sensory and motor
fibres, all of which have their own special centres in the brain, but
the proportions of these nerve components differ greatly in different
nerves. Certain preoral nerves (iii., iv., and vi.) are exclusively
somatic motor; others (i. and ii.) are special sensory nerves for the
olfactory and visual organs; but most of the other cranial nerves
include several components, and are therefore "mixed" nerves.
Besides these components some cranial nerves include also a quasi-
independent system of nerve-fibres, which converge from certain
cutaneous sense-organs to an independent centre in the medulla
oblongata, the tuber acusticum,[449] and is probably derived from
the general cutaneous system of nerve components. Such nerve
fibres, including also the auditory nerve, which has its origin from
the same centre, constitute the lateralis system. Perhaps the most
striking feature in the postoral cranial nerves is the predominance of
the visceralis or sympathetic system over the somatic. Omitting the
lateralis fibres and a relatively few somatic sensory fibres, visceral
fibres, sensory and motor, are the principal components of all these
nerves, including v. but excluding viii. The reason for this is to be
found in the fact that splanchnic or visceral muscles in relation with
the jaws and branchial arches have usurped the place of somatic
muscles in the muscular system of the head. For developmental and
other reasons the olfactory and optic nerves stand in a category of
their own, and the same may be said of the third, fourth, and sixth
nerves, which innervate the muscles of the eyeball. The remaining
nerves, all of which have their origin in the medulla oblongata,
possess certain features in common, and as they are related to the
gill-clefts in such a way that each forks over a cleft, they may be
conveniently distinguished as "branchial" or "branchiomeric nerves."
A typical branchial nerve consists of (1) a principal ganglion near the
origin of the nerve from the brain; (2) a main trunk which gives off
(3) a somatic sensory branch or dorsal nerve to the skin; (4) a
palatine nerve (visceral sensory) to the oral or pharyngeal mucous
membrane; (5) an epibranchial ganglion which is associated with a
transitory embryonic epibranchial sensory organ at the dorsal border
of a branchial cleft; (6) a pre-branchial nerve (visceral sensory),
skirting the anterior margin of a cleft in its ventral course; and (7) a
post-branchial branch (visceral motor) similarly related to the hinder
margin.

Fig. 218.—Diagram showing the principal branches of the cranial nerves

in a Fish, mk.c, Meckel's cartilage; ol.o, olfactory organ; p.q,
palato-quadrate; s, spiracle; i-v, branchial clefts; i, ii, iii, iv, vi, the
first, second, third, fourth, and sixth cranial nerves. The remaining
nerves are differently shaded. Black.—The trigeminal nerve: g.g,
Gasserian ganglion; md, mandibularis; mx, maxillaris; op.p,
ophthalmicus profundus. Oblique shading.—The lateralis system
and its centre (t.a), the tuber acusticum: bucc.vii, buccalis branch
of vii; md.ex, external mandibular branches of vii; l.n.x, lateralis
nerve, with its supra-temporal branch (s.t), and its commissural
connexion (c) with op.s.vii, the ophthalmicus superficialis of vii;
viii, auditory nerve. Dotted.—The facialis proper, including c.t,
chorda tympani; gn.g, geniculate ganglion; hy, hyomandibularis,
with its motor branches m, m, m; p.n.vii., palatine. Dark grey.—
The glossopharyngeal (ix), with its pre- and post-branchial
branches and its palatine nerve, p.n.ix; anastomosing with the
palatine branch of vii (Jacobson's anastomosis). White.—The
vagus: x1-4, the branchial nerves, ganglionated and forking over
clefts ii-v; v.n.x, visceral nerve; oc, occipito-spinal nerves; d.r and
v.r, the dorsal and ventral roots of the first two spinal nerves.
(Slightly modified after Wiedersheim.)

The first six cranial nerves resemble those of the higher Craniates in
their mode of origin from the brain, in the physiological nature of
their component fibres, and in their peripheral distribution, and
therefore they need not be specially referred to here. The principal
branches of the fifth or trigeminal nerve are shown in Fig. 218.
Comparing this nerve with a typical branchial nerve it would seem
that the profundus and superficialis ophthalmic nerves are dorsal
nerves; the maxillaris and mandibularis, pre- and post-branchial
branches, respectively, in relation with the modified gill-cleft which
forms the mouth, while the branch to the oral surface represents a
palatine nerve. The most important of the distinctive features in the
cranial nerves of Fishes are to be found in the relations of nerves vii.,
ix., and x. to branchial clefts, and in the lateralis system of nerve
components and its association with the lateral line sensory organs.
The seventh or facial nerve is an exceptionally interesting nerve.
Besides the usual components of a typical branchial nerve certain of
its so-called branches are wholly or largely derived from the lateralis
system. For this reason the nerve may be said to consist of two
portions, the facial proper, or those fibres which constitute the facial
nerve in air-breathing Craniates, and the lateralis branches which
solely innervate lateral line sense-organs, and are therefore peculiar
to aquatic forms. The facial proper has a ganglion (the facial or
geniculate ganglion) on its root, and on entering the orbit after
traversing the cranial wall it gives off a palatine nerve. Just over the
spiracle a pre-branchial nerve, the representative of the chorda
tympani of Mammals, leaves the main trunk, and passes ventrally in
relation with the anterior wall of the spiracle to its ultimate
distribution in the walls of the mouth cavity. The main trunk, now
called the ramus hyomandibularis, then pursues a ventral course
behind the spiracle as a post-branchial nerve, and certain of its
mainly motor branches which pass downwards in connexion with the
hyoid arch supply the muscles of that arch, and, if an operculum is
present, the opercular muscles as well. The lateralis portion of the
facial includes the following principal branches, each of which may
have a ganglion on its root: (1) an ophthalmicus superficialis; (2) a
buccalis nerve with its ramus oticus; and (3) external mandibular
nerves which course in the ramus hyomandibularis. The addition of
the great lateralis nerve, which is usually described as the lateral
branch of the tenth nerve, and of the eighth or auditory nerve which
supplies the auditory organ, completes the enumeration of the main
factors of the lateralis system. The ninth or glosso-pharyngeal nerve,
perhaps the most typical of all the branchial nerves, has pre- and
post-branchial branches which enclose the hyo-branchial cleft. Its
palatine nerve usually extends forwards and anastomoses with the
corresponding branch of the seventh, thus forming a connexion
(Jacobson's anastomosis) between the two cranial nerves. In some
Elasmobranchs and Teleosts fibres derived from the dorsal branch of
the ninth nerve innervate a few sense-organs of the lateral sensory
canal of the head, and hence that nerve sometimes contains lateralis
fibres. The tenth or vagus is a compound nerve. Besides the great
lateralis nerve generally associated with it, the vagus includes as
many typical branchial nerves as there are branchial clefts behind
the hyo-branchial cleft, and in Elasmobranchs and in Chimaera these
nerves have independent origins from the medulla oblongata. Each
nerve has the typical structure, a ganglionated trunk which forks
over a gill-cleft into the usual pre- and post-branchial branches, and
palatine branches to the pharyngeal walls. In the Dipnoi the lateralis
nerve is connected with the superficial ophthalmic branch of the
seventh nerve by a commisural nerve which curves across the outer
face of the auditory capsule. A somewhat similar anastomosis is also
present in Petromyzon. The vagus also includes a large ramus
intestinalis, which in Elasmobranchs, at all events, has a distinct
ganglionated root. The nerve forms characteristic plexuses on the
oesophagus and stomach, and in Cyclostomes its branches may
extend nearly the whole length of the intestine. In Ganoids and
Teleosts there is an interesting nerve known as the "lateralis
accessorius." It is a compound nerve, and owes its formation to the
union of somatic sensory fibres derived in succession from dorsal
branches of the v., vii., ix., and x. nerves, and also from the
corresponding branches of a variable number of spinal nerves. The
finer branches of the nerve are distributed to the skin of one or more
of the fins, or even, as in Gadus, to all the fins, especially to the
numerous "end-buds" which are present on those organs. In many
Fishes a variable number of the anterior spinal nerves (spino-
occipital) perforate the occipital region of the skull. They probably
represent the ventral roots only of the ordinary spinal nerves of this
region.
 Sense-Organs

The Cutaneous Sense-Organs.—These organs, the most

remarkable and certainly the most characteristic of the sense-organs
of Cyclostomes and Fishes, are bud-like groups of epidermic cells in
relation with the ends of sensory nerve fibres. Each consists of a
central core of sensory cells, provided with terminal cuticular sensory
hairs, and surrounded by a zone of supporting and mucus-secreting
cells which leave the hairs exposed at the apex of the bud. Two
kinds of these organs can be distinguished, which differ in their
innervation and in their position in the skin. Of the two, the so-called
end-buds are the more primitive. They occupy a superficial position
in the epidermis, and their sense-cells are as long as the supporting
cells. They are present in Cyclostomes and Elasmobranchs, and
especially in Teleosts, where they are irregularly distributed over the
surface of the body, on the fins, lips, and barbels, and also in the
epithelium of the mouth and pharynx. In the Dipnoi they are limited
to the oral cavity, and in the higher Craniates they become taste-
buds.[450] Their somatic sensory nerves[451] are derived from the
vii., ix., and x. cranial nerves, and the lateralis accessorius. In the
second type, usually called "nerve-eminences," the sensory cells are
shorter than the supporting cells, and they are always innervated by
the lateralis system. When first developed in the embryo they are
quite superficial, like end-buds, but later the epidermis in which they
lie sinks inwards so as to line a series of pits, closed sacs, tubes,
open grooves, or closed canals. Pit-organs, so abundant on the head
and trunk of Teleosts (Fig. 220), are simple epidermic pits with
insunken nerve-eminences, disposed in groups or in lines (accessory
lateral lines) or irregularly distributed. The "Spalt-papillen" of
Elasmobranchs are pit-organs in which the orifice of the pit is
reduced to a slit. The more deeply-seated Savi's vesicles on the
ventral surface of the Torpedo, and the nerve-sacs of Ganoids, are
similar organs converted into closed sacs and pinched off from the
rest of the epidermis. Lorenzini's ampullae or mucus canals, which
are found in definitely located groups on the lateral and upper
surfaces of the head in Elasmobranchs, may perhaps be compared
to pit-organs prolonged inwards to form subcutaneous tubes, each
of which terminates in a radially-septate, chambered dilatation or
ampulla, containing groups of sensory cells.

Besides the more diffusely scattered sense-organs there are others

which become disposed in definite lines along the sides of the body
and on the head, and, enclosed in grooves or closed canals,
constitute the highly characteristic lateral line system of Cyclostomes
and Fishes.[452] The auditory organ must also be included as a
specialised portion of this system. Both organs are innervated by the
lateralis system, and both arise from a common rudiment in the
embryonic epidermis in the position of the future auditory organ.
This rudiment grows backwards along the side of the body in the
form of a cord of cells differentiated from the epidermis, and also
forwards, where it soon divides into the rudiments of future supra-
orbital and infra-orbital canals. Sense-organs are differentiated at
intervals along the line of the cord; and in the body, but not on the
head, they frequently exhibit a segmental disposition. Each sensory
organ then sinks down into a short epidermic groove, which by the
subsequent meeting of its lips becomes a canal detached from the
epidermis. The short canals then become continuous, leaving,
however, an externally opening primary pore between every two
consecutive canals, and the result is a continuous canal having
sense-organs imbedded in its epidermic lining and connected with
the exterior by pores at intervals (Fig. 219).[453] The enclosure of
the canals in the scales of the lateral line of the trunk or in special
drain-pipe ossicles on the head, and the dichotomous subdivision of
the primary pores into groups of surface-pores, complete the
evolution of the system in its more advanced condition.
 Fig. 219.—Vertical longitudinal section through the lateral canal of Amia
calva. l.n, Lateralis nerve with its branches, n, n, to the sensory
organs, s.o, s.o; p, p, p, external pores; s.c, sensory canal; s, s,
scales of the lateral line. (From Wiedersheim, after Allis.)

Typically, the lateral line system consists of certain canals or

grooves, usually but not invariably continuous, and defined by their
innervation, (i.) a lateral canal extending along the side of the body
and the hinder part of the head, and having its sensory organs
supplied by the great lateralis nerve (Fig. 220); (ii.) a supra-orbital
canal passing forwards over the eye and innervated by the
superficial ophthalmic branch of the facial nerve; (iii.) an infra-orbital
canal supplied by the buccalis and otic branches of the same nerve;
and (iv.) a hyo-mandibular or operculo-mandibular canal, situated on
the outer side of the hyoid region, and thence prolonged downward
and forward in relation with the lower jaw, and innervated by the
external mandibular branches of the facial nerve. The hyo-
mandibular canal is sometimes distinct from the other canals, as in
Elasmobranchs and some Teleosts (Fig. 220); and in certain North
American Siluroids the same may be said of the supra-orbital. But,
as a rule, the infra-orbital is continuous behind both with the lateral
and the supra-orbital canals, while the hyo-mandibular canal joins
the infra-orbital, or, exceptionally, the supra-orbital canal. Transverse
commissural canals often connect the lateral and supra-orbital canals
of opposite sides across the dorsal surface of the head, and the
corresponding infra-orbital and hyo-mandibular canals may also be
continuous at the extremity of the snout or at the mandibular
symphysis.
 Fig. 220.—Sensory canals of the left side of the head of Gadus virens.
e, Eye; i.o, infra-orbital canal (dotted); l.c, lateral canal (oblique
shading); n, nasal apertures; op, operculum; op.m, operculo-
mandibular canal (longitudinal shading); p.o, pit-organs; s.o,
supra-orbital canal (cross-hatched); s.o.c, supra-orbital
commissure; s.t, supra-temporal branch; t.t, tubuli by which the
canals communicate with the exterior. (From Cole.)

Throughout their extent the canals communicate with the exterior by

pores, or short canals terminating in pores, or by branched canals
ending in groups of pores. In Cyclostomes[454] the lateral line
system is represented by pit-organs disposed as in Fishes, and
innervated by a true lateralis nerve. Some Elasmobranchs have the
lateral canal of the trunk represented by an open groove protected
by marginal denticles. Chimaera is more primitive still in this respect,
for on the head as well as on the body the sensory organs are in
open grooves. Amongst Fishes these organs are most primitive in
the Dipnoi, where they retain their superficial position in the
epidermis. In Teleostomes the lateral canals perforate the scales of
the lateral line, and at intervals they open externally by simple or
multiple pores which perforate the scales. On the head they are
more or less completely enclosed in special ossicles which either
remain distinct or fuse with certain of the adjacent dermal or
cartilage bones of the skull. The use of the lateral line organs is not
certainly known. They occur only in Fishes and Amphibia, and as
blind Fishes are able to avoid obstacles with the greatest ease when
swimming, it is possible that these organs enable their possessors to
appreciate undulatory movements in water in the shape of reflex
waves from contiguous surfaces or objects.[455] Their great antiquity
is shown by their existence in most of the Heterostraci, and in the
Antiarchi and Arthrodira, although they have not yet been discovered
in the Osteostraci.

The Auditory Organs.—In its more typical condition each auditory

organ consists of a membranous sac or vestibule, partially
constricted into an upper portion or utriculus and a lower or sacculus
(Fig. 221, A). Three semicircular canals are connected with the
utriculus, of which two are vertical and at right angles to one
another, and the third is horizontal. One end of each canal is dilated
into an ampulla. A slender tube, the ductus endolymphaticus, leaves
the sacculus, and ends in a sac-like swelling, the sinus
endolymphaticus, which apparently represents a portion of the
embryonic epidermic involution from which the auditory organ is
formed. A smaller sac-like outgrowth from the sacculus, the lagena,
corresponds to the cochlea of the higher Vertebrates. The epidermic
lining of this system of cavities is differentiated into patches or
ridges of sense-cells (maculae or cristae), separated by supporting
cells and innervated by the terminal branches of the auditory nerve.
There is a crista acustica in each ampulla; and maculae acusticae are
present in the utriculus, sacculus, and lagena. A fluid, the
endolymph, fills all the cavities, and a similar fluid or perilymph
occupies the spaces in the periotic capsule in which the various
chambers are lodged. Among the more notable deviations from this
type of auditory organ the Cyclostome Myxine, apparently, has but a
single semicircular canal with an ampulla at each end, and the
vestibule is a simple sac (Fig. 221, B). Petromyzon has two canals,
but lacks the horizontal canal. In Elasmobranchs, including Chimaera
(C), the ductus endolymphaticus retains its primitive connexion with
the exterior by means of a pore on the dorsal surface of the head. In
the Dipnoi (e.g. Protopterus) the paired endolymphatic sinuses
divide into a number of caecal branches containing otoliths, which
meet and interlace over the fourth ventricle (Fig. 217).[456] Otoliths,
either in the form of fine, mucus-connected, calcareous particles, as
in Elasmobranchs, or as massive solid concretions in Teleosts, are
present in relation with the sensory areas of the utriculus, sacculus,
and lagena.

Fig. 221.—Auditory organs of Fishes. A, of a typical Fish; B, of Myxine;

C, of Chimaera; and D, of Perca. a.c, Anterior canal; am', am",
am'", ampullae; am.n, nerves to ampullae; c, semicircular canal
(in Myxine); d.e, ductus endo-lymphaticus; h.c. horizontal canal;
l, lagena; mc, macula acustica; m.s, macula acustica of the
sacculus; n, nerves to ampullae; o, external aperture of the
ductus endo-lymphaticus; p.c, posterior canal; s, sacculus; s.e,
sinus endo-lymphaticus; sk, superficial skin; s.s, sinus superior; u,
utriculus; viii, auditory nerve. (From Wiedersheim, after Retzius.)

In a few marine and in a large number of freshwater Teleosts the

auditory organ enters into a more or less intimate connexion with
the air-bladder by one of three different methods.

Fig. 222.—Cavity of the air-bladder of a Siluroid (Macrones nemurus)

exposed by the removal of its ventral wall. a.c, Anterior chamber;
b.o, basioccipital; b.w, body wall, here reduced to the external
skin; cl, clavicle; l.c, lateral chamber; l.s, longitudinal septum; pt,
 post-temporal; tr.a, anterior portion of the tripus; tr.c, crescentic
portion of the tripus; t.s, transverse septum; t.s', shorter
transverse septum. (From Bridge and Haddon.)

The first and simplest is by the apposition of the extremities of a pair

of caecal tubular prolongations from the air-bladder to the outer
surfaces of the fibrous membranes which close a pair of vacuities in
the outer bony walls of the periotic capsules, the inner surfaces
being bathed by the perilymph surrounding the auditory organs. This
method is characteristic of certain Serranidae, Berycidae, Sparidae,
Gadidae, and Notopteridae,[457] and probably in the Hyodontidae. In
the second method, of which several Clupeidae (e.g. Herring,
Pilchard, etc.) furnish examples, the periotic vacuities are open
instead of closed, and the sac-like ends of the tubular extensions
from the air-bladder are in actual contact with protruding outgrowths
from the utriculus.[458] The third method, by far the most elaborate,
is by the intervention of a series of movably connected "Weberian"
ossicles, of which the most posterior on each side (the tripus) is
inserted into the dorsal wall of the air-bladder (Fig. 223), while the
anterior one (scaphium) forms the outer wall of a median backward
prolongation (sinus impar) of the perilymph-containing spaces
surrounding the two auditory organs. This in turn encloses a similar
median prolongation (sinus endolymphaticus) from the two sub-
cerebrally united endolymphatic ducts (Fig. 223).[459] This complex
mechanism is present in the Cyprinidae, Siluridae, Characinidae, and
Gymnotidae; and hence the term "Ostariophysi"[460] as a collective
name for these families.[461] The physiological raison d'être of the
connexion between the air-bladder and the auditory organ cannot
yet be regarded as satisfactorily determined. It is possible, as Weber
thought, that it may be an auxiliary to the function of hearing by
transmitting to the ear sound-waves impinging on the surface of the
body and affecting the gases in the air-bladder.[462] On the other
hand, it may be urged with perhaps greater probability that the
connexion exists for the purpose of conveying to the ear stimuli due
to the varying degrees of distension of the air-bladder, such as, it
may be presumed, are naturally brought about by the variations of
hydrostatic pressure which a Fish encounters in the course of its
ascent or descent in the water.[463] Whether regarded as an
accessory to hearing, or as a means of regulating the distension of
the air-bladder, the physiological value of the connexion must be
considerable, and on this point it is at least significant that the
Weberian mechanism is characteristic of the dominant families of
freshwater Teleosts at the present day.[464]

Fig. 223.—Diagram to show the Weberian ossicles and their relations to

the ear and the air-bladder. at, Atrium, an extension of the sinus
impar; a.v.c, anterior vertical canal; b.w, bony wall of the periotic
capsule; d.e, the medianly-united endolymphatic ducts; h.c,
horizontal canal; in, intercalarium, a third ossicle imbedded in the
ligament (i.lg) connecting the scaphium with the tripus; n, bony
nodules on the sides of the complex vertebral centrum; p.v.c,
posterior vertical canal; s, sacculus; sc, scaphium; s.e, sinus
endolymphaticus; s.i, sinus impar; tr.a, tr.c, the anterior and
crescentic parts of the tripus; ut, utriculus. The radial lines
represent the fibres of the dorsal wall of the air-bladder. (From
Bridge and Haddon.)

The Olfactory Organs.—These organs are essentially a pair of pit-

like inpushings of the skin of the ventral side of the head in front of
the mouth, with their lining epidermis differentiated into sensory
cells separated by supporting cells, and connected with the olfactory
lobes of the brain by olfactory nerves.
 Fig. 224.—Two stages in the development of the olfactory organ and
the pituitary involution in Petromyzon. A is the earlier, B a much
later stage. br, Brain; in, infundibulum; l.lp, lower lip; ms,
mesenteron; n, notochord; ol.o, olfactory organ; pn, pineal body;
pt.s, pituitary sac; st, stomodaeum; u.lp, upper lip. (From Parker
and Haswell, after Dohrn.)

The Cyclostomata are unique amongst Craniates in the apparently

unpaired condition of the olfactory organ, and in its remarkable
relation to the pituitary involution. In the embryo Lamprey the
median and ventral olfactory pit is carried inwards with the pituitary
invagination, so that the former appears as a dorsal outgrowth from
the latter, and the two have a common external opening, the naso-
pituitary aperture (Fig. 224). Later the extraordinary forward growth
of the upper lip to form the roof of the buccal funnel has the effect
of shifting the naso-pituitary involution and its aperture to a final
position on the dorsal side of the head. It is due to this dorsal
displacement that, as we shall see, the pituitary caecum reaches the
ventral surface of the brain by perforating the basis cranii from
above, instead of from below as in all other Craniates. The pituitary
body is pinched off from the dorsal side of the naso-pituitary
involution. In the adult Lamprey the olfactory organ appears as a
round sac divided by a median septum into two lateral chambers
(Fig. 225), the lining epithelium of which is raised into prominent
ridges. Behind the sac the pituitary involution is prolonged
backwards beneath the brain, and, after traversing the basi-cranial
fontanelle, it widens out into a spacious cul-de-sac and terminates
on the dorsal side of the pharynx, beneath the anterior end of the
notochord. In Myxine the pituitary involution ends by opening into
the pharynx.
 Fig. 225.—Side view of the brain of Petromyzon, with the olfactory
organ and the pituitary caecum in section. cblm, Cerebellum;
crb.h, cerebral hemisphere; dien, thalamencephalon; f, fold in the
nasal tube; gl, nasal glands; inf, infundibulum; l.gn.hb, left
ganglion habenulae; med.obl, medulla oblongata; na.ap, naso-
pituitary aperture; n.ch, notochord; Nv1-nv10, cranial nerves;
Nv12, first ventral spinal nerve; olf.cp, olfactory capsule; olf.l,
olfactory lobe; olf.m.m, olfactory mucous membrane; opt.l, optic
lobe; pn, pineal body; pn′, inferior pineal body; pn.e, parietal eye;
pty.b, pituitary body; pty.p, pituitary cul-de-sac; sp, median
septum of the olfactory sac; sp1, first dorsal spinal nerve. (From
Parker and Haswell, after Ahlborn and Kaenische.)

The apparently monorhinal condition of the Cyclostomes is probably

a secondary acquisition. At the earliest embryonic stage at which any
trace of an olfactory organ is apparent, there is a median thickening
of the epidermis, possibly a vestige of some older sensory organ
comparable, it may be, to the so-called olfactory organ of
Amphioxus; on each side of it there is a lateral thickening, the
rudiments of the paired organs.[465] The three thickenings, or
"plakodes," then sink inwards to form an olfactory pit. The partial
subdivision of the adult organ by a vertical septum, and the
presence of two olfactory nerves, point to the same conclusion.[466]
All Fishes possess olfactory organs which are obviously paired. In
Elasmobranchs and Dipnoi they retain their primitive ventral position.
Many Sharks and Dog-Fishes possess an oro-nasal groove leading
from each olfactory organ to the corresponding angle of the mouth.
The Dipnoi proceed a stage farther, and, by the conversion of the
grooves into short canals, the olfactory pits communicate with the
mouth by true internal nostrils, as in the higher Vertebrates. In the
adults of existing Teleostomi the orifice of each organ is usually
divided into two by the growth of a fold of skin across it, and the
two apertures rotate outwards and upwards on to the lateral or the
upper surface of the snout. Of the two nostrils the posterior one
probably corresponds to an external nostril, and the anterior one to
the internal nostril. Occasionally each olfactory organ has only a
single orifice. In the Crossopterygii and in some Teleostei the nostrils
become tubular. The lining epithelium of the olfactory pits is usually
produced into ridges, disposed longitudinally or transversely, or in
the form of radii from a central point in the roof. Many Teleosts have
each olfactory organ prolonged backwards into one or two sacs, the
nasal sacs, which are either simple reservoirs, or glandular and
mucus-secreting. In a species of Chinese Sole (Cynoglossus
semilaevis) the two sacs, one from each olfactory organ, unite over
the roof of the mouth in a common median sac, and in one unique
specimen the latter communicated with the mouth by a large naso-
pharyngeal aperture.[467]

The Eyes.—In essential structure the eyes of Cyclostomes and

Fishes resemble those of the higher Craniates. As a rule, in Fishes
they are relatively larger, however, and the lens is globular and the
cornea somewhat flatter. Ciliary processes and ciliary muscles are
absent. As the eyes are nearly always lateral in position it is probable
that monocular vision is the rule. In Teleosts and in Amia a "choroid
gland," consisting of a mass of capillary blood-vessels, surrounds the
optic nerve externally to the retina, and derives its blood from the
efferent artery of the pseudobranch (Fig. 226). In most Teleostomi,
but not in Cyclostomes, Elasmobranchs, and Dipnoi, there is a
singular prolongation of the choroid coat, known as the "processus
falciformis," which extends across the vitreous humour to the inner
face of the lens, where it ends in an expansion, the "campanula
Halleri" (Fig. 226). Accommodation to vision at different distances is
not effected by alterations in the convexity of the lens, but by a
change in its position with regard to the retina, apparently brought
about by the contraction of a special retractor muscle.[468] Some
oceanic pelagic Teleosts are remarkable for their curious telescopic
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