Вы находитесь на странице: 1из 99

CONTENTS

Serial

Topic

Page

no.

no.

01.

Acknowledgement

02

02.

Abstract

03

03.

SPAN Laboratories Pvt. Ltd. A Company Insight

05

04.

Products of SPAN Laboratories Pvt. Ltd.

08

05.

Sales and Marketing Strategy adopted by SPAN Laboratories

13

Pvt. Ltd.

06.

Ayurveda An Overview

18

07.

Scope of Ayurvedic Medicine

23

08.

How the Ayurvedic Tradition became a system of Empirical

27

Medicine

09.

Quality Control of Ayurvedic Drugs

30

10.

Protocols for testing the quality of Oils, Paste, Tablets Powder

54

& Syrup

11.

Results

88

12.

Conclusion

94

13.

Bibliography

99

1

ACKNOWLEDGEMENT

This project has been made possible with the help of a lot of people. My learning wou ld be incomplete if I don‘t acknowledge people who catalysed this process, Starting with the key stone of the edifice Mr. Subhash Chandra (Managing Director) who obliged me by giving an opportunity to undergo a short term summer training program under his esteemed guidance at Span Laboratories Private Limited, Lucknow.

I would also like to thanks Mr. Prabhash Chandra (Production Manager) and Mr. Guddu (Assistant) for their help during my training.

I would also like to extend my gratitude towards Mr. A. K. Saxena (Packaging Manager), Mr.

Swapnil Chandra (Marketing Head-OTC Division) and Mr. Saurabh (Assistant) for giving their valuable time, guidance suggestions and critical comments which aided my learning and work.

I would specifically like to thank all the employees of the Span Laboratories Private Limited for the help and cooperation provided.

I also thank my friends Hemant, Sudhanshu and Ashutosh for their cooperation and support.

I would also like to thanks my parents and God without whose blessings it would not have been so easy.

(Mudit Misra)

2

ABSTRACT

Medicinal plants constitute a source of raw materials for both traditional systems of medicine (e.g. Ayurvedic, Unani, Homeopathy, and Siddha) and modern medicine. Nowadays, plant materials are employed throughout the industrialized and developing world as home remedies, over-the-counter drugs, and ingredients for the pharmaceutical industry. As such, they represent a substantial proportion of the global drug market. Most rural populations, especially in India, depend on medicinal herbs as their main source of primary health care. Although most medicinal herbs are not, in their natural state, fit for administration, preparations suitable for administration are made according to pharmacopeia directions. The therapeutic potential of a ayurvedic drugs depends on its form: whether parts of a plant, or simple extracts, or isolated active constituents. Ayurvedic remedies consist of portions of plants or unpurified plant extracts containing several constituents, which often work together synergistically. The ayurvedic drug preparation in its entirety is regarded as the active substance and the constituents are either of known therapeutic activity or are chemically defined substances or group of substances generally accepted to contribute substantially to the therapeutic activity of the drug. Phytochemical screening involves botanical identification, extraction with suitable solvents, purification, and characterization of the active constituents of pharmaceutical importance. Qualitative chemical examination employing different analytical techniques is conducted to detect and isolate the active constituent(s). In general, all medicines, whether they are synthetic or of plant origin, should fulfill the basic requirements of being efficacious and safe. Ultimate proof of these can only be achieved by some form of quality testing. A defined and constant composition of the drug is therefore one of the most important prerequisites for any kind of clinical experiment.

Quality control for the efficacy and safety of ayurvedic products is essential. The quality control of phytopharmaceuticals may be defined as the status of a drug, which is determined either by identity, purity, content, and other chemical, physical or biological properties, or by the manufacturing process. Compared with synthetic drugs, the criteria and the approach for ayurvedic drugs are much more complex.

3

Phytopharmaceuticals are always mixtures of many constituents and are therefore very variable and difficult to characterize. The active principle(s) in phytopharmaceuticals are not always known. The quality criteria for ayurvedic drugs are based on a clear scientific definition of the raw material. Depending on the type of preparation, sensory properties, physical constants, moisture, ash content, solvent residues, and adulterations have to be checked to prove identity and purity. Microbiological contamination and foreign materials, such as heavy metals, pesticide residues, aflatoxins, and radioactivity, also need to be tested for. To prove the constant composition of ayurvedic preparations, appropriate analytical methods have to be applied and different concepts have to be used in order to establish relevant criteria for uniformity.

With many of these ayurvedic medicines we do not fully understand how they work. Nor do we always know which component is pharmacologically active. Even though ayurvedic remedies may be effective, do their benefits outweigh their risks? In some countries ayurvedic remedies are sold as food supplements, thus evading safety regulations. Can ayurvedic medicines save money? Not all plant-based medicines are cheap.

Even though global herbal resources have a great potential as natural drugs and are of great commercial importance, they are very often procured and processed without any scientific evaluation, and launched onto the market without any mandatory safety and toxicology studies because there is no effective machinery to regulate manufacturing practices and quality standards. Although some ayurvedic medicines are efficacious, there is unquestionably a need for more reliable information, a demand that must be met adequately by doctors, pharmacists, and other health care professionals.

4

SPAN LABORATORIES PRIVATE LIMITED A Company Insight

Span Laboratories Private Limited is a GMP certified company. It is a leading name in the field of pharmaceutical products. The company was established in 1996 at Gomti Nagar, Lucknow. Thereafter, the company was incorporated on 6th April, 1996.

The company has been in the field of manufacturing and marketing of ayurvedic pharmaceutical products since fourteen years. Span Laboratories carry a range of ayurvedic products to suit every task great or small. Span Laboratories supply a range of liver stimulants, products for digestive system, laxative drugs to suit every one. The company is well equipped with large number of state-of-art machineries and a dynamic and dedicated well trained workforce.

Span Laboratories is headed by Mr. Subhash Chandra, Mr. Prabhash Chandra and Mr. S.P.

Saxena as Directors. It is managed by a dedicated team of skilled professionals and highly

team.

trained

sales

The company is highly grateful to its team for its dedication and help. What we are is just because of the support and dedication of our team. The Span Laboratories team consists of internationally experienced sales executives, production managers and technology experts. Our team works in co-ordination with each other, which are enthusiastic as well as industrious in nature.\

team is headed by Mr.Suhail Sahid, Mr.Syed Ifrahim and Mr.Jay Prakash

Saxena who have more than 15 years of experience in pharmaceutical industry. Each of them possesses a specialized knowledge of industry. They carry out our global sales operations rivigorously and with great spirit. The unique combination of our team add to the value of our

company and opens new perspectives for making sound decisions.

Our Sales

MISSION AND VISION At Span Laboratories Private Limited our vision is to provide quality services to our customers. Our goal is to achieve open rewarding workplace environment which enables our colleagues to realize their full potential.

5

Our Mission is to achieve excellence in all respect. We will act with honesty in dealing with each other, our customers, our employees, our chemist and all other people who are in contact with us. Our vision revolves around quality production, teamwork, honesty, integrity and developing environment in all aspect of our performances and business strategies.

PRINCIPLES AND CORE VALUES OF SPAN

Ownership: Accept personal responsibility, and accountability to meet the business needs.

Passion for Winning: Be the leaders in area of responsibility, with a deep commitment to deliver results.

People Development: Our ethics are considered to be the most important asset. Value addition is done through result driven training, and they encourage reward excellence.

Consumer Focus: Give importance to customer needs and develop products to fulfil them better.

Team Work: Work together on the principle of mutual trust and transparency in a boundary less organization. Team members are intellectually honest in advocating proposals, including risks involved.

Innovations: Continuous innovations in products and processes are the basis of our success.

Integrity: Achievement of business success with integrity is another important principle. We believe in honesty with customers, business partners and also with each other.

STRATEGIC INTENT

Focus on growing core brands across categories, reaching out to new geographies, within and outside India, and improve operational efficiencies by leveraging technology.

6

Be the preferred company to meet the health and personal grooming needs of our target consumers with safe, efficacious, natural solutions by synthesizing our deep knowledge of ayurveda and herbs with modern science.

Build a platform to make SPAN Labs. an indian as well as a global ayurvedic leader.

Be a professionally managed employer of choice, attracting, developing and retaining quality personnel

Commitment towards environmental protection

Provide superior returns, relative to our peer group, shareholders.

QUALITY POLICY At SPAN, quality is a relentless commitment to continuous improvements in product, process and systems to provides consistent quality products to meet our customer‘s requirement worldwide. The management is fully committed to quality and ensures all resources to accomplish this task. Our Quality Objectives include:

Focus on our customers and successfully meet their needs and requirements.

Manufacture effective health care products at competitive prices and to improve the quality of life of the common masses.

Implement systems to ensure prevention of errors either than detection of errors.

Ensure global competitiveness by striving to achieve current good manufacturer practices (GMP).

Ensure safety in all operations by working according to the system in all areas of operations.

Increase productivity and reduce wastage with in the organization.

Provide appropriate training to the people to improve their skills and expertise, thus strenghthening our commitment to the quality process.

7

PRODUCTS OF SPAN LABORATORIES

DIGENORM SYRUP, CAPSULES & TABLETS

PRODUCTS OF SPAN LABORATORIES  DIGENORM SYRUP, CAPSULES & TABLETS  EVACUA POWDER 8

EVACUA POWDER

PRODUCTS OF SPAN LABORATORIES  DIGENORM SYRUP, CAPSULES & TABLETS  EVACUA POWDER 8

8

LIVGUARD SYRUP

 LIVGUARD SYRUP  REGAIN SYRUP 9

REGAIN SYRUP

 LIVGUARD SYRUP  REGAIN SYRUP 9

9

RHA OIL

 RHA OIL  SENSATION CAPSULE 10

SENSATION CAPSULE

 RHA OIL  SENSATION CAPSULE 10

10

SPANDRYL SYRUP

 SPANDRYL SYRUP  SPANLIV DS SYRUP, TABLETS & CAPSULES 11

SPANLIV DS SYRUP, TABLETS & CAPSULES

 SPANDRYL SYRUP  SPANLIV DS SYRUP, TABLETS & CAPSULES 11

11

UREE SYRUP

 UREE SYRUP  U TONE SYRUP & CAPSULES 12

U TONE SYRUP & CAPSULES

 UREE SYRUP  U TONE SYRUP & CAPSULES 12

12

SALES & MARKETING STRATEGY ADOPTED BY SPAN LABORATORIES

In theory, the purpose of selling is to help a customer realize his or her goals in an economic fashion. However, in reality this is not always the case. Customers can be influenced to purchase a product or service that initially was not of interest to them. Some salespeople are trained in the art of selling customers things they don't need.

Take for example the purchasing of a car: a consumer may have a set of cars in mind (called an evoked set) that she feels match her needs, wants and budget. She may seek the advice of a salesperson given that a salesperson can help her realize the right car given those criteria.

This can be a socially useful function; salespeople have specialized knowledge of products that can help consumers make an informed decision. However, a salesperson may also talk a consumer into purchasing a more expensive or perhaps larger car then she needs or can afford. In this context, the salesperson may have usefully helped the customer re-evaluate her needs, thereby establishing a new set of appropriate choices among which included the newer or large car. This again would be a helpful and useful service provided by the salesperson. However, it is sometimes the case that customers purchase a product or service that was not initially intended and remains an inappropriate purchase after the fact. On the other hand, the consumer in this scenario can be held partially responsible for the inappropriate purchase; indeed, "A fool and his money are soon parted."

This dysfunctional behavior is encouraged by:

Incentives from the manufactures of products or the companies of service providers to salespeople to sell their products where other similar products offered by competitors are offered.

The incentive to sell a customer a product that is in need of being cleared out, despite the fact that a customer may be better to wait for the new product.

Incentives of salespeople to increase their total number of sales especially where retailers keep track of sales or offer commission-based salaries.

13

DISTIRTUBTION CHANNEL OF SPAN LABORATORIES

Manufacturing Plant

CHANNEL OF SPAN LABORATORIES Manufacturing Plant Clearing and forwarding agent (different regions) Stockist A
CHANNEL OF SPAN LABORATORIES Manufacturing Plant Clearing and forwarding agent (different regions) Stockist A
CHANNEL OF SPAN LABORATORIES Manufacturing Plant Clearing and forwarding agent (different regions) Stockist A

Clearing and forwarding agent (different regions)

Plant Clearing and forwarding agent (different regions) Stockist A Stockist B Stockist C Retailers Retailers

Stockist A

Clearing and forwarding agent (different regions) Stockist A Stockist B Stockist C Retailers Retailers Retailers
Clearing and forwarding agent (different regions) Stockist A Stockist B Stockist C Retailers Retailers Retailers

Stockist B

forwarding agent (different regions) Stockist A Stockist B Stockist C Retailers Retailers Retailers Retailers
forwarding agent (different regions) Stockist A Stockist B Stockist C Retailers Retailers Retailers Retailers

Stockist C

agent (different regions) Stockist A Stockist B Stockist C Retailers Retailers Retailers Retailers Retailers
Retailers Retailers Retailers Retailers Retailers Retailers C O N S U M E R S
Retailers
Retailers
Retailers
Retailers
Retailers
Retailers
C
O
N
S
U
M
E
R
S

The above diagram it shows channel of distribution of SPAN LABS Products, here first the products are manufactured and from Manufacturing plants the packed goods are supplied to Clearing And Forwarding Agents(C&FA) and from here the goods are then further supplied

14

to number of Stockiest or Distributors, from here goods reaches to large number of Retailers and it is the duty of Stockiest to take orders from retailers and then supply the goods to them, this work is generally done by stockiest salesman through ready stock or by taking orders first and then placing the order. From here the goods finally reaches to Customers. Customer purchases the product from retailers.

This was the basic Channel of Distribution used by SPAN LABS Products, now I will throw light on each channel of distribution of SPAN LABS Products.

SUPPLY CHAIN MANAGEMENT:

Supply chain management starts before physical distribution: it involves procuring the right inputs (raw materials, components and capital equipment), converting them into finished products and dispatching them to the final destinations. The supply chain perspective can help identify superior suppliers and distributors and help them improve productivity, which ultimately brings down the company‘s costs.

A broader view sees a company at the center of a value network that includes its

suppliers, its immediate customers and their end customers. The value network includes valued relations with others such as university researchers, government approval agencies

and so on.

PROCUREMENT & TRANSPORT

Getting the raw material and packaging material requirement from the production unit

in charge.

Constant updates on the procurement of materials and transport details.

Production details and ingredient content information from the different personnel and coordinating this activity.

PACKAGING

Approval and coordination of the supply of packaging material to the production unit.

15

CLEARING AND FORWARDING AGENTS (C&FA)

From manufacturing plant the stock is transported or supplied to clearing and forwarding agents.

Clearing and Forwarding Agents is a third party and SPAN LABS Products gives contract to them, so company has nothing to do in building the relationship with them.

Here C&FA keep or stock the goods with them.

STOCKIEST OR DISTRIBUTORS

Stockiest store the products in their godowns, C&FA supplies the goods to them as per their order.

Stockiest has some sales men working under him, they are known as stockiest sales man. Their work is to place the products in the market and take order from retailers and then supply goods to them.

Sales man either take ready stock with them or they first take orders and then supply goods later on.

There is a beat which is a schedule route of sales man, means sales man has to daily cover the route as mention in the beat.

Merchandising, making products visible, pasting posters, putting banners, and seeing that goods are properly placed in the retail outlets is also the duty of stockiest sales man.

Companies‘ sales officer keeps a check on the stockiest and monthly report is also prepared which is further analyzed by ASM & ZSM.

RETAILERS

Retailers are backbone of the company as they are the one who can take the product on new heights or can bring it down to toes.

Stockiest supplies goods to retailers and tries Persuading retailers to give the brand special displays (using merchandising tools) to get affective brand presence, and arranging it in more noticeable manner.

16

Classification of outlets in different type of markets is different according to their sales volume.

17

AYURVEDA An Overview

Ayurveda is an ancient science of life, a traditional and the oldest and most holistic medical system available on the planet today. Its major premise involves the symbiosis of mind, body and spirit. Any imbalance in this synthesis results in physical ailments. This ancient Indian medicine seeks to reestablish the harmony between the mind, body and its environment. It was placed in written form over 5,000 years ago in India, and was said to be a world medicine dealing with both body and the spirit. Before the advent of writing, the ancient wisdom of this healing system was a part of the spiritual oral tradition of the Vedic tradition. This has been handed down to us by means of ancient venerable scripts as palm leaf books, leather leaves, etc. The oldest works in Ayurveda still available are the Charaka Samhita, Sushuta Samhita and Ashtanga Samgraha, among others.

"Ayu" means life and "Veda" means knowledge from the Vedic texts. This holistic science is the knowledge of complete balance of the Body, Mind and Spirit, including emotions and psychology, on all levels. Ayurveda includes in its consideration, longevity, rejuvenation and self-realization therapies through herbs, diet, exercise, yoga, aromas, tantras, mantras, and meditation.

It is said to have originated from Lord Brahma (Creator of the Universe, according to Indian mythology) and descended to the earth through various generations of gods and saints. The sage-physician-surgeons of the time were the same sages or seers, deeply devoted holy people, who saw health as an integral part of spiritual life. It is said that they received their training of Ayurveda through direct cognition during meditation. In other words, the knowledge of the use of various methods of healing, prevention, longevity and surgery came through Divine revelation (Cosmic Intelligence); there were no guessing or testing and harming animals. These revelations were transcribed from the oral tradition into book form, interspersed with the other aspects of life and spirituality.

The original ancient Ayurvedic scholars also comprehended a true method to study and fully understand Ayurveda. The Vedic Way to study and understand Ayurveda is the same Vedic Way one takes to study life itself.

18

The Vedas describe life as a series of experiences. Each experience can be seen as the fundamental unit of life. It may be overwhelming to try to understand life as a whole, but by understanding each experience, one can, therefore, understand life itself.

Every experience has three components: (1) an experiencer, (2) an object of experience, and (3) the experience itself.

The experiencer is you.

The object of the experience can be any stimulus or environmental influence, e.g. a sight, a sound, a thought, a material object, a circumstance, a situation, a person, etc.

The experience is the interaction between the experiencer and the object.

An experience takes place only with all three components.

The Vedic approach to understand each experience is threefold:

Sravanam (receiving Truth),

Mananam (contemplating Truth), and

Nidhidhyasanam (absorbing Truth)

These are the three aspects of the study and understanding Ayurveda. It is from this approach

that Ayurveda derives its name, for it is a true path for learning the meaning of life. Every

popular book written by the burgeoning numbers of uninitiated Ayurvedic ‗authors‘ translates

the term Ayurveda as ―knowledge or science (ved) of life (ayu).‖ However it is clear that

very few individuals realize the profound meaning of this appellation.

Sravanam can be achieved through exposure to the primary Ayurvedic scriptures or other

numinous literature and through the teachings of knowledgeable professors.

Mananam can be individual or through discussion and debate with other students of

Ayurveda.

19

Nidhidhyasanam can be approached through a specific form meditation, sometimes referred

to as reflection, which allows the truthful understanding to become absorbed Only by

absorbing Truth, can one truly understand these principles and put them to use.

It is not necessary to approach sravanam, mananam, and nidhidhyasanam in a serial order

(i.e. one after the other), but rather one can incorporate different levels of each throughout

one's study of Ayurveda. We can constantly aspire to incorporate all three in our studies and

in our lives and perhaps glimpse the same truths as did the ancient Vedic seers.

Ayurveda is a holistic system of healing which evolved among the Brahmin sages of ancient

India some 3000-5000 years ago. There are several aspects of this system of medicine which

distinguish it from other approaches to health care:

1. It focuses on establishing and maintaining balance of the life energies within us, rather than focusing on individual symptoms.

2. It recognizes the unique constitutional differences of all individuals and therefore recommends different regimens for different types of people. Although two people may appear to have the same outward symptoms, their energetic constitutions may be very different and therefore call for very different remedies.

3. Ayurveda is a complete medical system which recognizes that ultimately all intelligence and wisdom flows from one Absolute source (Paramatman). Health manifests by the grace of the Absolute acting through the laws of Nature (Prakriti). Ayurveda assists Nature by promoting harmony between the individual and Nature by living a life of balance according to her laws.

4. Ayurveda describes three fundamental universal energies which regulate all natural processes on both the macrocosmic and microcosmic levels. That is, the same energies which produce effects in the various galaxies and star systems are operating at the level of the human physiology--in your own physiology. These three universal energies are known as the Tridosha.

5. Finally, the ancient Ayurvedic physicians realized the need for preserving the alliance of the mind and body and offers mankind tools for remembering and nurturing the

20

subtler aspects of our humanity. Ayurveda seeks to heal the fragmentation and disorder of the mind-body complex and restore wholeness and harmony to all people.

HISTORY OF AYURVEDA

To see the world from which Ayurveda developed it is necessary to go back 4000 years. Ayurveda had not yet been established. People lived close to the cycles of nature in a thriving agrarian society on the banks of the River Indus, dependent on the abundance of the harvest and the bounty of water for its survival. It was also a world in which the people were subjected to the full force of the power of nature; torrents of rain and the fierce heat of the sun, as well as the reassurance of spring returning and the joy of reaping a mature harvest. Subservience to the power that controls these natural extremes was at the centre of everyday life in a religious world full of rituals. Regular fire sacrifices were carried out to supplicate the deities upon whose favour the world depended. Ritual performance was as central to maintaining health as eating enough food; both were needed to live and flourish. To treat disease, herbs and potions were used alongside the incantations of the priests. In fact the priests were both doctors and religious specialists. Disease spread fast in these warm and humid climes. Fear of illness and of the death of loved ones was an everyday reality. According to their belief system disease could be imposed from the spiritual world, from an accident, or from the natural world. Here is the world in which the eternal tradition and the empirical experience of everyday life could meet and intermingle. It was out of such a cultural context that Ayurveda developed. Here was a fast-changing society that was exploring its ideals of religion, royalty, leadership, law, medicine, and family. Philosophical insight expanded as agrarian culture flourished.

21

22

22

SCOPE OF AYURVEDIC MEDICINE

Classically, Ayurvedic Medicine was conceptualized and practiced as eight major clinical

subspecialties of medicine in addition to numerous adjunctive specialties. The eight major

subspecialties continue to be taught today and they include:

1. Internal Medicine (Kayachikitsa)

2. General Surgery (Shalya Tantra)

3. Otorhinolaryngology (Shalakya)

4. Pediatrics and Obstetric/Gynecology (Kaumarabhrtya)

5. Psychiatry (Bhutavidya)

6. Toxicology (Agada Tantra)

7. Nutrition, Detoxification and Rejuvenation (Rasayana Tantra)

8. Fertility and Virility (Vajikarana)

For every disease, there is information about: definition, etiology, prodrome, clinical symptoms, pathophysiology, and prognosis, principles of treatment, medicines, diet, lifestyle recommendations, and even etymology. This approach is strikingly similar to that of modern medicine and even more comprehensive. Over the last century, Ayurvedic Medicine has experienced a rebirth and has continued to evolve its holistic approach to health in accordance with modern needs and scientific advances of the day. Today, modern Ayurveda also includes:

1. Kulam Svastyam Kutumbakam: Principles of Preventative Healthcare For the Entire Family

2. Sangakara Chikitsa: Treatment of Addictions. Includes strategies for defeating addictions to alcohol, tobacco, sexual behavior, and food.

3. Panchakarma Chikitsa: Purification and Rejuvenation Treatments. Prescribed with respect to one's individual nature, work, social circumstance, age, and season.

4. Sthaulya Chikitsa: The Ayurvedic Approach To Diet and Weight Loss. This discipline covers practical and effective approaches to maintain a healthy weight through constitutionally-determined diet, exercise, herbs, spices, teas, breathing, and psychological aids.

23

5. Vatavyadhi Chikitsa: Specific treatment plans for the diseases of Vata origin which affect the musculoskeletal system and nervous system (joints, bones, muscles, nerves) Examples include but are not limited to: osteoarthritis, osteoporosis, osteopenia, multiple sclerosis, spondylosis, sciatica, fibromyalgia, and chronic fatigue syndrome.

6. Svabhaavoparamavaada: Promotion of self-healing and resistance to disease (i.e. immunity) as per your age, sex, occupation, nature, daily routine, medical history, mental status, season, and region.

7. Vajikarana: Specific remedies for Male infertility and impotence as well as Female infertility.

8. Saundarya Sadhana: Beauty and cosmetic treatments for men and women, including skin, hair, eyes, posture, body odor, and general appearance.

BASIC PRINCIPLES OF AYURVEDA

The Three Doshas The Five Elements Everything in the universe is made up of combinations of the Five Elements (Pancha Mahabhutas). This includes the human being which also acquires a soul or spirit. These five elements are known as:

Space or Akasha

Air or Vayu

Fire or Tejas

Water or Apa

Earth or Prithvi

These five elements, it should be understood, derive from and are expressions of an unmanifest and undifferentiated Creative Principle, which is one. These five elements are to be understood in a material sense as well as a subtle sense. By earth we are to understand not only the terrain of our planet or the iron in our red blood cells and spleen, but also the quality of steadfastness of mind, strength of one‘s moral fiber, one‘s slow and quiet undeterred advancement towards a goal, and the resistance to the manifestations of others. By water we mean to imply the cohesive aspects of reality which flows into and holds things together, perfectly and simply witnessed in the ubiquitous H 20 molecule. And the other elements too

24

were intended by the ancient vaidyas (physicians) to communicate the essential universal principle inherent in a particular element. By fire we mean the universal force in nature that produces heat and radiates light; it is our passion to pursue despite obstacles and delays; it is what burns away the cloak of ignorance (avidya) and allows the Truth to shine with brilliance. Fire removes doubt from the mother-substance of human heart and replaces it with joy. Air is that transparent, rarefied, kinetic force which sets the universe in motion; it moves the blood through the vessels, wastes from the body, thoughts through the mind; it moves the birds to warmer climates in winter, it moves the planets around their suns. Space is the subtlest of all elements which is everywhere and touches everything; in the mind it is the vessel which receives all impressions, in the heart space accepts love; space is receptivity and non-resistance to what is true.

Thus these Five Subtle Elements (Pancha Mahabhutas) form the basis for all things found in the material creation, from a grain of sand to the complex physiology of every human being. Balancing these elements in just the right way for each unique individual is the key to maintaining health and treating disease should it arise, whether it is physical, mental, or spiritual.

The Tridosha

The five elements can be seen to exist in the material universe at all scales both organic and inorganic, from peas to planets. When they enter into the biology of a living organism, man for example, they acquire a biological form. This means that the five elements are coded into three biological forces which govern all life processes. These three forces are known as the

The tridosha regulates every physiological and

psychological process in the living organism. The interplay among them determines the qualities and conditions of the individual. A harmonious state of the three doshas creates balance and health; an imbalance, which might be an excess (vrddhi) or deficiency (ksaya), manifests as a sign or symptom of disease.

three doshas, or simply thetridosha

The three doshas are known as Vata, Pitta, and Kapha.

25

You can think of these three doshas as fundamental biological energies which regulate all the life processes of an individual. And as we will discuss later, although all individuals are made up of these same three energies, we all have them in unique proportions. The doshas obtain their qualities by virtue of their elemental composition as we can see in the simple diagram below. Each of the three doshas is composed of two elements as shown here:

Elements Composing the Tridosha

Vata

Space (Akasha)

Air (Vayu)

Pitta

Fire (Tejas)

Water (Apa)

Kapha

Water (Apa)

Earth (Prithvi)

Thus, Vata is composed of space and air, Pitta of fire and water, and Kapha of water and

earth

qualities of fire and water; Kapha dosha the stability and solidity of water and earth. Interestingly, the Sanskrit entomology of the word dosha gives it the meaning of "blemish, that which darkens". This alerts us to the fact that when in balance these force are life- supporting but when imbalanced they are the agents of disease and misery.

Vata dosha has the mobility and quickness of space and air; Pitta dosha the metabolic

26

HOW THE AYURVEDIC TRADITION BECAME A SYSTEM OF EMPIRICAL MEDICINE

Any history of ayurvedic development requires discussing two different perspectives; a linear religio-historical approach and a circular organic expansion. The first perceives Ayurveda as a timeless system of medicine where its knowledge is perfect and divinely inspired; the second view is that ayurvedic medical knowledge has developed out of ritualistic healing into an empirical medicine system that is grounded in clinical exper ience. The introductory verses of ayurvedic texts reflect the perspective that Ayurveda is an eternal revelation. They all start with a mythological account of the gods passing ayurvedic knowledge down to humans. This divine stamp is a well-known Indian method of authenticating a text and making it orthodox (Wujastyk 2003). It is a way of bringing formerly untraditional and perhaps unaccepted ideas into mainstream culture. Much of the secondary and modern ayurvedic literature also implies a consistent tradition that is divinely inspired and eternal. But, as you untangle the web of influences that have affected Ayurveda the evidence clearly reveals an expanding tradition that has accumulated knowledge over time and through experience. This latter organic perspective, first introduced by Jan Meulenbeld, holds that Ayurveda is a science of unfolding truth and as a path of discovery it has not and will not remain static. These developments are not necessarily mutually exclusive, but it is useful to understand the roots of different ayurvedic traits. The concept of a timeless tradition has great appeal, for the insights of Ayurveda are incredible and they do appear to be divinely inspired. How else have we learnt about the properties of so many herbs and minerals? How was it discovered, for example, that brahmi (Bacopa monniera) is so effective at improving the intellect and guggulu (Commiphora mukul) so useful at reducing tumours? How did the pioneers of Ayurveda learn to diagnose illness with only the five senses at their disposal? Having said this, the idea of human knowledge growing through experience, logic and insight has great value. Human development is firmly grounded in endeavour. For Indian minds this duality causes no conflict as Ayurveda can be two things at the same time: both divinely inspired and open to human adaptation. This is a powerful medium for expression of the truth as it is both reductionist and holistic.

Taking the first paradigm, while there is nothing inherently wrong with the claim of eternal divine origins there are some potential problems with this perspective. It could potentially

27

stifle new ideas within Ayurveda as, in order to gain validity, there is a tendency for clinical experience to be referenced back to a divine eternal source. Humble that this approach is, new ideas are not easily propagated. There is an element of this attitude displayed by the core theoretical ground of Ayurveda, having remained very similar over the last 2000 years. The relative lack of modern innovative ayurvedic literature generating improved methods of treatment, in comparison to Chinese and Western herbal medicine, is perhaps partially a result of this. It may be that the inherent theories of Ayurveda are already complete, but effective clinical insights are always of benefit as new diseases and cultural habits arise.The insistence on divine origins has stagnated this process of valuing both clinical experience and theory. It is not therefore surprising that as Ayurveda has been under continual threat from certain Moghul, British and, currently, allopathic forces in the last 400 years, it has in some quarters been necessary to fall back on its ancient roots in order to validate and justify its presence. This has protected but also weakened Ayurveda. Its strength is really in its present clinical excellence and the ayurvedic community should be harnessing powerful social forces and speaking with confidence about its ability to help our society. However, this is made difficult when Ayurveda is presently only recognised as an adjunctive medical system, where ayurvedic doctors can only hold the position of a third medical officer at primary health centres in India, and complementary medicine the world over holds a similarly lowly position in the medical hierarchy. As a literature base of over 2000 years, hundreds of thousands of expert physicians, millions of healed patients and numerous positive clinical trials attest, ayurvedic treatment works and practitioners and professional registers should promote this, researchers should publish clinical data and governments should support it enthusiastically. Although Ayurveda has its roots in the past, its practitioners must embrace the present. Ayurveda and ayurvedic physicians deserve greater recognition than they receive today. Another, and potentially more serious, problem of relying on a doctrine that holds its origins as divinely and infallibly inspired, is that it can and has resulted in right -wing fundamental political groups utilising it to their own ends. This is clearly the case in India today with the current rise in popularity of right-wing fundamental Hindu groups, and shows how the struggle for political supremacy can infect religion (and vice versa).

This insistence of the divine origins of Ayurveda may unwittingly reinforce this political doctrine if it continues to ignore modern Indological historical knowledge. By this, it is referred to certain quarters of the academic community promoting this ideology as though

28

Vedic knowledge has remained eternally and statically predominant in all aspects of Indian culture for all time. The point is that while religion, medicine and politics are interrelated, the potential repercussions of an ideology must be considered; in this case, pandering to extreme political causes that oppose the central tenet of Ayurvedacaring for all humanity.

The second paradigm, the scientific dependence on empirical evidence, can also be taken too far to the extreme, with similar detriment. This has occurred within the modern medical paradigm of ‗evidencebased medicine‘ requiring ethically dubious double-blind clinical trials and animal experiments with a heavy dependence on single active ingredients, synthesised medicines, separate chemical pathways and a reductionist methodology that has lost the holistic view.

Holding onto the primacy of either of these two paradigms means that the complete picture is missed. As we shall see, Ayurveda can offer a balance to these extremes as it contains both paradigms within it. I think this inner debate between tradition and progression is mirrored in our everyday lives and specifically experienced when using natural medicine. The question is how to respect tradition while integrating personal experience. Internally it is a case of communication between heart and head where intuition and intellect are both valid. As we shall see, intuition and intellect are both essential for medicine to be, as Ayurveda is, truly holistic.

29

QUALITY CONTROL OF AYURVEDIC DRUGS

Quality control for efficacy and safety of ayurvedic products is of paramount importance. Quality can be defined as the status of a drug that is determined by identity, purity, content, and other chemical, physical, or biological properties, or by the manufacturing processes. Quality control is a term that refers to processes involved in maintaining the quality and validity of a manufactured product. For the quality control of a traditional medicine, the traditional methods are procured and studied, and documents and the traditional information about the identity and quality assessment are interpreted in terms of modern assessment. In general, all medicines, whether they are of synthetic or of plant origin, should fulfill the basic requirements of being efficacious and safe, and this can be achieved by suitable clinical trials. This applies both to the multinational pharmaceutical company conducting a multi-center, double-blind placebo-controlled study with a herbal extract, and to the health practitioner in a rural village who applies a locally produced ayurvedic mixture. Natural products in medicine constitute a vast array of ―raw materials,‖ making clear definitions important. Quality criteria are based on clear scientific definitions of the raw material. The term ―ayurvedic drugs‖ denotes plants or plant parts that have been converted into phytopharmaceuticals by means of simple processes involving harvesting, drying, and storage. Hence they are capable of variation. This variability is also caused by differences in growth, geographical location, and time of harvesting. A practical addition to the definition is also to include other crude products derived from plants, which no longer show any organic structure, such as essential oils, fatty oils, resins, and gums. Derived or isolated compounds in the processed state such as extracts or even isolated purified compounds (e.g. strychnine from Strychnos nux-vomica) or mixtures of compounds (e.g. abrin from Abrus precatorius) are, as a rule, not included in the definition. Combinations with chemically defined active substances or isolated constituents, and homeopathic preparations which frequently contain plants, are not regarded as ayurvedic medicines. Their production is already based on adequate quality control of the respective starting materials. The following paragraphs will focus on quality control of ayurvedic drugs in compliance with the above definition.

In general, quality control is based on three important pharmacopeial definitions:

Identity: Is the herb the one it should be?

30

Purity: Are there contaminants, e.g., in the form of other herbs which should not be there?

Content or assay: Is the content of active constituents within the defined limits?

It is obvious that the content is the most difficult one to assess, since in mo st ayurvedic drugs the active constituents are unknown. Sometimes markers can be used which are, by definition, chemically defined constituents that are of interest for control purposes, independent of whether they have any therapeutic activity or not. To prove identity and purity, criteria such as type of preparation sensory properties, physical constants, adulteration, contaminants, moisture, ash content and solvent residues have to be checked. The correct identity of the crude herbal material, or the botanical quality, is of prime importance in establishing the quality control of ayurvedic drugs. Identity can be achieved by macro- and microscopical examinations. Voucher specimens are reliable reference sources. Outbreaks of diseases among plants may result in changes to the physical appearance of the plant and lead to incorrect identification. At times an incorrect botanical quality with respect to the labeling can be a problem. For example, in the 1990s, a South American product labeled as ―Paraguay Tea‖ was associated with an outbreak of ant cholinergic poisoning in New York. Subsequent chemical analysis revealed the presence of a class of constituents that was different from the metabolites normally found in the plant from which Paraguay tea is made.

Purity is closely linked with the safe use of drugs and deals with factors such ash values, contaminants (e.g. foreign matter in the form of other herbs), and heavy metals. However, due to the application of improved analytical methods, modern purity evaluation also includes microbial contamination, aflatoxins, radioactivity, and pesticide residues. Analytical methods such as photometric analysis, thin layer chromatography (TLC), high performance liquid chromatography (HPLC), and gas chromatography (GC) can be employed in order to establish the constant composition of herbal preparations. Depending upon whether the active principles of the preparation are known or unknown, different concepts such as ―normalization versus standardization‖ have to be applied in order to establish relevant criteria for uniformity.

Content or assay is the most difficult area of quality control to perform, since in most ayurvedic drugs the active constituents are not known. Sometimes markers can be used. In all

31

other cases, where no active constituent or marker can be defined for the ayurvedic drug, the percentage extractable matter with a solvent may be used as a form of assay, an approach often seen in pharmacopeias. The choice of the extracting solvent depends on the nature of the compounds involved, and might be deduced from the traditional uses. For example, when an ayurvedic drug is used to make a tea, the hot water extractable matter, expressed as milligrams per gram of air-dried material, may serve this purpose. A special form of assay is the determination of essential oils by steam distillation. When the active constituents (e.g. sennosides in Senna) or markers (e.g. alkydamides in Echinacea) are known, a vast array of modern chemical analytical methods such as ultraviolet/visible spectroscopy (UV/VIS), TLC, HPLC, GC, mass spectrometry (MS), or a combination of GC and MS (GC/MS), can be employed. Several problems not applicable to synthetic drugs influence the quality of ayurvedic drugs:

Ayurvedic drugs are usually mixtures of many constituents.

The active principle(s) is (are), in most cases unknown.

Selective analytical methods or reference compounds may not be available commercially.

Plant materials are chemically and naturally variable.

Chemo-varieties and chemo cultivars exist.

The source and quality of the raw material are variable.

The methods of harvesting, drying, storage, transportation, and processing (for example, mode of extraction and polarity of the extracting solvent, instability of constituents, etc.) have an effect. Strict guidelines have to be followed for the successful production of a quality ayurvedic drug. Among them are proper botanical identification, phytochemical screening, and standardization. Quality control and the standardization of ayurvedic medicines involves several steps. The source and quality of raw materials, good agricultural practices and manufacturing processes are certainly essential steps for the quality control of ayurvedic medicines and play a pivotal role in guaranteeing the quality and stability of ayurvedic preparations.

The quality of a plant product is determined by the prevailing conditions during growth, and accepted Good Agricultural Practices (GAP) can control this. These include seed selection, growth conditions, and use of fertilizers, harvesting, drying and storage. In fact, GAP

32

procedures are, and will be, an integral part of quality control. Factors such as the use of fresh plants, age and part of plant collected, period, time and method of collection, temperature of processing, exposure to light, availability of water, nutrients, drying, packing, transportation of raw material and storage, can greatly affect the quality, and hence the therapeutic value of ayurvedic medicines.

Apart from these criteria, factors such as the method of extraction, contamination with microorganisms, heavy metals, and pesticides can alter the quality, safety, and efficacy of ayurvedic drugs. Using cultivated plants under controlled conditions instead of those collected from the wild can minimize most of these factors.

Sometimes the active principles are destroyed by enzymic processes that continue for long periods from collection to marketing, resulting in a variation of composition. Thus proper standardization and quality control of both the raw material and the ayurvedic preparations should be conducted. Standardization involves adjusting the ayurvedic drug preparation to a defined content of a constituent or a group of substances with known therapeutic activity by adding excipients or by mixing ayurvedic drugs or herbal drug preparations. Botanical extracts made directly from crude plant material show substantial variation in composition, quality, and therapeutic effects. Standardized extracts are high-quality extracts containing consistent levels of specified compounds, and they are subjected to rigorous quality controls during all phases of the growing, harvesting, and manufacturing processes. No regulatory definition exists for standardization of dietary supplements. As a result, the term ―standardization‖ may mean many different things. Some manufacturers use the term standardization incorrectly to refer to uniform manufacturing practices; following a recipe is not sufficient for a product to be called standardized. Therefore, the presence of the word ―standardized‖ on a supplement label does not necessarily indicate product quality. When the active principles are unknown, marker substance(s) should be established for analytical purposes and standardization. Marker substances are chemically defined constituents of a ayurvedic drug that are important for the quality of the finished product. Ideally, the chemical markers chosen would also be the compounds that are responsible for the botanical‘s effects in the body.

33

There are two types of standardization. In the first category, ―true‖ standardization, a definite phytochemical or group of constituents is known to have activity. Ginkgo with its 26% ginkgo flavones and 6% terpenes is a classic example. These products are highly concentrated and no longer represent the whole herb, and are now considered as phytopharmaceuticals. In many cases they are vastly more effective than the whole herb. However the process may result in the loss of efficacy and the potential for adverse effects and herbdrug interactions may increase. The other type of standardization is based on manufacturers guaranteeing the presence of a certain percentage of marker compounds; these are not indicators of therapeutic activity or quality of the herb. In the case of ayurvedic drug preparations, the production and primary processing of the medicinal plant or herbal drug has

a direct influence on the quality of the active pharmaceutical ingredients (APIs). Due to the

inherent complexity of naturally growing medicinal plants and the limited availability of simple analytical techniques to identify and characterize the active constituents solely by chemical or biological means, there is a need for an adequate quality assurance system. This assurance is also required during cultivation, harvesting, primary processing, handling, storage, packaging, and distribution. Deterioration and contamination through adulteration, especially microbial contamination, can occur at any one of these stages. It is extremely important to establish good agricultural, harvesting, and manufacturing practices for ayurvedic starting materials in order to minimize these undesirable factors.

In this regard producers, processors, and traders of medicinal plants or ayurvedic drugs have an obligation and a role to play. The manufacturers and suppliers of ayurvedic products should adhere to quality control standards and good manufacturing practices. Currently, only

a few manufacturers adhere to complete quality control and good manufacturing procedures

including microscopic, physical, chemical, and biological analysis. Organizations such as Department of Ayush help safeguard our health by carrying out premarket reviews of all drugs before they are authorized for sale. The products available in the market are analyzed

regularly to ensure that they are free of unsafe ingredients and that the products actually contain the ingredients indicated on the labels.

The potency and quality of an individual ayurvedic product may be unclear because of lack of regulation. It is obvious that for a given plant product its quality will also be determined by the prevailing conditions during the growth cycle of the plant. Therefore, for cultivated plants

34

the GAP system has been introduced, under which each step, including seed selection, growing conditions, use of fertilizers, and optimization of harvest time, harvesting, and drying, has to adhere to a set of criteria. It is likely that GAP procedures will become an integral part of quality control in the near future.

Microscopic Evaluation

Quality control of ayurvedic drugs has traditionally been based on appearance and today microscopic evaluation is indispensable in the initial identification of herbs, as well as in identifying small fragments of crude or powdered herbs, and detection of foreign matter and adulterants. A primary visual evaluation, which seldom needs more than a simple magnifying lens, can be used to ensure that the plant is of the required species, and that the right part of the plant is being used. At other times, microscopic analysis is needed to determine the correct species and/or that the correct part of the species is present. For instance, pollen morphology may be used in the case of flowers to identify the species, and the presence of certain microscopic structures such as leaf stomata can be used to identify the plant part used. Although this may seem obvious, it is of prime importance, especially when different parts of the same plant are to be used for different treatments. Stinging nettle (Urtica urens) is a classic example where the aerial parts are used to treat rheumatism, while the roots are applied for benign prostate hyperplasia.

Determination of Foreign Matter

Ayurvedic drugs should be made from the stated part of the plant and be devoid of other parts of the same plant or other plants. They should be entirely free from moulds or insects, including excreta and visible contaminant such as sand and stones, poisonous and harmful foreign matter and chemical residues. Animal matters such as insects and ―invisible‖ microbial contaminants, which can produce toxins, are also among the potential contaminants of ayurvedic medicines. Macroscopic examination can easily be employed to determine the presence of foreign matter, although microscopy is indispensable in certain special cases (for example, starch deliberately added to ―dilute‖ the plant material). Furthermore, when foreign matter consists, for example, of a chemical residue, TLC is often needed to detect the contaminants.

35

Determination of Ash

To determine ash content the plant material is burnt and the residual ash is measured as total

and acid-insoluble ash. Total ash is the measure of the total amount of material left after burning and includes ash derived from the part of the plant itself and acid-insoluble ash. The latter is the residue obtained after boiling the total ash with dilute hydrochloric acid, and

burning the remaining insoluble matter. The second procedure measures the amount of silica present, especially in the form of sand and siliceous earth.

Determination of Heavy Metals

Contamination by toxic metals can either be accidental or intentional. Contamination by

heavy metals such as mercury, lead, copper, cadmium, and arsenic in ayurvedic remedies can

be attributed to many causes, including environmental pollution, and can pose clinically

relevant dangers for the health of the user and should therefore be limited. The potential

intake of the toxic metal can be estimated on the basis of the level of its presence in the product and the recommended or estimated dosage of the product. This potential exposure

can then be put into a toxicological perspective by comparison with the so-called Provisional

Tolerable Weekly Intake values (PTWI) for toxic metals, which have been established by the

Food and Agriculture Organization of the World Health Organization (FAO-WHO).

A simple, straightforward determination of heavy metals can be found in many

pharmacopeias and is based on color reactions with special reagents such as thioacetamide or diethyldithiocarbamate, and the amount present is estimated by comparison with a standard. Instrumental analyses have to be employed when the metals are present in trace quantities, in admixture, or when the analyses have to be quantitative. The main methods commonly used are atomic absorption spectrophotometry (AAS), inductively coupled plasma (ICP) and neutron activation analysis (NAA).

Determination of Microbial Contaminants and Aflatoxins

Medicinal plants may be associated with a broad variety of microbial contaminants, represented by bacteria, fungi, and viruses. Inevitably, this microbiological background

36

depends on several environmental factors and exerts an important impact on the overall quality of ayurvedic products and preparations. Risk assessment of the microbial load of medicinal plants has therefore become an important subject in the establishment of modern Hazard Analysis and Critical Control Point (HACCP) schemes.

Ayurvedic drugs normally carry a number of bacteria and molds, often originating in the soil. Poor methods of harvesting, cleaning, drying, handling, and storage may also cause additional contamination, as may be the case with Escherichia coli or Salmonella spp. While a large range of bacteria and fungi are from naturally occurring microflora, aerobic spore- forming bacteria frequently predominate.

Laboratory procedures investigating microbial contaminations are laid down in the well- known pharmacopeias, as well as in the WHO guidelines. Limit values can also be found in the sources mentioned. In general, a complete procedure consists of determining the total aerobic microbial count, the total fungal count, and the total Enterobacteriaceae count, together with tests for the presence of Escherichia coli, Staphylococcus aureus, Shigella, and Pseudomonas aeruginosa and Salmonella spp. The European Pharmacopoeia also specifies that E. coli and Salmonella spp. should be absent from ayurvedic preparations. However it is not always these two pathogenic bacteria that cause clinical problems. For example, a fatal case of listeriosis was caused by contamination of alfalfa tablets with the Grampositive bacillus Listeria monocytogenes.

Materials of vegetable origin tend to show much higher levels of microbial contamination than synthetic products and the requirements for microbial contamination in the Indian Pharmacopoeia allow higher levels of microbial contamination in ayurvedic remedies than in synthetic pharmaceuticals. The allowed contamination level may also depend on the method of processing of the drug. For example, higher contamination levels are permitted if the final ayurvedic preparation involves boiling with water.

The presence of fungi should be carefully investigated and/or monitored, since some common species produce toxins, especially aflotoxins. Aflatoxins in ayurvedic drugs can be dangerous to health even if they are absorbed in minute amounts. Aflatoxin-producing fungi sometimes build up during storage. Procedures for the determination of aflatoxin contamination in

37

ayurvedic drugs are published by the WHO. After a thorough clean-up procedure, TLC is used for confirmation. In addition to the risk of bacterial and viral contamination, ayurvedic remedies may also be contaminated with microbial toxins, and as such, bacterial endotoxins and mycotoxins, at times may also be an issue. There is evidence that medicinal plants from some countries may be contaminated with toxigenic fungi (Aspergillus, Fusarium). Certain plant constituents are susceptible to chemical transformation by contaminating microorganisms.

Withering leads to enhanced enzymic activity, transforming some the constituents to other metabolites not initially found in the herb. These newly formed constituent(s) along with the molds such as Penicillium nigricans and P. jensi may then have adverse effects.

Determination of Pesticide Residues

Even though there are no serious reports of toxicity due to the presence of pesticides and fumigants, it is important that herbs and ayurvedic products are free of these chemicals or at least are controlled for the absence of unsafe levels. ayurvedic drugs are liable to contain pesticide residues, which accumulate from agricultural practices, such as spraying, treatment of soils during cultivation, and administering of fumigants during storage. However, it may be desirable to test ayurvedic drugs for broad groups in general, rather than for individual pesticides. Many pesticides contain chlorine in the molecule, which, for example, can be measured by analysis of total organic chlorine. In an analogous way, insecticides containing phosphate can be detected by measuring total organic phosphorus. Samples of ayurvedic material are extracted by a standard procedure, impurities are removed by partition and/or adsorption, and individual pesticides are measured by GC, MS, or GC/MS. Some simple procedures have been published by the Department of Ayush and the Indian Pharamacopoeia has laid down general limits for pesticide residues in medicine.

Analytical Methods

Published monographs in a pharmacopeia are the most practical approach for quality control of ayurvedic drugs and there are many available. When pharmacopeial monographs are unavailable, development and validation of analytical procedures have to be carried out by the manufacturer. The best strategy is to follow closely the pharmacopeial definitions of

38

identity, purity, and content or assay. Valuable sources for general analytical procedures are included in the pharmacopeias, in guidelines published by the Department of Ayush . Additional information, especially on chromatographic and/or spectroscopic methods can be found in the general scientific literature. The plant or plant extract can be evaluated by various biological methods to determine pharmacological activity, potency and toxicity. A simple chromatographic technique such as TLC may provide valuable additional information to establish the identity of the plant material. This is especially important for those species that contain different active constituents.

Qualitative and quantitative information can be gathered concerning the presence or absence of metabolites or breakdown products. TLC fingerprinting is of key importance for ayurvedic drugs made up of essential oils, resins, and gums, which are complex mixtures of constituents that no longer have any organic structure. It is a powerful and relatively rapid solution to distinguish between chemical classes, where macroscopy and microscopy will fail.

Chromatograms of essential oils, for example, are widely published in the scientific literature, and can be of invaluable help in identification.

The instruments for UV-VIS determinations are easy to operate, and validation procedures are straightforward but at the same time precise. Although measurements are made rapidly, sample preparation can be time consuming and works well only for less complex samples, and those compounds with absorbance in the UV-VIS region.

HPLC is the preferred method for quantitative analysis of more complex mixtures. Though the separation of volatile components such as essential and fatty oils can be achieved with HPLC, it is best performed by GC or GC/MS.

The quantitative determination of constituents has been made easy by recent developments in analytical instrumentation. Recent advances in the isolation, purification, and structure elucidation of naturally occurring metabolites have made it possible to establish appropriate strategies for the determination and analysis of quality and the process of standardization of ayurvedic preparations. Classification of plants and organisms by their chemical constituents is referred to as chemotaxonomy. TLC, HPLC, GC, quantitative TLC (QTLC), and high-

39

performance TLC (HPTLC) can determine the homogeneity of a plant extract. Over- pressured layer chromatography (OPLC), infrared and UV-VIS spectrometry, MS, GC, liquid chromatography (LC) used alone, or in combinations such as GC/MS, LC/MS, and MS/MS, and nuclear magnetic resonance (NMR), electrophoretic techniques, especially by hyphenated chromatographies, are powerful tools, often used for standardization and to control the quality of both the raw material and the finished product. The results from these sophisticated techniques provide a chemical fingerprint as to the nature of chemicals or impurities present in the plant or extract.

Based on the concept of photoequivalence, the chromatographic fingerprints of ayurvedic medicines can be used to address the issue of quality control. Methods based on information theory, similarity estimation, chemical pattern recognition, spectral correlative chromatograms (SCC), multivariate resolution, the combination of chromatographic fingerprints and chemometric evaluation for evaluating fingerprints are all powerful tools for quality control of ayurvedic products.

Validation

The validation of ayurvedic products is a major public health concern both in developed and resource-poor countries, where fake businesses selling adulterated ayurvedic medicines are common. In this regard, there is no control by the government agencies, despite the existence of certain guidelines in some individual countries and those outlined by the WHO. If the ayurvedic products are marketed as therapeutic agents, and irrespective of whether the products really have any positive effects to cure and reduce the severity of the disease, it is necessary to ensure scientific validation and periodic monitoring of the quality and efficacy by drug control administrators.

It is feasible that the introduction of scientific validation would control the production of impure or adulterated ayurvedic products and would eventually ensure their rational use. This could also lead to the regulation of the industry so that only qualified physicians and health providers are allowed to prescribe the medication.

40

Several of the principal pharmacopeias contain monographs outlining standards for ayurvedic drugs. The major advantage of an official monograph published in a pharmacopeia is that standards are defined and available, and that the analytical procedures used are fully validated. This is of major importance, since validation can be a rather time-consuming process.

By definition, validation is the process of proving that an analytical method is acceptable for its intended purpose for pharmaceutical methods. Guidelines from the United States Pharmacopeia (USPC, 19942001), the International Conferenceon Harmonization (ICH), and the US Food and Drug Administration (FDA) provide a framework for performing such validations. In general, validation investigations must include studies on specificity, linearity, accuracy, precision, range, detection, and quantitative limits, depending on whether the analytical method used is qualitative or quantitative. Also of utmost importance is the availability of standards. For macroscopic and microscopic procedures in general this means that reliable reference samples of the plant must be available. A defined botanical source (e.g. voucher specimens) will normally solve this problem. Standards for chromatographic procedures are less easy to obtain. Characteristic plant constituents, either active or markers, are seldom available commercially. Sometimes an LC/MS approach can be referred to as a mode of characterization. Going one step further, after isolation of such a compound, elucidations to prove its definite structure will not be easy. The method often employed is to use readily available compounds that behave similarly in the chosen chromatographic systems, and to calculate retention values and/or times towards these compounds as a standard.

Qualitative chemical examination is designed to detect and isolate the active ingredient(s). TLC and HPLC are the main analytical techniques commonly used. In cases when active ingredients are not known or too complex, the quality of plant extracts can be assessed by a ―fingerprint‖ chromatogram.

41

Ayurvedic Supplements

A botanical is a plant or part of a plant valued for its medicinal or therapeutic properties, flavor, and/or scent. Herbs are subsets of botanicals. To be classified as a dietary supplement, a botanical must meet the following criteria:

1. It is intended to supplement the diet.

2. It contains one or more dietary ingredients (including amino acids, vitamins, minerals, herbs, or other botanicals, etc.).

3. It is intended to be taken orally as a pill, capsule, tablet, or liquid.

4. It is labeled as being a dietary supplement.

An ayurvedic supplement labeled ―Natural‖ does not mean it is safe or without any harmful effects. Ayurvedic products can act the same way as drugs. Their safety depends on factors such as their chemical make-up, how they work in the body, method of preparation, and dosage. In the US, the FDA regulates herbal and other dietary supplements. This means that they do not have to meet the same standards as drugs and over-the-counter medications, they are not required to be standardized, and no legal or regulatory definitions exist for standardization. As a result, manufacturers are not required to demonstrate the safety and effectiveness of their products before they reach the market. In addition, they do not have to adhere to any of the quality control measures applicable to drugs; hence the composition may vary greatly from one batch to another. The use of some ayurvedic supplements has been reported to be associated with ailments such as oral manifestations, including swelling, irritation, and bleeding of the tongue. These potential effects of ayurvedic supplements, in conjunction with factors related to regulation restrictions, suggest that the use of these products may be associated with various adverse reactions that can affect health. The active ingredient (s) in many ayurvedic supplements is not known and some have been found to be contaminated with metals, unlabeled prescription drugs, and microorganisms. Under its current regulatory authority, the Department of Ayush can remove a ayurvedic supplement from the market only after it has been shown to be unsafe. There has been an increase in the number of Internet websites that sell and promote ayurvedic supplements. Unfortunately, some of them make inaccurate claims and statements regarding their products and claim unsubstantiated effects in curing disease and disease conditions.

42

Adulteration of Ayurvedic drugs

Direct or intentional adulteration of drugs usually includes practices in which a ayurvedic drug is substituted partially or fully with other inferior products. Due to morphological resemblance to the authentic herb, many different inferior commercial varieties are used as adulterants. These may or may not have any chemical or therapeutic potential. Substitution by ―exhausted‖ drugs entails adulteration of the plant material with the same plant material devoid of the active constituents. This practice is most common in the case of volatile oil-containing materials, where the dried exhausted material resembles the original drug but is free of the essential oils. Foreign matter such as other parts of the same plant with no active ingredients, sand and stones, manufactured artifacts, and synthetic inferior principles are used as substitutes. The practice of intentional adulteration is mainly encouraged by traders who are reluctant to pay premium prices for herbs of superior quality, and hence are inclined to purchase only the cheaper products. This encourages producers and traders to sell herbs of inferior qualit y. Rarity of an herbal product is another factor that influences adulteration. Sometimes sale of inferior products may be unintentional.

In the absence of proper means of evaluation, an authentic drug partially or fully devoid of the active ingredients may enter the market. Factors such as geographical sources, growing conditions, processing, and storage are all factors that influence the quality of the drug. Deterioration may contribute to indirect adulteration, and crude drugs are often prone to deterioration, especially during storage, leading to the loss of the active ingredients, production of metabolites with no activity and, in extreme cases, the production of toxic metabolites. Physical factors such as air (oxygen), humidity, light, and temperature can bring about deterioration directly or indirectly. These factors, alone or in combination, can lead to the development of organisms such as molds, mites, and bacteria. Oxidation of the constituents of a drug can be brought about by oxygen in the air, causing some products, such as essential oils, to resinify or to become rancid. Moisture or humidity and elevated temperatures can accelerate enzymatic activities, leading to changes in the physical appearance and decomposition of the herb.

43

Dried herbs are particularly prone to contamination with spores of bacteria and fungi present in the air. Bacterial growth is usually accompanied by the growth of molds, whose presence is evidenced by changes in appearance; break down of the plant material, and smell. Mites, nematode worms, insects/moths, and beetles can also destroy ayurvedic drugs during storage.

Control measures to protect against deterioration include the use of airtight containers made of materials that will not interact physically or chemically with the material being stored. Storage in ventilated, cool, dry areas and periodic spraying of the stored area with insecticides will help to prevent the spread of infestation.

Sterilization of crude drugs is achieved by treatment of bulk consignments with ethylene oxide, and methyl bromide under controlled conditions and complying with acceptable limits for toxic residues. Markets from time to time experience wild fluctuations in the price of herbals. One reason for this is indiscriminate harvesting which leads to the extinction of natural populations still the only source of bioresources. This in turn encourages producers to replace the required herb with other supplements.

Contamination of Ayurvedic drugs and HerbDrug Interactions

Conventional synthetic pharmaceuticals such as synthetic corticosteroids, nonsteroidal anti- inflammatory drugs and other prescription drugs, potent drugs such as phenylbutazone, in fact examples of almost every therapeutic drug class have been found in certain herbal remedies as contaminants. A recent study by Ramsay et al. found that potent corticosteroids had been deliberately added to ayurvedic creams in order increase their efficacy. This problem is widespread, and occurs in both Asian and European countries. These ―adulterated‖ ayurvedic medicines sometimes result in serious ailments such as acute renal failure.

Many people, especially those living with HIV/AIDS, use both ayurvedic medicines and prescription drugs. A number of clinically significant interactions between prescribed and ayurvedic medicines have been identified. When these medications are used together, they can interact in the body, causing changes in the way the herbs and/or the drug works. Such changes are called herbdrug interactions. Concurrent use of ayurvedic or homeopathic

44

remedies alongside prescribed or over-the-counter medicines are frequent, and may mimic, magnify, or oppose the effect of the drug.

Herbdrug interactions are not chemical interactions between a drug and a ayurvedic component to produce something toxic. Instead, the interactions generally cause either an increase or decrease in the amount of drug in the bloodstream. As with conventional medicines, ayurvedic medicines interact with drugs in two general ways: pharmacokinetically and pharmacodynamically. Pharmacokinetic interactions result in alterations in the absorption, distribution, metabolism, or elimination of the drug or natural medicine. These interactions affect drug action by quantitative alterations, either increasing or decreasing the amount of drug available to have an effect. Pharmacodynamic interactions cause alterations in the way a drug or natural medicine affects a tissue or organ system. These actions affect drug action in a qualitative way, either through enhancing or antagonizing effects.

Herbdrug interactions change the effectiveness of the treatment, sometimes resulting in potentially dangerous side effects, possibly leading to toxicity, and/or reduced benefits. They can modify the mode of action of the drug, leading to unexpected complications or enhancement of the therapeutic effect, possibly leading to overmedication and an impact on health. Drug interactions are a significant problem in association with the use of St John‘s wort.

The risk of herbdrug interactions is not limited to synthetic drugs. Ayurvedic supplements and certain foods can interact with medications. Unfortunately very little is known about these interactions and there is little available scientific research on herbdrug interactions. When combining ayurvedic therapies with other medications, it is important to watch for potential symptoms and to inform health care providers. It is essential to train doctors to appreciate that drug interactions exist and to emphasize the importance of the need fo r physicians and naturopathic doctors to work together. Currently, there is very little information published on herbdrug interactions. Controlled clinical studies are needed to clarify and determine their clinical importance and more research is required to define them.

45

Toxicity of Ayurvedic drugs

For several reasons it is not possible to establish absolute safety standards for ayurvedic preparations based solely on epidemiological studies. First, these types of studies would be costly. Second, there is little published data in countries where the major use of medicinal plants occurs and thus general standards based on a limited number of reports would have little meaning. Third, the exact identification of the products implicated in side effects claimed for medicinal plants is usually lacking. In spite of these inadequacies, there are a number of general comments that can be made with regard to avoiding potential serious side effects from ayurvedic medicines. The definition of ―toxic‖ is ultimately a matter of viewpoint. Traditionally, herbs and ayurvedic products have been considered to be nontoxic and have been used by the general public and traditional medicinal doctors worldwide to treat a range of ailments. The fact that something is natural does not necessarily make it safe or effective. The active ingredients of plant extracts are chemicals that are similar to those in purified medications, and they have the same potential to cause serious adverse effects. Whilst the literature documents severe toxicity resulting from the use of herbs, on many occasions the potential toxicity of herbs and ayurvedic products has not been recognized. In India, herbs can be obtained from temples, night markets, street vendors, herbal stores, neighborhoods, or relatives, and from traditional medicine practitioners. Ordinary people recommend the medicines to others without safety considerations. The general public and many practitioners also believe that the herbs are nontoxic. Apparently, this cultural style/concept needs more attention in terms of drug safety education. Herbs and ayurvedic preparations can cause toxic adverse effects, serious allergic reactions, adverse drug interactions, and can interfere with laboratory tests. High-risk patients such as the elderly, expectant mothers, children, those taking several medications for chronic conditions, those with hypertension, depression, high cholesterol or congestive heart failure, should be more cautious in taking ayurvedic medicine. It is axiomatic that pregnancy should be a time of minimal medical intervention, and vaids in particular regard pregnancy as a ―contraindication‖ to taking ayurvedic medicines.

Two kinds of side effects have been reported for ayurvedic medicines. The first, considered to be intrinsic to ayurvedic drugs themselves, is mainly related to predictable toxicity due to toxic constituents of the ayurvedic ingredients and overdosage, and the second is allergy. Many cases of allergic reactions have been reported for ayurvedic drugs. It is impossible to

46

completely eliminate the possibility of any substance, including prescription drugs, ayurvedic remedies, or cosmetics, producing an allergic response in people exposed to them. Ayurvedic medicines do not present any more of a problem in this respect than any other class of widely used foods or drugs. Based on published reports, the side effects or toxic reactions associated with ayurvedic medicines in any form are rare. This could be due to the fact that ayurvedic medicines are generally safe, that adverse reactions following their use are underreported, or because the nature of the side effects or minor allergic reactions is such that they are not reported.

Perhaps the major problem with regard to the safety of ayurvedic medicines is related to the manufacturing practice, including contamination, substitution, incorrect preparation and dosage, intentional addition of unnatural toxic substances, interactions involving synthetic prescriptions, drugs, and ayurvedic medicines, either intentional or unintentional mislabeling, and the presence of natural toxic contaminants.

Many ordinary foods contain constituents that could be regarded as poisonous. Alpha gliadin produced by gluten in wheat, oats, and rye, the cyanogenic glycosides in many fruit skins and seeds, thiocyanates of the brassica vegetables, and lectins of many pulses including soya and red kidney bean are such examples. Cyanogenetic glycodides present in the kernel of many fruits can undergo gastric hydrolysis, resulting in the release of hydrogen cyanide. Viscotoxins, which are constituents of mistletoe, are both cytoxic and cardiotoxic. Nonetheless, these foods are generally regarded as safe. Similarly, both water and oxygen can kill in excessive amounts! So quantity is often an important consideration. A number of cases have been reported in the literature in which ayurvedic medicines, used for a number of years with safety, suddenly appear to be unsafe, and to date there has been no satisfactory explanation for these adverse effects. In this context herbs can be broadly classified into three major categories:

The food herbs medicines such as peppermint, ginger, garlic, hawthorn, nettles, lemon, and balm are gentle in action, have low toxicity, and are unlikely to cause any adverse response. They can be consumed in substantial quantities over long periods of time without any acute or chronic toxicity. However they may bring about allergic reactions in certain individuals.

47

The medicinal herbs – these are not daily ―tonics‖ and need to be used with greater knowledge (dosage and rationale for use) for specific conditions (with a medical diagnosis) and usually only for a limited period. They have a greater potential for adverse reactions and in some cases drug interactions. They include aloe vera, black cohosh, comfrey, echinacea, ephedra, ginkgo biloba, ginseng, kava kava, milk thistle, and senna.

The poisonous herbs have a strong potential for either acute or chronic toxicity and should only be prescribed by trained clinicians who understand their toxicology and appropriate use.

Fortunately, the vast majority of these herbs is not available to the public and is not sold in health food or ayurvedic stores. Aconite, Arnica spp., Atropa belladonna, digitalis, datura, male fern, gelsemium, and veratrum are some examples.

There are herbs such as Lobelia and Euonymus spp. that have powerful actions, often causing nausea or vomiting, although they are safe under appropriate conditions. There is also an idiosyncratic grouping of herbs that have been alleged, with some scientific support, to exhibit specific kinds of toxicity. The best known example is the hepatotoxicity of pyrrolizidine alkaloid-containing plants such as Symphytum (comfrey), Dryopteris (male fern), Viscum (mistletoe), and Corynanthe (Yohimbe).

Screening of Ayurvedic drugs

Once the botanical identity of a herb is established, the next step is phytochemical creening, which involves bioassays, extraction, purification, and characterization of the active constituents of pharmaceutical importance. The herb or ayurvedic drug preparation in its entirety is regarded as the active substance. These constituents are either of known therapeutic activity or are chemically defined substances or a group of substances generally accepted to contribute substantially to the therapeutic activity of a ayurvedic drug. In any program in which the end product is to be a drug, some type of pharmacological screening, or evaluation, must obviously be done.

48

Pharmacological screening programs are not without problems. Ideally the active principles should be isolated, preferably using bioassay guided isolation processes, which can be problematic. The ideal pharmacological screen would be to identify those extracts or pure compounds that are highly active and nontoxic. Such a screen is rare to find. Failure to duplicate pharmacological results is another problem. There are many pharmacological screening tests available. In the random selection program of the National Cancer Institute (NCI) in the US, plants are randomly selected, extracted, and the extracts are evaluated against one or more in vitro tumor systems and in vitro cytotoxicity tests. An extension of this procedure is to isolate metabolites or ―active compounds‖ from the plant that had shown most promising activity and subject them to pharmacological tests. In another approach, plants containing specific types or classes of chemical compounds, for example alkaloids, are tested. Simple tests such as color reactions are carried out on various parts of the plant in the field, and assays are carried out in the laboratories. In terms of costbenefit ratio, these ―shotgun‖ approaches are considered to be very unsatisfactory. Another method involves random collection of plants and subjection of their extracts to several broad screening methods and pharmacological tests. The success of this method depends on the number of samples assayed, adequate funding, and appropriate predictable bioassay protocols. Broad-based empirical screening, which is time consuming and expensive, can detect novel activities but is not suited for screening large numbers of samples.

Diagnosis by observation, a method introduced by the ―father‖ of medicine, Hippocrates, is still one of the most powerful tools of today‘s physicians. In vitro screening methods, though restricted to the detection of defined activities, are simpler and more useful. Recently, biochemical and receptorligand binding assays have gathered momentum. This has been made possible by the increasing availability of human receptors from molecular cloning, and extracts and compounds can be tested for binding directly to the presumed therapeutic target protein. Clone receptors can be expressed in a functional state linked to receptor proteins in cells such as yeast, and this has been made possible by applications of molecular biology. Combined with automated instrumentation and computer databases, hundreds of such assays can be completed in relatively short periods of time. These screening processes are successfully used by international agencies such as the National Cancer Institute (NCI) in the United States and the Central Drug Research Institute in India.

49

The technology of plant medicinal screening processes has even advanced to enzyme isolation. The enzymes that cause the disease are first isolated and the plant extracts are tested to determine if they block enzyme action. An enzyme immunoassay for the quantification of femtomole quantities of therapeutically important alkaloids has been established. Ethanolic extracts, tinctures, and pure plant compounds from commercially available herbs have been analyzed for their in vitro cytochrome P450 3A4 (CYP3A4) inhibitory capability via a fluorometric microtiter plate assay. These studies indicate that high-throughput screening methods for assessing CYP3A4 inhibition by natural products have important implications for predicting the likelihood of potential herbdrug interactions.

Higher plants contain both mutagens and antimutagens and are susceptible to mutagenesis, but screening programs for the detection of antimutagenesis rarely employ higher plant systems. However, using modified screening tests to detect antimutagenic agents, higher plants have been shown to contain a variety of structurally novel antimutagenic agents. Short- term bacterial and mammalian tissue culture systems are the standard methods employed.

Labeling of Ayurvedic Products

The quality of consumer information about the product is as important as the finished Ayurvedic product. Warnings on the packet or label will help to reduce the risk of inappropriate uses and adverse reactions. The primary source of information on ayurvedic products is the product label. Currently, there is no organization or government body that certifies an herb or a supplement as being labeled correctly.

It has been found that ayurvedic remedy labels often cannot be trusted to reveal what is in the container. Studies of ayurvedic products have shown that consumers have less than a 50% chance of actually getting what is listed on the label, and published analyses of ayurvedic supplements have found significant differences between what is listed on the label and what is in the bottle. The word ―standardized‖ on a product label is no guarantee of higher product quality, since there is no legal definition of the word ―standardized.‖ Consumers are often left on their own to decide what is safe and effective for them and the lack of consistent labeling on ayurvedic products can be a source of consumer frustration.

50

Certain information such as ―the product has been manufactured according to Pharmacopoeia standards,‖ listing of active ingredients and amounts, directions such as serving quantity (dosage) and frequency of intake of the drug, must be included on the labels of all ayurvedic products and packages. The label should also indicate the method of extraction and relative amount of macerate and menstruum used, and possible side effects. It should indicate that the product‘s content has been standardized to contain a particular amount of a specified biochemical constituent. Standardization gives the buyers a measure of potency by which to judge the quality of the product and to compare dosage with those indicated by clinical trials. This will also ensure that the correct herb has been used. In addition to the above information, the label should include the name and origin of the product, its intended use, net quantity of contents, other ingredients such as herbs and amino acids, and additives, for which no daily values have been established, storage conditions, shelf life or expiry date, warnings, disclaimer, and name and address of manufacturer, packer or distributor. A herb categorized as a nutritional supplement cannot claim any health benefits or ―disease claims‖ on the label, leaving the consumer with little information.

Marketing plays a big role in the use of ayurvedic products and the media help significantly to provide information about natural health products. One of the problems with mass media ―propaganda‖ is scientific inconsistency. Unless the packaging contains a medical claim, ayurvedic products are not reviewed by any government agency. Food and drug administrations that regulate prescription drugs only review a ayurvedic product if the item is suspected of being harmful or if the label contains medical claims. Scientists use several approaches to evaluate botanical dietary supplements for their potential health benefits and safety risks, including their history of use and laboratory studies using cell or animal models. Studies involving people can provide information that is relevant as to how botanical dietary supplements are used.

Policies and Regulations

It is a widely held myth that modern drugs are dangerous foreign chemicals with side effects, while ayurvedic medicines are natural, gentle and safe. The truth is that some herbs can be dangerous and can bring about serious diseases and even lead to death. Unlike conventional

51

drugs, ayurvedic products are not regulated for purity and potency and this could cause adverse effects and can even lead to drug interactions. There are fewer studies on ayurvedic medicines than on conventional drugs, mainly because, unlike synthetic chemicals, herbs cannot be patented, so there is little money to be made by funding such research. It is important that consumers are made aware of interactions herbs might have with other drugs they are taking. Unfortunately this information is not available with ayurvedic medicines. Ayurvedic medicines are also frequently adulterated with prescription drugs. In certain countries, ayurvedic products used for diagnosis, cure, mitigation, treatment, or prevention of disease are normally treated as drugs, and hence regulated by legislation.

However, in most countries, including the United States, such legislation does not exist and in fact, most botanical products are marketed as dietary supplements. Ayurvedic products categorized as nutritional or dietary supplements are not regulated. In many countries these medicines are not required to pass any regulatory analysis to be sold as health food supplements. It is clear that the ayurvedic industry needs to follow strict guidelines and that regulations are needed. The food and drug administration‘s that regulate prescription drugs only review a ayurvedic product if the item is suspected of being harmful or if the label contains a medical claim. Although research is being done, it is very limited and only a few ayurvedic drugs have been studied adequately by well-controlled clinical trials. Even though evidence should always be presented to support claims of products, most herbs are still marketed with little or no research. To be registered as drugs, these products need to be tested to prove their safety and clinical efficacy. However, so far, few programs have been established to study the safety and efficacy of ayurvedic medicines as originally proposed in the WHO guidelines for the assessment of herbal medicines.

The future of ayurvedic drugs is overshadowed by the pervading lack of regulatory control. In 1993, the WHO sponsored a symposium on the use of medicinal plants. The result was a standard guideline for the assessment of herbal medicines and a recommendation that governments of the world should protect medicinal plants, improve regulation of herbal medicines, and respect traditional medicine approaches.

More recently the Department of Ayush developed a new regulatory framework for natural health products, which came into effect in 1995.

52

Among other things, the new regulations call for improved labeling, good manufacturing practices, product and site licensing, and provision of a full range of health claims that will be supported by evidence. However, even in India, the only regulatory requirements enforced are that all products intended for medicinal use, including natural health products, are issued a Drug Identification Number (DIN). These numbers are not required for raw materials such as bulk herbs. In the US, access to herbal medicines is restricted by FDA regulations. Before any new chemical or ayurvedic drug is approved, research must prove that it is both safe and effective. As a result of these restrictions, packages of ayurvedic medicines are labeled as food supplements, which do not require pre-approved testing. Food supplements cannot make any healing claims or issue warnings about potential risks. In the US, plant-based derivatives already appear in a quarter of the prescription medicines produced. However, many other plants with healing properties are shunned by the medical community despite scientific data from other countries showing their effectiveness. The misconception that herbs are old fashioned and unscientific has helped to promote a general distrust of phytotherapy. The Central Drug Research Institute contends that, in many cases, ayurvedic medicines are safer than prescription drugs. According to the CDRI, ayurvedic medicines react more slowly and often include their own antidotes to counteract any toxic effects.

With proper enforcement of regulations, more products that are legitimate will enter the market and the consumers will see justifiable claims on labels. In fact, it is predicted that appropriate regulations will rejuvenate the indian market in response to growing concerns about the regulatory environment for ayurvedic remedies.

53

PROTOCOLS FOR TESTING THE QUALITY OF OILS, PASTE, TABLETS, POWDER AND SYRUP

OILS

1. Physical Properties

a) Description

b) Colour:

c) Odour

d) Viscosity

2. Test for heavy metal

a) Lead

b) Cadmium

c) Mercury

d) Arsenic

LEAD: The following method is based on the extraction of lead by solutions of dithizone.

All reagents used for the test should have as low a content of lead as practicable. All reagent

solutions should be stored in containers of borosilicate glass. Glassware should be rinsed

thoroughly with warm dilute nitric acid, followed by water.

Special Reagents

(1) Ammonia-cyanide solution Sp. Dissolve 2 g of potassium cyanide in 15 ml of strong

ammonia solution and dilute with water to 100 ml.

(2) Ammonium citrate solution Sp. Dissolve 40 g of citric acid in 90 ml water. Add two

drops of phenol red solution then add slowly strong ammonia solution until the solution

acquires a reddish colour. Remove any lead present by extracting the solution with 20 ml

quantities of dithizone extraction solution until the dithizone solution retains its orange-green

colour.

54

(3) Dilute standard lead solution Dilute 10.0 ml of standard lead solution with sufficient 1 per cent v/v solution of nitric acid to produce 100.0 ml. Each ml of this solution contains 1 μg of lead per ml.

(4) Dithizone extraction solution Dissolve 30 mg of diphenylthiocarbazone in 1000 ml of chloroform and add 5 ml of alcohol. Store the solution in a refrigerator. Before use, shake a suitable volume of the solution with about half its volume of 1 per cent v/v solution of nitric acid and discard the acid. (5) Hydroxylamine hydrochloride solution Sp. Dissolve 20 g of hydroxylamine hydrochloride in sufficient water to produce about 65 ml. Transfer to separator, add five drops of thymol blue solution, add strong ammonia solution until the solution becomes yellow. Add 10 ml of a 4 per cent w/v solution of sodium diethyldithiocarbamate and allow standing for five minutes. Extract with successive quantities, each of 10 ml, of chloroform until a 5 ml portion of the extract does not assume a yellow colour when shaken with dilute copper sulphate solution. Add dilute hydrochloric acid until the solution is pink and then dilute with sufficient water to produce 100 ml.

(6) Potassium cyanide solution Sp. Dissolve 50 g of potassium cyanide in sufficient water to produce 100 ml. Remove the lead from this solution by extraction with successive quantities, each of 20 ml of dithizone extraction solution until the dithizone solution retains its orange-green colour. Extract any dithizone remaining in the cyanide solution by shaking with chloroform. Dilute this cyanide solution with sufficient water to produce a solution containing 10 g of potassium cyanide in each 100 ml.

(7) Standard dithizone solution Dissolve 10 ml of diphenyl thiocarbazone in 1000 ml of chloroform. Store the solution in a glass-stoppered, lead-free bottle, protected from light and in a refrigerator.

(8) Citrate-cyanide wash solution To 50 ml of water add 50 ml of ammonium citrate solution Sp. and 4 ml of potassium cyanide solution Sp., mix, and adjust the pH, if necessary, with strong ammonia solution to 9.0.

55

(9) Buffer solution pH 2.5 To 25.0 ml of 0.2 M potassium hydrogen phthalate add 37.0 ml of 0.1 N hydrochloric acid and dilute with sufficient water to produce 100.0 ml.

(10) Dithizone-carbon tetrachloride solution Dissolve 10 mg of diphenylthiocarbazone in 1000 ml of carbon tetrachloride. Prepare this solution fresh for each determination.

(11) pH 2.5 wash solution To 500 ml of a 1 per cent v/v nitric acid add strong ammonia solution until the pH of the mixture is 2.5, then add 10 ml of buffer solution pH 2.5 and mix.

(12) Ammonia-cyanide wash solution To 35 ml of pH 2.5 wash solution add 4 ml of ammonia-cyanide solution Sp., and mix.

Method Transfer the volume of the prepared sample directed in the monograph to a separator and unless otherwise directed in monograph, adds 6 ml of ammonium citrate solution Sp., and 2 ml hydroxylamine hydrochloride solution Sp., (For the determination of lead in iron salts use 10 ml of ammonium citrate solution Sp.). Add two drops of phenol red solution and make the solution just alkaline (red in color) by the addition of strong ammonia solution. Cool the solution if necessary, and add 2 ml of potassium cyanide solution Sp. Immediately extract the solution with several quantities each of 5 ml, of dithizone extraction solution, draining off each extract into another separating funnel, until the dithizone extraction solution retains its green colour. Shake the combinedithizone solutions for 30 seconds with 30 ml of a 1 per cent w/v solution of nitric acid and discard the chloroform layer. Add to the solution exactly 5 ml of standard dithizone solution and 4 ml of ammonia-cyanide solution Sp. and shake for 30 seconds; the color of the chloroform layer is of no deeper shade of violet than that of a control made with a volume of dilute standard lead solution equivalent to the amount of lead permitted in the sample under examination.

CADMIUM: Determination Conditions: - Reference condition: dry temperature: 100- 120ºC, maintain 20 seconds; ash temperature: 300-500ºC, maintain 20-25 seconds; atomic temperature: 1500-1900ºC, maintain 4-5 seconds; measurement wavelength: 228.8 nm; background calibration: deuterium lamp (D lamp) or Zeeman Effect.

56

II Preparation of Cd standard stock solution: - Measure accurately a quantity of Cd

single-element standard solution to prepare standard stock solution Cd with 2% HNO 3 , which containing 0.4 μg per ml Cd, stored at 0-5ºC. III Preparation of calibration curve: - Measure accurately a quantity of cadmium standard stock solutions, diluted to the concentration of 1.6, 3.2, 4.8, 6.4 and 8.0 ng per ml with 2% HNO3 respectively. Pipette accurately 10 μl the above solutions respectively, inject them into the graphite oven, determine their absorbance, and then draw the calibration curve with absorbance as vertical axis and concentration as horizontal ordinate.

IV

Preparation of test solution: - Reference to method of ―Preparation of test solution‖ of

Pb

in the above.

V

Determination: - Pipette accurately 10-20 μl of the test solution and its corresponding

reagent blank solution respectively, determine their absorbance according to the above method of ―Preparation of calibration curve‖ (If interference occurs, weigh accurately respectively 1 ml of the standard solution, blank solution and test solution, add 1 ml of a solution containing 1% NH4H2PO4 and 0.2% Mg (NO3)2, shake well, determine their absorbance according to the method above, calculate the content of Cd in the test solution from the calibration curve.

MERCURY: Determination Conditions: - Apparatus suitable hydride generator device; reducing agent: a solution containing 0.5% sodium borohydride and 0.1% sodium hydroxide; carrier liquid: 1% hydrochloric acid; carrier gas: nitrogen; measurement wavelength: 253.6 nm; background calibration: deuterium lamp (D lamp) or Zeeman Effect.

II Preparation of mercury standard stock solution: - Measure accurately a proper quantity

of mercury single-element standard solution to prepare standard stock solution with 2% nitric acid solution, which containing 1.0 μg per ml Hg, stored at 0-5ºC.

III Preparation of calibration curve: - Measure accurately 0, 0.1, 0.3, 0.5, 0.7 and 0.9 ml of

mercury standard stock solution, transfer into a 50 ml volumetric flask respectively, add 40

ml 4% sulfuric acid solution and 0.5 ml of 5% potassium permanganate solution, shake well,

57

drop 5% hydroxylamine hydrochloride solution until the violet red just disappears, dilute with 4% sulfuric acid solution to the volume, shake well. A quantity of each solution is injected to the hydride generator device, determine the absorbance, and then plot the calibration curve with peak area (absorbance) as vertical axis and concentration as horizontal ordinate.

IV Preparation of test solution

Method :- Transfer 1 g of the coarse powder of the substance being examined, accurately weighed, into a casparian flask, add 5-10 ml of the mixture solution of nitric acid solution (HNO3) and perchloric acid (HCIO 4 ) (4 : 1), mix well, fix a small hopper on the flask-top, immerse overnight, heat to slake on the electric hot plate at 120-140ºC for 4-8 hours until slaking completely, cool, add a quantity of 4% sulfuric acid solution and 0.5 ml of 5% potassium permanganate solution, shake well, drop 5% hydroxylamine hydrochloride solution until the violet red colour just disappears, dilute with 4% H 2 SO 4 solution to 25 ml, shake well, centrifugate if necessary, the supernatant is used as the test solution. Prepare synchronally the reagent blank solution based on the same procedure.

V Determination :- Pipette accurately a quantity of the test solution and its corresponding

reagent blank solution, respectively, proceed as described under ―Preparation of calibration curve‖ beginning at the words ―add 1 ml of 25% potassium iodide solution‖. Calculate the content of mercury (Hg) in the test solution from the calibration curve.

ARSENIC: Determination Conditions: - Apparatus suitable hydride generator device, reducing agent: a solution containing 1% sodium borohydride and 0.3% sodium hydroxide; carrier liquid: 1% hydrochloric acid; carrier gas: nitrogen; measurement wavelength: 193.7 nm; background calibration: deuterium lamp (D lamp) or Zeeman Effect.

II Preparation of As standard stock solution :- Measure accurately a quantity of As single-

element standard solution to prepare standard stock solution with 2% nitric acid solution (HNO3), which containing 1.0 μg per ml As, stored at 0-5ºC.

58

III Preparation of calibration curve: - Measure accurately proper quantity of arsenic standard stock solutions, diluted with 2% HNO3 to the concentration of 2, 4, 8, 12 and 16 ng per ml respectively. Accurately transfer 10 ml of each into 25 ml volumetric flask respectively, add 1 ml of 25% potassium iodide solution (prepared prior to use), shake well, add 1 ml of ascorbic acid solution (prepared prior to use), shake well, dilute with hydrochloric acid solution (20-100) to the volume, shake well, close the stopper and immerse the flask in a water bath at 80ºC for 3 minutes. Cool, transfer proper quantities of each solution respectively into the hydride generator device, determine the absorbance, then plot the calibration curve with peak area (absorbance) as vertical axis and concentration as horizontal ordinate.

IV Preparation of test solution: - Reference to method of ―Preparation of test solution‖ of Pb in the above.

V Determination :- Pipette accurately 10 ml of the test solution and its corresponding reagent blank solution respectively, proceed as described under ―Preparation of calibration curve‖ beginning at the words ―add 1 ml of 25% potassium iodide solution‖. Calculate the content of As in the test solution from the calibration curve.

3. Microbial contamination

The tests for microbial contamination are carried out on the same sample of the preparations being examined using the above-stated media. When the quantity in a single container is insufficient to carry out the tests, the combined contents of the two or more containers are used to inoculate the above-stated media.

Remove the liquid from the test containers with a sterile pipette or with a sterile syringe or a needle. Aseptically transfer the specified volume of the material from each container to a vessel of the culture medium. Mix the liquid with the medium but do not aerate excessively. Incubate the inoculated media for not less than 14 days, unless otherwise specified in the monograph, at 30º to 35º in the case of fluid thioglycollate medium and at 20º to 25º in the case of soyabean-casein digest medium. When the material being examined renders the medium turbid so that the presence or absence of microbial growth cannot be determined readily by visual examination, transfer suitable portions of the medium to fresh vessels of the

59

same medium between the third and seventh days after the test is started. Continue incubation of the transfer vessels for not less than 7 additional days after the transfer and for a total of not less than 14 days.

For oils and oily solutions : Use media to which have been added 0.1% w/v of (4-tert- octylphenoxy) polyethoxyethanol, 1% w/v of polysorbate 80 or other suitable emulsifying agent, in an appropriate concentration, shown not to have any antimicrobial properties under the conditions of test. Carry out the test as described under For aqueous solutions and suspensions.

Cultures containing oily preparations should be shaken gently each day. However, when fluid thioglycollate medium is used for the detection of anaerobic micro-organisms, shaking or mixing should be kept to a minimum to maintain anaerobic conditions.

For ointments : Prepare by diluting ten-fold in a sterile diluent such as fluid B or any other aqueous vehicle capable of dispersing the test material homogeneously throughout the fluid mixture (Before use, test the dispersing agent to ascertain that in the concentration used it has no significant antimicrobial effects during the time interval for all transfers). Mix 10 ml of the fluid mixture so obtained with 80 ml of the medium and proceeds as directed under for aqueous solutions and suspensions.

For solids: Transfer the quantity of the preparation to be examined to the quantity of medium specified in Table 5 and mix, the conditions of incubation being the same as For aqueous solutions and suspensions. Proceed as directed under for aqueous solutions and suspensions beginning at the words ―When the material being examined‖. 3.8.8.5. For sterile devices : For articles of such size and shape as permit complete immersion in not more than 1000 ml of culture medium test the intact article, using the appropriate media, and incubating as directed under For aqueous solutions and suspensions. For transfusion or infusion assemblies or where the size of an item makes immersion impracticable and only the liquid pathway must be sterile, flush the lumen of each of twenty units with a sufficient quantity of fluid thioglycollate medium and the lumen of each of twenty units with a sufficient quantity of soyabean-casein digest medium to yield a recovery of not less than 15 ml of each medium, and incubate with not less than 100 ml of each of the two media as directed under For

60

aqueous solutions and suspensions. For devices in which the lumen is so small that fluid thioglycollate medium will not pass through, substitute alternative thioglycollate medium for the fluid thioglycollate medium and incubate that inoculated medium an aerobically. Where the presence of the specimen being tested, in the medium interferes with the test because of bacteriostatic or fungistatic action, rinse the article thoroughly with the minimum amount of fluid A. Recover the rinsed fluid and test as described For sterile devices under Method A.

Observation and Interpretation of Results At intervals during the incubation period, and at its conclusion, examine the media for macroscopic evidence of microbial growth. If no evidence of growth is found, the preparation being examined passes the tests for sterility. If evidence of microbial growth is found, reserve the containers showing this and, unless it is demonstrated by any other means that their presence is due to causes unrelated to the preparation being examined and hence that the tests for sterility are invalid and may therefore be recommenced, perform a retest using the same number of samples, volumes to be tested and the media as in the original test. If no evidence of microbial growth is then found, the preparation being examined passes the tests for sterility. If evidence of microbial growth is found, isolate and identify the organisms. If they are not readily distinguishable from those growing in the containers reserved in the first test, the preparation being examined fails the tests for sterility. If they are readily distinguishable from those growing in the containers reserved in the first test, perform a second retest using twice the number of samples. If no evidence of microbial growth is found in the second retest, the preparation being examined passes the tests for sterility. If evidence of growth of any micro-organisms is found in the second retest the preparation being examined fails the tests for sterility.

4. TOTAL BACTERIAL COUNT /FUNGAL COUNT OBJECTIVE Test for Total Microbial Count is provided to determine compliance with the requirements given in individual monograph / specifications.

PRINCIPLE: Total microbial count is the estimation of the number of viable aerobic micro- organisms present in the pharmaceutical articles of all kinds, from raw materials to the

61

finished

forms.

BUFFERS AND MEDIA 1. pH 7.2 Phosphate Buffer Stock Solution: Dissolve 34 g of Monobasic Potassium Phosphate in about 500 mL of water contained in a 1000 mL volumetric flask. Adjust to pH 7.2 ± 0.1 by the addition of 4 % w/v aqueous solution of Sodium Hydroxide (about 175 mL), add water to volume, and mix. Dispense and sterilize. Store under refrigeration. For use, dilute the Stock Solution with water in the ration of 1 to 800, and sterilize in an autoclave at 121 0 C , 15 lb pressure for about 15 min.

2. Fluid Soyabean Casein Digest Medium

Pancreatic Digest of Casein

17.0

g

Papacy

Digest

of

Soybean

3.0

g

Meal

 

Sodium Chloride

 

5.0

g

Dibasic Potassium Phosphate

2.5

g

Dextrose (C 6 H 12 O 6 . H 2 O)

2.5

g

Distilled Water

 

1000 mL

Final pH after Sterilization

7.3

± 0.2

Dissolve the solids in the water, warming slightly to effect solution. Cool to room temperature and adjust pH. 7.3 ± 0.2 and Filter, if necessary. Distribute the media into suitable containers and sterilize in an autoclave at 121 OC for about 15min.

3. Fluid Casein Digest-Soy Lecithin-Polysorbate 20 Medium

Pancreatic Digest of Casein

20.0

g

Soy Lecithin

5.0

g

Polysorbate

20 -40 mL

Water

960 mL

62

Dissolve the pancreatic digest of casein and soy lecithin in 960 mL of water, heating in a water batch at 48 - 50 OC for about 30 min. to effect solution. Add 40 mL of Polysorbate 20. Mix, and dispense as desired and sterilize in an autoclave at 121 OC for about 15min

4. Sabouraud Glucose Agar with Antibiotics

Peptones (meat and Casein)

10.0

g

D- Glucose Monohydrate

40.0

g

Agar

15.0

g

Water

1000 mL

Water Adjust the pH to 5.4 +2. Sterilize by heating in an autoclave at 121 0 C for 15 min. immediately before use, add 0.10 g of Benzylpenicillin Sodium and 0.10 g of Tetracycline per liter of medium as sterile solutions or, alternatively, add 50 mg of Chloramphenicol per liter of medium.

5. Peptone Water

(Buffered Sodium Chloride - Peptone Solution pH 7.0)

Potassium Dihydrogen Orthophosphate

3.56

g

Disodium Hydrogen Orthophosphate

7.23

g

Sodium Chloride

4.30

g

Peptone(meat and Casein)

1.0 g

Water

1000 mL

0.1 to 1.0 % w/v Polysorbate 20 or Polysorbate 80 may be added. Sterilize by heating in an autoclave at 121 0 C for 15 min.

6. Potato Dextrose Agar Medium

63

Cook 300 g of peeled and diced potatoes in 500 mL of water prepared by distillation, filter through cheesecloth, add water prepared by distillation to make 1000 mL and add the following:

Agar

15

mg

Glucose

20

mg

pH after sterilization

5.6 ± 0.2

Dissolve by heating, and sterilize.

For use, just prior to pouring the plates, adjust the melted and cooled medium to 45 0 C with Sterile Tartaric Acid solution (1 in 10) to a pH of 3.5 ± 0.1 Do not reheat the pH 3.5 medium. All the above media should be incubated for 24 hours at 37 0 C before use. Any contaminated media should be discarded.

Instead of preparing media, one can also use dehydrated media of Hi media / Difco and rehydrate the required quantity as per instructions on the bottle label, dispense in required quantities and sterilize.

Preliminary Testing The methods given herein are invalid unless it is demonstrated that the test specimens to which they are applied to not, of themselves, inhibit the multiplication, under the test conditions, of micro-organisms that may be present. Therefore, prior to doing the tests,

inoculate diluted specimens of the substance being examined, with separate viable cultures of Escherichia coli, B. subtilis and Staphylococcus aureus. Add 1 mL of not less than 10 -3 dilution of a 24 hour broth culture of the micro-organism to the first dilution (in buffer solution pH 7.2, Fluid Soybean-Casein Digest medium, of the test material and following the test procedure. If the organisms fail to grow in the relevant medium the procedure should be

modified

a. increasing the volume of diligent, the quantity of test material remaining the same,

b. incorporating a sufficient quantity of a suitable inactivating agent in the diluents or by,

64

by

c. combining the afore-mentioned modifications so as to permit growth in the media.

If inhibitory substances are present in the sample 0.5 % of soy lecithin and 4 % of the

Alternatively, repeat the test as

described in the previous paragraph, using fluid casein digest - soy lecithin - polysorbate 20 medium to demonstrate neutralization of preservatives or other anti-microbial agents in the test material.

Polysorbate 20 may be added to the culture medium

PRE - TREATMENT OF THE PREPARATION BEING EXAMINED

Use separate 10 mL or 10 g specimens for testing.

Water - Soluble Products:

Dissolve or dilute 10 g or 10 mL of the preparation being examined, unless otherwise prescribed, in Peptone Water or another suitable medium shown not to have anti-microbial activity under the conditions of the test and adjust the volume to 100 mL with the same medium. If necessary, adjust the pH to about 7.

Non - Fatty Products Insoluble in Water:

Suspend 10 g or 10 mL of the preparation being examined, unless otherwise prescribed, in Peptone Water or another suitable medium shown not to have anti-microbial activity under the conditions of the test and dilute to 100 mL with the same medium. If necessary divide the preparation being examined and homogenize the suspension mechanically.

A suitable surface - active agent such as 0.1 % w/v of Polysorbate 80 may be added to assist

the suspension of poorly wettable substances. If necessary, adjust the pH of the suspension to about 7

Fatty Products:

Homogenize 10 g or 10 mL of the preparation being examined, unless otherwise prescribed with 5 g of Polysorbate 20 or Polysorbate 80. If necessary, heat to not more than 40 0 C. Mix carefully while maintaining the temperature in a water bath or in an oven. Add 85 mL of Peptone Water or another suitable medium shown not to have anti-microbial activity n the conditions of the test, heated to not more than 40 OC if necessary. Maintain this temperature

65

for the shortest time necessary for formation of an emulsion and in any case for not more than 30 min. If necessary, adjust the pH of the emulsion to about 7.

PROCEDURE As per IP

1. Dissolve or suspend 10 g of the substance being examined in sufficient buffer solution, pH 7.2, Fluid Soybean - Casein Digest Medium, or Fluid Casein Digest-Soy lecithin - Polysorbate 20 medium to produce 100 mL.

2. Perform the test for the absence of inhibitory (anti-microbial) properties as described under Preliminary Testing before the determination of Total Microbial Count.

3. Add the substance being examined, to the medium not more than 1 hour after

preparing the appropriate dilution‘s for inoculation.

A. Plate Method

1. This method is applicable for substances that are sufficiently soluble or translucent.

Dilute further, if necessary, the sample liquid prepared as described above, so that 1

mL will be expected to yield between 30 to 300 colonies.

2. Pipette 1 mL of the final dilution into each of two sterile petri dishes. Immediately add to each dish 15 to 20 mL of Soybean-Casein digest agar medium

that has previously been melted and cooled to about 45 0 C.

3. Cover the petridishes, mix the sample with the agar by tilting or rotating the dishes,

and allow the contents to solidify to room temperature.

4. Invert the petri dishes, and incubate for 48 to 72 hours at 37 0 C.

5. After incubation examine the plates of growth, count the number of colonies, and

express the average for the two plates in terms of number of micro-organisms per g of

the

If no colonies are recovered from the dishes representing the initial 1: 10 dilution of

the substance, express the results as ―less than 10 micro-organisms per g of substance‖.

substance.

66

B. Multiple - Tube Method

1. This method is applicable for substances that are insoluble or not translucent. Into each of the fourteen test tubes of similar size place 9 mL of sterile Fluid Soyabean casein digest medium.

2. Arrange 12 of the tubes in four sets of three tubes each. Put aside one set of three tubes to serve as the controls.

3. Into each of three tubes of one set [―100‖ ] and into fourth tube (A) pipette 1 mL of the Solution of suspension of the specimen, and mix.

4. From tube A pipette 1 mL of its contents into the one remaining tube (B) not included in a set, and mix.

5. These two tubes contain 100 mg (or 100 µl) and 10 mg (10 µl) of the specimen respectively.

6. Into each of the second set [―10‖] of three tubes pipette 1 mL from tube A and into each tube of the third set [―1‖] pipette 1 mL from tube B.

7. Discard the unused contents of tubes A and B.

8. Close well, and incubate all of the tubes.

9. Following the incubation period, examine the tubes for growth; the three control tubes remain clean and the observations in the tubes containing the specimen, when interpreted by reference to Table I. indicate the most probable number of micro- organisms per g or per mL of specimen.

As per BP

Determine the total viable aerobic count of the preparations being examined by the membrane filtration method, the plate count method or the serial dilution method as prescribed. Suitable degrees of dilution should be used so that the number of colony forming units is within the limits suggested for the method to be used.

A. Membrane Filtration

1. Use membrane filters having a normal pore size not greater than 0.45 µm and 50 nm in diameter the effectiveness of which in retaining bacteria has been established. For

67

example, Cellulose nitrate filters are used for aqueous, oily and weakly alcoholic solutions and Cellulose acetate filters for strongly alcoholic solutions.

2. The filtration apparatus and membrane are sterilized by appropriate means and are designed so that the solution being examined can be introduced and filtered under

aseptic conditions and so as to permit the removal of the membrane for transfer to the

medium.

culture

Transfer 10 mL or a quantity to each of two membrane filters and filter immediately. If necessary, dilute the pretreated preparation so that a colony count of 10 to 100 may be expected.

3. Wash each membrane by filtering through it three or more successive quantities , each of approximately 100 mL, of a suitable liquid such as buffered Sodium Chloride - peptone pH 7.0. For fatty substances, this liquid may contain a suitable surface active agent such as polysorbate 20 or polysorbate 80. Transfer one of the membrane filters, intended primarily for the enumerate of bacteria, to the surface of the plate of casein soybean digest agar and the other, intended primarily for the surface of a plate of sabouraud glucose agar with antibiotics.

4. Incubate the plates for 5 days, unless a more reliable count is obtained in a shorter time, at 30 - 35 0 C in the test intended to detect bacteria and at 20 - 25 OC in the test

intended

Count the number of colonies that are formed. Calculate the number of micro- organisms per gram or per milliliter of the preparation being examined, if necessary counting bacteria and fungi separately.

to

detect

fungi.

B. Plate Count

i. For Bacteria

1. Using petri dishes 9 to 10 cm in diameter, add to each dish a mixture of 1 mL of the pretreated preparation and about 15 mL of liquefied casein soyabean digest agar at not more than 45 OC .

2. Alternatively, spread the pretreated preparation on the surface of the solidified medium in a petri dish of the same diameter.

3. If necessary, dilute the pre-treated preparation as described above so that a colony count of not more than 300 may be expected.

68

4. Prepare at least two such petri dishes using the same dilution and incubate at 30 - 35OC for unless a more reliable count is obtained in a shorter time.

5. Count the number of colonies that are formed.

6. Calculate the results using plates with the greatest number of colonies but taking 300 colonies per plate as the maximum consistent with good evaluation.

ii. For Fungi

1. Using petri dishes 9 - 10 cm in diameter, add to each dish a mixture of 1 mL of the pre-treated preparation and about 15 mL of liquefied Sabouraud glucose agar with antibiotics at not more than 45 0 C.

2. Alternatively, spread the pretreated preparation on the surface of the solidified medium in a petri dish of the same diameter. If necessary, diluted the pretreated preparation as described above so that a colony count of not more than 100 may be expected.

3. Prepare atleast two such plates using the same dilution and incubate at 20 - 25 OC for 5 days, unless a more reliable count is obtained in a shorter time.

4. Count the colonies that are formed. Calculate the results using plates with not more than 100 colonies.

Serial Dilution

1. Prepare a series of 12 tubes each containing 9 - 10 mL of caseing soyabean digest broth.

2. To each of the first three tubes add 1 mL of the preparation diluted, dissolved or homogenized in the proportion 1 in 10, as described above.

3. To the next three tubes add 1 mL of a 2 in 100 dilution of the preparation and to the next three tubes add 1 mL of a 1 in 100 dilution of the preparation.

4. To the last three tubes add 1 mL of the diligent.

5. Incubate the tubes at 30 - 35 OC for at least 5 days.

6. The last three tubes should show no microbial growth.

7. If the reading of the results is difficult or uncertain owing to the nature of the preparation being examined, sub-culture on a liquid or solid medium and read the results after a further period of incubation.

69

8. Determine the most probable number of micro-organisms per gram or per milliliter of the preparation being examined from Table I.

As per USP

1.

For

specimens that are sufficiently soluble or translucent to permit use of the plate

Method, otherwise, use Multiple-tube Method.

2.

With either method, first dissolve or suspend 10 g of the specimen if it is a solid, or 10

mL

, accurately measured if the specimen is a liquid, in pH 7.2 Phosphate Buffer,

Fluid Soyabean-Casein Digest Medium, or Fluid Casein Digest-soy Lecithin - Polysorbate 20 Medium to make 100 mL.

3.

For

viscous specimens that cannot be pipetted at this initial 1:10 dilution, dilute the

specimen until a suspension is obtained i.e., 1:50 or 1: 100 etc., that can be pipetted.

4.

Perform the test for absence of inhibitory (anti-microbial) properties as described under Preparatory Testing before the determination of Total Aerobic Microbial Count.

5.

Add the specimen to the medium not more than 1 h after preparing the appropriate

dilution‘s

Plate Method

for

inoculation.

1. Dilute further, if necessary, the fluid so that 1mL will be expected to yield between 30

and 300 colonies.

2. Pipette 1 mL of the final dilution onto each of two sterile petridishes.

3. Promptly add to each dish 15 - 20 mL of Soyabean Casein Digest Agar Medium when testing for bacteria and add 15 - 20 mL Sabouraud Dextrose Agar medium or Potato

Dextrose Agar medium for molds and yeasts,. that previously has been melted and cooled to approx. 45 0 C.

4. Cover the petridishes, mix the sample with the agar by tilting or rotating the dishes,

and allow the contents to solidify at room temperature.

5. Invert the petridishes, and incubate for 48 - 72 hours at 37 OC for bacteria and incubate for 5 - 7 days at 20 - 25 0 C for molds and yeasts.

6. Following incubation, examine the plates for growth, count the number of colonies,

and express the average for the two plates in terms of the number of micro-organisms

per g or per mL of specimen.

70

7. If no microbial colonies are recovered from the dishes representing the initial 1: 10 dilution of the specimen, express the results as ―less than 10 micro-organisms per g or per mL of specimen.‖

CALCULATIONS

Take the average count of each dilution and multiply it by the dilution factor. Calculate the average form the readings obtained. This gives the count per gram or per mL of the sample/ Interpretation of Results :

If a limit is prescribed, it is to be interpreted as follows :

10

2 micro-organisms, MAXIMUM LIMIT OF ACCEPTANCE: 5 x 10 2

10

3 micro-organisms, MAXIMUM LIMIT OF ACCEPTANCE: 5 x 10 3

NOTE: Use the method as per the pharmacopoeial status (grade) of the material. In case of In- house specifications, follow the method as per the IP or as specified.

5. TEST FOR SPECIFIC PATHOGEN

a) E. coli

b) salmonella spp.

c) s.aureus

d) pseudomonas aeruginosa

For specified micro-organisms: Grow separately the test strains of Staphylococcus aureus and Pseudomonas aeruginosa in fluid soyabean-casein digest medium and Escherichia coli and Salmonella typhimurium at 30º to 35º for 18 to 24 hours. Dilute portions of each of the cultures using buffered sodium chloride-peptone solution pH 7.0to make test suspensions containing about 103 viable micro-organisms per ml. Mix equal volume of each suspension and use 0.4 ml (approximately 102 micro-organisms of each strain) as an inoculum in the test for E. coli, salmonellae, P. aeruginosa and S.aureus, in the presence and absence of the preparation being examined, if necessary. A positive result for the respective strain of micro- organism should be obtained.

71

6.

Pyrethroids

Pesticide

Residue,

Organochlorine

Pesticides,

Organophosphorus

Pesticides,

DEFINITION: For the purposes of the Pharmacopoeia, a pesticide is any substance or mixture of substances intended for preventing, destroying or controlling any pest, unwanted species of plants or animals causing harm during or otherwise interfering with the production, processing storage, transport or marketing of vegetable drugs. The item includes substances intended for use as growth-regulators, defoliants or desiccants and any substance applied to crops either before or after harvest to protect the commodity from deterioration during storage and transport.

Limits: Unless otherwise indicated in the monograph, the drug to be examined at least complies with the limits indicated in Table -1, The limits applying to pesticides that are not listed in the table and whose presence is suspected for any reason comply with the limits set by European Community directives 76/895 and 90/642, including their annexes and successive updates.

Limits for pesticides that are not listed in Table- 1 nor in EC directives are calculated using the following expression:

ADI x M ———————— MDD x 100 ADI = acceptable daily intake, as published by FAO-WHO, in milligrams per kilogram of body mass, M = body mass in kilograms (60 kg), MDD = daily dose of the drug, in kilograms.

If the drug is intended for the preparation of extracts, tinctures or other pharmaceutical forms whose preparation method modifies the content of pesticides in the finished product, the limits are calculated using the following expression:

ADI x M x E ———————— MDD x 100

72

E = extraction factor of the method of preparation, determined experimentally. Higher limits can also be authorised, in exceptional cases, especially when a plant requires a particular cultivation method or has a metabolism or a structure that gives rise to a higher than normal content of pesticides. The competent authority may grant total or partial exemption from the test when the complete history (nature and quantity of the pesticides used, date of each treatment during cultivation and after the harvest) of the treatment of the batch is known and can be checked precisely.

SAMPLING Method: For containers up to 1 kg, take one sample from the total content, thoroughly mixed, sufficient for the tests. For containers between 1 kg and 5 kg, take three samples,

equal in volume, from the upper, middle and lower parts of the container, each being sufficient to carry out the tests. Thoroughly mix the samples and take from the mixture an amount sufficient to carry out the tests. For containers of more than 5 kg, take three samples, each of at least 250 g from the upper, middle and lower parts of the container. Thoroughly

mix the samples and take from the mixture an amount sufficient to carry out the tests.

Size of sampling: If the number (n) of containers is three or fewer, take samples from each container as indicated above under Method. If the number of containers is more than three, take n+1 samples for containers as indicated under Method, rounding up to the nearest unit if necessary.

The

samples are to be analysed immediately to avoid possible degradation of the residues. If

this

is not possible, the samples are stored in airtight containers suitable for food contact, at a

temperature below 0ºC, protected from light.

Reagents: All reagents and solvents are free from any contaminants, especially pesticides

that

might interfere with the analysis. It is often necessary to use special quality solvents or, if

this

is not possible, solvents that have recently been re-distilled in an apparatus made entirely

of glass. In any case, suitable blank tests must be carried out.

73

Apparatus: Clean the apparatus and especially glassware to ensure that they are free from pesticides, for example, soak for at least 16 h in a solution of phosphate-free detergent, rinse with large quantities of distilled water R and wash with acetone and hexane or heptane.

Qualitative and quantitative analysis of pesticide residues The analytical procedures used are validated according to the regulations in force. In particular, they satisfy the following criteria:

The chosen method, especially the purification steps, are suitable for the combination

pesticide residue/substance to be analyzed and not susceptible to interference from co- extractives; the limits of detection and quantification are measured for each pesticide-matrix combination to be analysed.

Between 70 per cent to 110 per cent of each pesticide is recovered.

The repeatability of the method is not less than the values indicated

The reproducibility of the method is not less than the values indicated.

The concentration of test and reference solutions and the setting of the apparatus are such that a linear response is obtained from the analytical detector.

TEST FOR PESTICIDES

Organochlorine, organophosphorus and pyrethroid insecticides: The following methods may be used, in connection with the general method above, depending on the substance being examined, it may be necessary to modify, sometimes extensively, the procedure described hereafter. In any case, it may be necessary to use, in addition, another column with a different polarity or another detection method (mass spectrometer) or a different method (immunochemical methods) to confirm the results obtained. This procedure is valid only for the analysis of samples of vegetable drugs containing less than 15 per cent of water. Samples with a higher content of water may be dried, provided it has been shown that the drying procedure does not affect significantly the pesticide content.

EXTRACTION

To 10 g of the substance being examined, coarsely powdered, add 100 ml of acetone R and allow standing for 20 mim. Add 1 ml of a solution containing 1.8 μg/ ml of carbophenothion

74

R in toluene R. Homogenise using a high-speed blender for 3 min. Filter and wash the filter cake with two quantities, each of 25 ml, of acetone R.

Combine the filtrate and the washings and heat using a rotary evaporator at a temperature not exceeding 40ºC until the solvent has almost completely evaporated. To the residue add a few milliliters of toluene R and heat again until the acetone is completely removed. Dissolve the residue in 8 ml of toluene R. Filter through a membrane filter (45 μm), rinse the flask and the filter with toluene R and dilute to 10.0 ml with the same solvent (solution A).

PURIFICATION

Organochlorine, organophosphorus and pyrethroid insecticides: Examine by size- exclusion chromatography. The chromatographic procedure may be carried out using:

A stainless steel column 0.30 m long and 7.8 mm in internal diameter packed with styrene- divinylbenzene copolymer R (5 μm).

As mobile phase toluene R at a flow rate of 1 ml/min. Performance of the column. Inject 100 μl of a solution containing 0.5 g/l of methyl red R and 0.5 g/l of oracet blue 2R R in toluene R and proceed with the chromatography. The column is not suitable unless the colour of the eluate changes from orange to blue at an elution volume of about 10.3 ml. If necessary calibrate the column, using a solution containing, in toluene R, at a suitable concentration, the insecticide to be analysed with the lowest molecular mass (for example, dichlorvos) and that with the highest molecular mass (for example, deltamethrin). Determine which fraction of the eluate contains both insecticides.

Purification of the test solution: Inject a suitable volume of solution A (100 μl to 500 μl) and proceed with the chromatography. Collect the fraction as determined above (solution B). Organophosphorus insecticides are usually eluted between 8.8 ml and 10.9 ml. Organochlorine and pyrethroid insecticides are usually eluted between 8.5 ml and 10.3 ml.

Organochlorine and pyrethroid insecticides: In a chromatography column, 0.10 m long and 5 mm in internal diameter, introduce a piece of defatted cotton and 0.5 g of silica gel treated as follows: heat silica gel for chromatography R in an oven at 150ºC for at least 4 h.

75

Allow to cool and add dropwise a quantity of water R corresponding to 1.5 per cent of the mass of silica gel used; shake vigorously until agglomerates have disappeared and continue shaking for 2 h using a mechanical shaker. Condition the column using 1.5 ml of hexane R. Prepacked columns containing about 0.50 g of a suitable silica gel may also be used provided they are previously validated.

Concentrate solution B in a current of helium for chromatography R or oxygenfree nitrogen R almost to dryness and dilute to a suitable volume with toluene R (200 μl to 1 ml according to the volume injected in the preparation of solution B). Transfer quantitatively onto the column and proceed with the chromatography using 1.8 ml of toluene R as the mobile phase. Collect the eluate (solution C).

QUANTITATIVE ANALYSIS

1. Organophosphorus insecticides: Examine by gas chromatography,using carbophenothion R as internal standard. It may be necessary to use a second internal standard to identify possible interference with the peak corresponding to carbophenothion.

2. Test solution: Concentrate solution B in a current of helium for chromatography R almost to dryness and dilute to 100 μl with toluene R.

3. Reference solution: Prepare at least three solutions in toluene R containing the insecticides to be determined and carbophenothion at concentrations suitable for plotting a calibration curve.

4. The chromatographic procedure may be carried out using:

A fused-silica column 30 m long and 0.32 mm in internal diameter the internal wall of which is covered with a layer 0.25 μm thick of poly (dimethyl) siloxane R.

Hydrogen for chromatography R as the carrier gas. Other gases such as helium for chromatography R or nitrogen for chromatography R may also be used provided the chromatography is suitably validated.

A phosphorus-nitrogen flame-ionisation detector or a atomic emission spectrometry detector.

76

Maintaining the temperature of the column at 80ºC for 1 min, then raising it at a rate of 30ºC/min to 150ºC, maintaining at 150ºC for 3 min, then raising the temperature at a rate of 4ºC/min to 280ºC and maintaining at this temperature for 1 min and maintaining the temperature of the injector port at 250ºC and that of the detector at 275ºC. Inject the chosen volume of each solution. When the chromatograms are recorded in the prescribed conditions, the relative retention times are approximately those listed in Table-3. Calculate the content of each insecticide from the peak areas and the concentrations of the solutions.

3.2. Organochlorine and pyrethroid insecticides Examine by gas chromatography, using carbophenothion as the internal standard. It may be necessary to use a second internal standard to identify possible interference with the peak corresponding to carbophenothion.

Test solution. Concentrate solution C in a current of helium for chromatography R or oxygen-free nitrogen R almost to dryness and dilute to 500 μl with toluene R. Reference solution. Prepare at least three solutions in toluene R containing the insecticides to be determined and carbophenothion at concentrations suitable for plotting a calibration curve.

The chromatographic procedure may be carried out using:

A fused silica column 30 m long and 0.32 mm in internal diameter the internal wall of

which is covered with a layer 0.25 μm thick of poly (dimethyl) (diphenyl) siloxane R.

Hydrogen for chromatography R as the carrier gas. Other gases such as helium for

chromatography R or nitrogen for chromatography R may also be used, provided the chromatography is suitably validated.

An electron-capture detector.

A device allowing direct cold on-column injection. maintaining the temperature of the

column at 80ºC for 1 min, then raising it at a rate of 30ºC/min to 150ºC, maintaining at 150ºC

for 3 min, then raising the temperature at a rate of 4ºC/min to 280ºC and maintaining at this temperature for 1 min and maintaining the temperature of the injector port at 250ºC and that of the detector at 275ºC. Inject the chosen volume of each solution.

77

PASTE

1.

Physical properties

a) Description

b) Colour

c) Taste

2.

Total ash Incinerate about 2 to 3 g accurately weighed, of the ground drug in a

tared platinum or silica dish at a temperature not exceeding 450º until free from

carbon, cool and weigh. If a carbon free ash cannot be obtained in this way, exhaust

the charred mass with hot water, collect the residue on an ash less filter paper,

incinerate the residue and filter paper, add the filtrate, evaporate to dryness, and ignite

at a temperature not exceeding 450º. Calculate the percentage of ash with reference to

the air-dried drug.

3.

Acid insoluble ash: Boil the ash obtained for 5 minutes with 25 ml of dilute

hydrochloric acid; collect the insoluble matter in a Gooch crucible or on an ash less

filter paper, wash with hot water and ignite to constant weight. Calculate the

percentage of acid-insoluble ash with reference to the air dried drug.

4.

Total sugar: Heat 5 g with 1 ml of dilute sulphuric acid for five minutes on a

waterbath. Add 2 ml of dilute sodium hydroxide solution and 1 ml of copper sulphate

solution. A clear, blue coloured solution is produced. Continue heating on the water-

bath for five minutes. The solution remains blue and no precipitate is formed

5.

Test for Heavy Metals: Same as Oils

a) Lead

b) Cadmium

c) Mercury

d) Arsenic

6.

Total Bacterial Count /Fungal Count: Same as Oils

7.

Test for specific pathogen: Same as Oils

a) E. coli

78

b) Salmonella spp.

c) S. aureus

d) Pseudomonas aeruginosa

8.

Pesticide residue: Same as Oils

a) Organochlorine Pesticides

b) Organophosphorus Pesticides

c) Pyrethroids

TABLETS

1. Physical properties

a) Description

b) Colour

c) Odour

2. Disintegration time

Not more than 15 minutes

Not more than 60 minutes Guggulu tablets

3. ASSAY

The assay of the element being examined is tested by determing the decreased degree

of light intensity of radiation. Atomic absorption obeys the general rule for absorption

spectrophotometry. The assay is carried out by comparing the abosorbance of the test

preparation with that of the reference preparation.

Apparatus

An atomic absorption spectrophotometer consists of a light source, an atomic generator, a

monochromator and a detector system. Some are equipped with a background compensation

system and automatic sampling system, etc.

1. Light Source: A hollow-cathode discharge lamp is usually used. The cathode is made of

the element being examined.

79

2. Atomic Generator: There are four main types : flame atomizer, graphite furnace atomizer,

hydride-generated atomizer and cold vapour atomiser.

(1) Flame atomizer It mainly consists of a nebulizer and a burner. Its function is to nebulize the test solution into aerosol, which is mixed with combustion gas and the mixture is introduced into the flame generated by the burner. So that the substance being examined is to be dried, evaporated to form the ground state atoms of the element being examined. The burning flame is generated by different mixtures of gases; acetylene-air is mostly used. By modifying the proportion of combustion gas, the temperature of the flame can be controlled and a better stability and a better sensitivity can be obtained.

(2) Furnace atomizer It consists of electric furnace and a power supply. Its function is to dry and incinerate the substance being examined. During the stage of high temperature atomization, the ground state atoms of the element being examined are to be formed. Graphite is commonly used as the heater. Protection gas is introduced into the furnace to avoid oxidation and used to transfer the sample vapor.

(3) Hydride-generated atomizer It consists of hydride generator and atomic absorption cell. It is used for the determination of the elements such as arsenic, selenium, stannum and antimony etc. Its function is to reduce the element to be examined in acidic medium to the low-boiling and easily pyrolyzed hydride. And then the hydride is swept by a stream of carrier gas into the atomic absorption cell which consists of quartz tube and heater etc., in which the hydride is pyrolyzed by heating to form the ground-state atom.

(4) Cold vapor atomizer It consists of a mercury vapor atomizer and an absorption cell. It is suitable for the determination of mercury. Its function is to reduce the mercuric ion into mercury vapor which is swept into the quartz absorption cell by carrier gas.

3. Monochromator Its function is to separate the specified wavelength radiation from the

electromagnetic radiations eradiated from the light source. The optical path of the apparatus should assure the good spectra resolution and has the ability to work well at the condition of narrow spectral band (0.2 nm). The commonly used wavelength region is 190.0-900.0 nm.

80

4. Detector system It consists of a detector, a signal processor and a recording system. It

should have relatively higher sensitivity and better stability and can follow the rapid change

of the signal absorption.

5. Background compensation system System employed for the correction of atmospheric

effects on the measuring system. Four principles can be utilized for background compensation: continuous spectrum sources (a deuterium lamp is often used in the UV region), the Zeeman Effect, the self inversion phenomena and the non resonance spectrum. In the analysis using atomic absorption spectrophotometry, the interference to the determination caused by background and other reasons should be noticed. Changes of some experimental conditions, such as the wavelength, the slit width, the atomrizing condition, etc., may affect the sensitivity, the stability and the interference. If it is flame, the suitable wavelength, slit width and flame temperature, the addition of complexing agents and releasing agents and the use of Standard addition method may eliminate interference. If it is furnace, system, the selection of suitable background compensation system and the addition of suitable matrix modifying agents, etc may remove the interference. Background compensation method shall be selected as specified in the individual monograph.

Procedure: Method (direct calibration method): Prepare not less than 3 reference solutions of the element being examined of different concentrations, covering the range recommended by the instrument manufacturer and add separately the corresponding reagents as that for the test solution and prepare the blank solution with the corresponding reagents. Measure the absorbances of the blank solution and each reference solution of different concentrations separately, record the readings and prepare a calibration curve with the average value of 3 readings of each concentration on the ordinate and the corresponding concentration on the abscissa.

Prepare a test solution of the substance being examined as specified in the monograph; adjust the concentration to fall within the concentration range of the reference solution. Measure the absorbance 3 times, record the readings and calculate the average value. Interpolate the mean value of the readings on the calibration curve to determine the concentration of the element.

81

When used in the test for impurities, prepare two test preparations of the same concentration as specified in the monograph. To one of the test preparation add an amount of the reference substance equivalent to the limit of the element specified in the monograph. Proceed as directed above and measure this solution to give an appropriate reading a; then measure the test preparation without the addition of the reference substance under the same condition and record the reading b; b is not greater than (a-b).

3. Test for heavy metals: Same as Oils

a) Lead

b) Cadmium

c) Mercury

d) Arsenic

4. Microbial contamination: Same as in Oils

5. Total bacterial count /fungal count: Same as in Oils

6. Test for specific pathogen: Same as in Oils

a) E. coli

b) Salmonella spp.

c) S.aureus

d) Pseudomonas aeruginosa

7. PESTICIDE RESIDUE: Same as in Oils

a)

Organochlorine pesticides

b)

Organophosphorus pesticides

c)

Pyrethroids

POWDER

1.

Physical properties:

a) Description

b) Colour

c) Odour

d) Taste

82

2.

ASSAY OF ELEMENT (S): Same as in paste

3.

LOSS ON DRYING AT 105 ºC:

Procedure set forth here determines the amount of volatile matter (i.e., water drying off from the drug). For substances appearing to contain water as the only volatile constituent, the procedure given below, is appropriately used.

Place about 10 g of drug (without preliminary drying) after accurately weighing (accurately weighed to within 0.01 g) it in a tarred evaporating dish. For example, for underground or un powdered drug, prepare about 10 g of the sample by cutting shredding so that the parts are about 3 mm in thickness.

Seeds and fruits, smaller than 3 mm should be cracked. Avoid the use of high speed mills in preparing the samples, and exercise care that no appreciable amount of moisture is lost during preparation and that the portion taken is representative of the Official sample. After placing the above said amount of the drug in the tarred evaporating dish dry at 105º for 5 hours, and weigh. Continue the drying and weighing at one hour interval until difference between two successive weightings corresponds to not more than 0.25 per cent. Constant weight is reached when two consecutive weightings after drying for 30 minutes and cooling for 30 minutes in a desiccator, show not more than 0.01 g difference.

4.

TOTAL-ASH: Incinerate about 2 to 3 g accurately weighed, of the ground drug in a tared platinum or silica dish at a temperature not exceeding 450º until free from carbon, cool and weigh. If a carbon free ash cannot be obtained in this way, exhaust the charred mass with hot water, collect the residue on an ashless filter paper, incinerate the residue and filter paper, add the filtrate, evaporate to dryness, and ignite at a temperature not exceeding 450º. Calculate the percentage of ash with reference to the air-dried drug.

5.

ACID INSOLUBLE ASH Boil the ash obtained for 5 minutes with 25 ml of dilute hydrochloric acid; collect the insoluble matter in a Gooch crucible, or on an ashless

83

filter paper, wash with hot water and ignite to constant weight. Calculate the percentage of acid-insoluble ash with reference to the air dried drug.

6.

Particle size mesh size: 125-150

7.

Test for heavy metals: Same as Oils

a) Lead

b) Cadmium

c) Mercury

d) Arsenic

8.

Microbial contamination: Same as in oils

9.

Total Bacterial Count /Fungal Count: Same as in Oils

10.

Test For Specific Pathogen: Same as in Oil

a) E. coli

b) salmonella spp.

c) s. aureus

d) pseudomonas aeruginosa

11.

PESTICIDE RESIDUE: Same as in Oils

a) Organochlorine pesticides

b) Organophosphorus pesticides

c) Pyrethroids

SYRUP

1. PHYSICAL PROPERTIES

a) Description

b) Colour

c) Odour

84

2. TOTAL ASH: Same as in tablets

3. ACID INSOLUBLE ASH: Same as in tablets

4. WATER-SOLUBLE EXTRACTIVE :The quantitative tests e.g. total ash, acid- insoluble ash, water-soluble ash, alcohol soluble extractive, water- soluble extractive, ether-soluble extractive, moisture content, volatile oil content and assays are the methods upon which the standards of Pharmacopoeia depend. The methods for assays are described in their respective monographs and for other quantitative tests, methods are not repeated in the text of monographs but only the corresponding reference of appropriate appendix is given. The analyst is not precluded from employing an alternate method in any instance if he is satisfied that the method, which he uses, will give the same result as the Pharmacopoeial Method. In suitable instances the methods of microanalysis, if of equivalent accuracy, may be substituted for the tests and assays described. However, in the event of doubt or dispute the methods of analysis of the Pharmacopoeia are alone authoritative.

5. ALCOHOL SOLUBLE EXTRACTIVE: The quantitative tests e.g. total ash, acid-insoluble ash, water-soluble ash, alcohol soluble extractive, water- soluble extractive, ether-soluble extractive, moisture content, volatile oil content and assays are the methods upon which the standards of Pharmacopoeia depend. The methods for assays are described in their respective monographs and for other quantitative tests, methods are not repeated in the text of monographs but only the corresponding reference of appropriate appendix is given. The analyst is not precluded from employing an alternate method in any instance if he is satisfied that the method, which he uses, will give the same result as the Pharmacopoeial Method. In suitable instances the methods of microanalysis, if of equivalent accuracy, may be substituted for the tests and assays described. However, in the event of doubt or dispute the methods of analysis of the Pharmacopoeia are alone authoritative.

6. pH Between 3.0 and 4.0, determined in a 5.0 per cent w/v solution.

85

7. TOTAL SUGAR CONTENT Heat 5 g with 1 ml of dilute sulphuric acid for five minutes on a waterbath. Add 2 ml of dilute sodium hydroxide solution and 1 ml of copper sulphate solution. A clear, blue coloured solution is produced. Continue heating on the water-bath for five minutes. The solution remains blue and no precipitate is formed

8. VISCOSITY: Viscosity is a property of a liquid, which is closely related to the resistance to flow. In C.G.S. system, the dynamic viscosity (n) of a liquid is the tangential force in dryness per square centimeter exerted in either of the two parallel planes placed, 1 cm apart when the space between them is filled with the fluid and one of the plane is moving in its own plane with a velocity of 1 cm per second

relatively to the other. The unit of dynamic viscosity is the poise (abbreviated p). The centi poise (abbreviated cp) is 1/100th of one poise. While on the absolute scale, viscosity is measured in poise or centi poise, it is most convenient to use the kinematic scale in which the units are stokes (abbreviated S) and centi-stokes (abbreviated CS). The centistokes is 1/100th of one stoke. The kinematic viscosity of

a liquid is equal to the quotient of the dynamic viscosity and the density of the liquid at the same temperature. Viscosity of liquid may be determined by any method that will measure the resistance to shear offered by the liquid.

Absolute viscosity can be measured directly if accurate dimensions of the measuring

instruments are known but it is more common practice to calibrate the instrument with

a liquid of known viscosity and to determine the viscosity of the unknown fluid by comparison with that of the known.

9. Procedure The liquid under test is filled in a U tube viscometer in accordance with the expected viscosity of the liquid so that the fluid level stands within 0.2 mm of the filling mark of the viscometer when the capillary is vertical and the specified temperature is attained by the test liquid. The liquid is sucked or blown to the specified weight of the viscometer and the time taken for the meniscus to pass the two specified marks is measured. The kinematic viscosity in centistokes is calculated from the following equation:

86

Kinematic viscosity = kt

Where k = the constant of the viscometer tube determined by observation on liquids of known kinematic viscosity. T = time in seconds for meniscus to pass through the two specified marks.

10. TEST FOR HEAVY METALS: Same as Oils

a) Lead

b) Cadmium

c) Mercury

d) Arsenic

11. MICROBIAL CONTAMINATION: Same as in oils

12. TOTAL BACTERIAL COUNT /FUNGAL COUNT: Same as in Oils

13. TEST FOR SPECIFIC PATHOGEN: Same as in Oils

a) E. coli

b) Salmonella spp.

c) S. aureus

d) Pseudomonas aeruginosa

14. PESTICIDE RESIDUE: Same as in Oils

a) Organochlorine Pesticides

b) Organophosphorus Pesticides

c) Pyrethroids

87

OILS

RESULTS

1.

TEST FOR HEAVY METALS LEAD- Not more than 10 ppm Result : 8ppm CADMIUM- Not more than 0.3 ppm Result : 0.1ppm MERCURY- Not more than 01 ppm Result : 0.5ppm ARSENIC- Not more than 03 ppm Result : 01ppm

 

TOTAL BACTERIAL COUNT

2.

Not more than 1 x 105 CFU/gm Result : 1 x 104 CFU/gm

TOTAL FUNGAL COUNT Not more than 1 x 103 CFU/gm Result : 1 x 102 CFU/gm

 

PESTICIDE RESIDUE

3.

ORGANOCHLORINE PESTICIDES ORGANOPHOSPHORUS PESTICIDES PYRETHROIDS

Not more than 1 ppm Result : 0.8 ppm

 

TEST FOR AFLATOXINS

4.

(B1,B2,G1,G2)

Not more than B1 - 0.5 ppm Result 0.2 ppm Not more than G1 - 0.5 ppm

88

Result 0.4 ppm Not more than B2 - 0.1 ppm Result 0.73 ppm Not more

Result 0.4 ppm Not more than B2 - 0.1 ppm Result 0.73 ppm Not more than G2 - 0.1 ppm Result 0.63 ppm

PASTE

 

TEST FOR HEAVY METALS

1.

LEAD -Not more than 10 ppm

Result : 5ppm

CADMIUM- Not more than 0.3 ppm

Result : 0.1ppm

MERCURY- Not more than 01 ppm

Result : 0.93ppm

ARSENIC - Not more than 03 ppm

Result : 01 ppm

 

TOTAL BACTERIAL COUNT

2.

Not more than 1 x 105 CFU/gm

Result : 0.78 x 104 CFU/gm

TOTAL FUNGAL COUNT

Not more than 1 x 103 CFU/gm

Result : 0.58 x 103 CFU/gm

 

PESTICIDE RESIDUE

3.

ORGANOCHLORINE PESTICIDES

ORGANOPHOSPHORUS PESTICIDES

89

PYRETHROIDS

Not more than 1 ppm

Result : 0.38 ppm

POWDER

1.

PARTICLE SIZE MESH SIZE: 125-150

 

TEST FOR HEAVY METALS

2.

LEAD- Not more than 10 ppm

Result : 03 ppm

CADMIUM- Not more than 0.3 ppm

Result : 0.89 ppm

MERCURY- Not more than 01 ppm

Result : 0.97 ppm

ARSENIC-

Not more than 03 ppm

Result : 03 ppm

 

TOTAL BACTERIAL COUNT:

Not more than 1 x 105 CFU/gm

3.

Result : 0.89 x 102 CFU/gm

TOTAL FUNGAL COUNT: Not more than 1 x 103 CFU/gm

Result : 0.73 x 103 CFU/gm

 

PESTICIDE RESIDUE

4.

ORGANOCHLORINE PESTICIDES

ORGANOPHOSPHORUS PESTICIDES

90

 

PYRETHROIDS

Not more than 1 ppm

Result : 0.85 ppm

 

TEST FOR AFLATOXINS (B1,B2,G1,G2)

5.

B1 - Not more than 0.5 ppm

Result 0.25 ppm

G1 - 0.5 Not more than ppm

Result : 0.35 ppm

B2 - Not more than 0.1 ppm

Result : 0.21 ppm

G2 - Not more than 0.1 ppm

Result : 0.19 ppm

SYRUP

1.

pH - Between 3.0 and 4.0, determined in a 5.0 per cent w/v solution.

2.

TEST FOR HEAVY METALS

LEAD- Not more than 10 ppm Result : 8ppm CADMIUM- Not more than 0.3 ppm Result : 0.1ppm MERCURY- Not more than 01 ppm Result : 0.5ppm ARSENIC- Not more than 03 ppm Result : 01ppm

91

3.

TOTAL BACTERIAL COUNT

TOTAL BACTERIAL COUNT Not more than 1 x 105 CFU/gm Result : 1 x 104 CFU/gm

TOTAL FUNGAL COUNT Not more than 1 x 103 CFU/gm Result : 1 x 102 CFU/gm

4.

TOTAL BACTERIAL COUNT Not more than 1 x 105 CFU/gm Result : 1 x 104 CFU/gm

TOTAL FUNGAL COUNT Not more than 1 x 103 CFU/gm Result : (1 x 102 CFU/gm)

TABLETS

1.

TEST FOR HEAVY METALS

LEAD -Not more than 10 ppm

Result : 5ppm

CADMIUM- Not more than 0.3 ppm

Result : 0.1ppm

MERCURY- Not more than 01 ppm

Result : 0.93ppm

ARSENIC - Not more than 03 ppm

Result : 01 ppm

92

2.

TOTAL BACTERIAL COUNT

Not more than 1 x 105 CFU/gm

Result : 0.78 x 104 CFU/gm

TOTAL FUNGAL COUNT

Not more than 1 x 103 CFU/gm

Result : 0.58 x 103 CFU/gm

3.

PESTICIDE RESIDUE

ORGANOCHLORINE PESTICIDES

ORGANOPHOSPHORUS PESTICIDES

PYRETHROIDS

Not more than 1 ppm

Result : 0.38 ppm

4.

TEST FOR AFLATOXINS (B1,B2,G1,G2)

B1 - Not more than 0.5 ppm

Result 0.25 ppm

G1 - 0.5 Not more than ppm

Result : 0.35 ppm

B2 - Not more than 0.1 ppm

Result : 0.21 ppm

G2 - Not more than 0.1 ppm

Result : 0.19 ppm

93

CONCLUSIONS

Plant materials are used throughout the developed and developing world as home remedies, in over-the-counter drug products, and as raw material for the pharmaceutical industry, and they represent a substantial proportion of the global drug market. Therefore, it is essential to establish internationally recognized guidelines for assessing their quality. Certain herbs have become popular over the years, but the public, medical practitioners and the media still have a poor understanding of ayurvedic medicine. Evidence is emerging on the dangers of herbs. As in most situations, the truth lies hidden under the media hype, poorly understood science, and exaggerated claims. Seeing ayurvedic medicines as either panaceas or poisons blinds us to the reality that in most cases they are neither! Lack of experience, information, and education about herbs make consumers, physicians, and other orthodox health care provider‘s easy victims of market exploitation and ayurvedic myths.

There is no rational reason behind the tendency to equate ―natural‖ with ―harmlessness.‖ The fact that something is natural does not necessarily make it safe or effective. In addition, a lack of knowledge of phytochemistry leads to misinterpretation and misunderstanding. It is very likely that some herbs will have side effects, interact with other medications, and be toxic. Information on isolated constituents should not be applied directly to the whole herb and studies on in vitro forms should not be confused with oral administration. The gold standard for proof of efficacy for a medication is the controlled double-blind trial, which can offer proof of activity and effectiveness. In addition to this, well-designed unblended and clinical trials, epidemiological, animal, and phytochemical studies can provide useful information on the ayurvedic drug. It is not uncommon for studies to be carried out on animals and the results extrapolated to humans even though they have different metabolic processes. Many herbs have not been subjected to this type of study. We do not fully understand how many of these neither ayurvedic medicines work, nor do we know which component is pharmaceutically active. Even though ayurvedic remedies may be effective, do their benefits outweigh the risks? With rationing looming in virtually all health care systems, the question whether ayurvedic medicines can save money is important. Not all plant medicines are cheap. Botanicals are not patentable (they can be patented for use); hence ayurvedic remedies are not viable candidates for the existing drug approval processes. Pharmaceutical companies will not risk a loss, and

94

ayurvedic producers, especially in developing countries, lack the financial resources even to consider conducting research or seeking approval. In contrast to the United States, many European and Asian countries have taken a more holistic approach to researching the efficacy of ayurvedic remedies.

Companies supplying standardized extracts with the greatest degree of quality control typically offer the highest quality products. Most standardized extracts are currently made under strict guidelines set forth by individual members of the THE DEPARTMENT OF INDIAN SYSTEM OF MEDICINE & HOMEOPATHY as well as those proposed by The Department of Ayush. The The Department of Ayush production of standardized extracts serves as a model for quality control processes for all forms of ayurvedic preparations. Ayurvedic products and nutritional supplements are not the same. Thus, some manufacturers evade regulation of their safety.

The possibility of herbdrug interactions is important but ―under-research‖ is an issue. The World Health Assembly in resolutions has emphasized the need to ensure the quality of medicinal plant products by using modern control techniques and applying suitable standards. These resolutions describe a series of tests for assessing the quality of medicinal plant materials. The tests are designed primarily for use in national drug quality control laboratories in developing countries, and complement those described in the international pharmacopeia, which provide quality specifications only for the few plant materials that are included in the Protocol for Testing of Ayurvedic, Siddha and Unani Medicines by PHARMACOPEIAL LABORATORY FOR INDIAN MEDICINES. This manual does not constitute a herbal pharmacopeia, but a collection of test procedures to support the development of national standards based on local market conditions, with due regard to existing national legislation and national and regional norms.

The test procedures cannot take account of all possible impurities. Common sense and good pharmaceutical practice should be applied in deciding whether an unusual substance not detectable by the prescribed tests can be tolerated. The indian pharmacopeia provides quality specifications only for the few plant materials that are included in the WHO Model List of Essential Drugs.

95

There is a lack of open interpretation in the area of safety and efficacy, especially for bibliographic studies. Such interpretations are particularly relevant for ayurvedic medicinal products because they have been used for long periods of time, sometimes over centuries, and a wealth of literature is available. It is desirable that this documented knowledge is exploited in order to avoid unnecessary tests with animals and clinical trials. Scientific evaluation of the traditional knowledge is needed. In many societies much of the knowledge resides in the hand of the healers, where oral transmission of information is the unwritten rule. In most cases, the information is not documented. As a result, in many regions, this knowledge is endangered because the younger generation is unwilling to carry on the profession of the elders. Knowledge that has been refined over thousands of years of experimentation with ayurvedic medicine is being lost. A major research opportunity in this field would be to catalogue information on ayurvedic medicines by traditional healers in cultures where these skills are normally transmitted through an apprentice system.

Opinion about the safety, efficacy, and the appropriateness of medicinal herbs varies widely among medical and health professionals in countries where ayurvedic remedies are used. In most cases the safety and efficacy of drugs of ayurvedic origin cannot be attributed to one single chemical constituent. Various pharmaceutical particulars, including the production and collection of the starting material and the extraction procedures, need to be assessed. Some professionals, however, accept historical, empirical evidence as the only necessary criterion for the efficacy of ayurvedic medicines. Others would ban all ayurvedic remedies as dangerous or of questionable value. ayurvedic medicines have the potential for improving public health at low cost. Phytomedicines, if combined with preventive medical practice, could be a cost-effective, practical way to shift modern health care from treatment to prevention.

Manufacturers and distributors should attempt to certify that the ayurvedic medicines available to the public meet certain standards by answering questions such as: Does the product meet recognized standards of quality? Does the label accurately reflect what is in the product? Is the product reasonably free of contaminants such as heavy metals or pesticides? Was the product produced and packaged under clean and safe conditions? Good housekeeping is required to prove that a product is safe and effective. To obtain this certification, a manufacturer must submit research-based evidence that the product does what

96

it claims to do and that it does so without harming the consumer. Clinical trials should be conducted to establish facts such as average effective dose for any drug, as well as potential side effects a compound may cause. Recommendations on product information such as dosage limits and any warnings should also be supplied to the consumer.

Two paradigm shifts in medicine characterize the beginning of the twenty-first century: the gradual renunciation of the long-standing reliance on monosubstance therapy in favor of a multidrug therapy and the transition to a new kind of multitarget therapy, through which the interference of drugs with protective, repair, and immunostimulatory mechanisms of the human body, rather than with single disease-causing agents, gains more and more importance. Phytomedicine research has a good chance of contributing to these new strategies through the development of new and better drugs for an evidence-based and rational phytotherapy. One major concern will be to investigate the multivalent and multitarget actions of plant constituents and standardized extracts, with the aim of rationalizing the therapeutic superiority of many plant extracts over single isolated constituents. Phytomedicine and chemosynthetic pharmaceutical research find themselves in a race to develop new medicines, with fewer or no side effects, for therapeutic and preventive application in illness for which causality-based treatments are nonexistent or imperfect.

It has now become evident that there is need for a holistic approach to health care, and the untapped potential of traditional medicines should be utilized. However, this will not be easy, as it requires a thorough search for medicinal plants, proper guidelines for their identification, validation of the scientific methods of isolation of active ingredients, preclinical evaluation of their pharmacological and toxicological profiles, and clinical evidence of their usefulness. Clinical trials should be conducted to establish facts such as the average effective dose for any drug, as well as potential side effects a compound may cause. In short, the ayurvedic drugs need to be analyzed in the same way as any modern drug that is with randomized controlled clinical trials.

As doctors and researchers continue to explore the safety and effectiveness of ayurvedic medicines, more is learned about both their promises and their pitfalls. At the same time, legislators at the national level should continue to press for effective laws to protect consumers from potentially harmful ayurvedic drugs. In the mean time, your own scrutiny

97

and curiosity are your best protection. Quality control for efficacy and safety of ayurvedic products is of utmost importance. The assurance of the safety of a herbal drug requires monitoring of the quality of the finished product as well as the quality of the consumer information on the ayurvedic remedy.

98

BIBLIOGRAPHY

1. Manuals and handouts of SPAN Laboratories (P) Ltd.

2. The Ayurvedic Pharmacopeia of India by THE DEPARTMENT OF INDIAN SYSTEM OF MEDICINE & HOMEOPATHY.

3. Protocol for Testing of Ayurvedic, Siddha and Unani Medicines by PHARMACOPEIAL LABORATORY FOR INDIAN MEDICINES.

4. Production of ISM Drugs with Current Good Manufacturing Practices by THE DEPARTMENT OF INDIAN SYSTEM OF MEDICINE & HOMEOPATHY.

5. GMP for Botanicals: Regulatory and Quality Issues on Phytomedicines by Mukherjee P.K., Verpoorte R., Business Horizons, New Delhi, 2003.

6. Quality Control, Screening, Toxicity, and Regulation of Herbal Drugs by Wickramasinghe M. Bandaranayake.

7. Reflections on the basic concepts of Indian pharmacology by Jan Meulenbeld, 1987

8. Indian J. Exp. Biol. 1984 Articles by M. L. Sharma,N. Chandokhe Ray, B.J. Ghatak, K. S. Jamwal,O.P. Gupta,G. B. Singh,M.M. Ali, R.S. Thakur,K.L. Handa,P. R. Rao,Y. K. Sareen.

9. Sebastian Pole - Ayurvedic Medicine The Principles of Traditional Practice

10. http://www.niam.com

99