Quality Assurance of Herbal Formulations

Quality Assurance of Herbal FormulationsINTRODUCTION Herbal drugs have been used since ancient times as medicines for the treatment of a range of diseases. Medicinal plants have played a key role in world health. In spite of the great advances observed in modern medicine in recent decades, plants still make an important contribution to health care. Over the past decade, interest in drugs derived from higher plants, especially the phytotherapeutic ones, has increased expressively.1 Herbs have created in interest among the people by its clinically proven effects like immunomodulation, adaptogenic and antimutagenic etc.2 it is estimated that about 25% of all modern medicines are directly or indirectly derived from higher plant. According to the World Health Organization (WHO), because of poverty and lack of access to modern medicine, about 65-80% of the world's population which lives in developing countries depends essentially on plants for primary health care.1 Also the overuse of synthetic drugs which results in higher incidence of adverse drug reactions, have motivated the humans to go back to nature for safer remedies.2 India is the 8th largest country having a total of around 47,000 plant species, out of which more than 7,500 species have medicinal values. Among these medicinal plants only 800 species are claimed to be in use and around 120 species are used in large quantities.3 Currently the major pharmaceutical companies have demonstrated renewed interest in investigating higher plants as a source for new lead structures and also for the development of standardized phytotherapeutic agents with proved efficacy, safety and quality. 1 It is now increasingly accepted worldwide that screening natural products is a more effective strategy for discovering new chemical entities as natural product libraries have a broader distribution of molecular properties such as molecular mass, octanol-water coefficient and diversity of ring systems when compared to synthetic and combinatorial counter parts.3 The expanded use of herbal medicines worldwide has led to concerns relating to its safety, quality and effectiveness. The quality control of herbal crude drugs and there formulations is of paramount importance in justifying their acceptability in modern system of medicine. One of the major problem faced by user industry is non- availability of rigid quality control profiles and evaluationparametersforherbalformulations.4 1 Quality Assurance of Herbal Formulations HISTORY Herbal medicine is the oldest form of healthcare known to mankind. Herbs had been used by all cultures throughout history. It was an integral part of the development of modern civilization. The plants provided food, clothing, shelter, and medicine. Much of the medicinal use of plants seems to have been developed through observations of wild animals, and by trial and error. They methodically collected information on herbs and developed well-defined herbal pharmacopoeias. Indeed, well into the 20th century much of the pharmacopoeia of scientific medicine was derived from the herbal lore of native peoples. Many drugs commonly used today are of herbal origin. Indeed, about 25% of the prescription drugs dispensed in the United States contain at least one active ingredient derived from plant material. Some are made from plant extracts; others are synthesized to mimic a natural plant compound. The use of plants as medicine is older than recorded history. As mute witness to this fact marshmallow root, hyacinth, and yarrow have been found carefully tucked around the bones of a Stone Age man in Iraq. These three medicinal herbs continue to be used today. Marshmallow root is a demulcent herb, soothing to inflamed or irritated mucous membranes, such as a sore throat or irritated digestive tract. Hyacinth is a diuretic that encourages tissues to give up excess water. The first U.S. Pharmacopoeia was published in 1820. This volume included an authoritative listing of herbal drugs, with descriptions of their properties, uses, dosages, and tests of purity. It was periodically revised and became the legal standard for medical compounds in 1906. But as Western medicine evolved from an art to a science in the nineteenth century, information that had at one time been widely available became the domain of comparatively few. Once scientific methods were developed to extract and synthesize the active ingredients in plants, pharmaceutical laboratories took over from providers of medicinal herbs as the producers of drugs. The use of herbs, which for most of history had been mainstream medical practice, began to be considered unscientific, or at least unconventional, and to fall into relative obscurity. 2 Quality Assurance of Herbal Formulations Early humans recognized their dependence on nature in both health and illness. Led by instinct, taste, and experience, primitive men and women treated illness by using plants, animal parts, and minerals that were not part of their usual diet. Physical evidence of use of herbal remedies goes back some 60,000 years to a burial site of a Neanderthal man uncovered in 1960 .4 In a cave in northern Iraq, scientists found what appeared to be ordinary human bones. An analysis of the soil around the bones revealed extraordinary quantities of plant pollen that could not have been introduced accidentally at the burial site. Someone in the small cave community had consciously gathered eight species of plants to surround the dead man. Seven of these are medicinal plants still used throughout the herbal world.4 All cultures have long folk medicine histories that include the use of plants. Even in ancient cultures, people methodically and scientifically collected information on herbs and developed well-defined herbal pharmacopoeias. Indeed, well into the 20th century much of the pharmacopoeia of scientific medicine was derived from the herbal lore of native peoples. Many drugs, including strychnine, aspirin, vincristine, taxol, curare, and ergot, are of herbal origin. About one-quarter of the prescription drugs dispensed by community pharmacies in the United States contain at least one active ingredient derived from plant material. 5 Middle East medicine. The invention of writing was a focus around which herbal knowledge could accumulate and grow. The first written records detailing the use of herbs in the treatment of illness are the Mesopotamian clay tablet writings and the Egyptian papyrus. About 2000 B.C., King Assurbanipal of Sumeria ordered the compilation of the first known materia medica--an ancient form of today's United States Pharmacopoeia--containing 250 herbal drugs (including garlic, still a favorite of herbal doctors). The Ebers Papyrus, the most important of the preserved Egyptian manuscripts, was written around 1500 B.C. and includes much earlier information. It contains 876 prescriptions made up of more than 500 different substances, including many herbs.5Greece and Rome. One of the earliest materia medica was the Rhizotomikon, written by Diocles of Caryotos, a pupil of Aristotle. Unfortunately, the book is now lost. Other Greek and Roman compilations followed, but none was as important or influential as that written by Dioscorides in the 1st century A.D., better known by its Latin name De Materia Medica. This text contains 950 curative substances, of which 600 are plant products and the rest are of animal 3 Quality Assurance of Herbal Formulations or mineral origin.5 Each entry includes a drawing, a description of the plant, an account of its medicinal qualities and method of preparation, and warnings about undesirable effects. Muslim world. The Arabs preserved and built on the body of knowledge of the Greco-Roman period as they learned of new remedies from remote places. They even introduced to the West the Chinese technique of chemically preparing minerals. The principal storehouse of the Muslim materia medica is the text of Jami of Ibn Baiar (died 1248 A.D.), which lists more than 2,000 substances, including many plant products. Eventually this entire body of knowledge was reintroduced to Europe by Christian doctors traveling with the Crusaders. Indeed, during the middle Ages, trade in herbs became a vast international commerce. East India. India, located between China and the West, underwent a similar process in the development of its medicine. The healing that took place before India's Ayurvedic medical corpus was similar to that of ancient Egypt or China (i.e., sickness was viewed as a punishment from the gods for a particular sin). Ayurvedic medicine emerged during the rise of the philosophies of the Upanishads, Buddhism, and other schools of thought in India. Herbs played an important role in Ayurvedic medicine. The principal Ayurvedic book on internal medicine, the Characka Samhita, describes 582 herbs.8 By the Later Han Dynasty (25-220 A.D.), medicine had changed dramatically in China. People grew more confident of their ability to observe and understand the natural world and believed that health and disease were subject to the principles of natural order. However, herbs still played an important part in successive systems of medicine. The Classic of the Materia Medica, compiled no earlier than the 1st century A.D. by unknown authors, was the first Chinese book to focus on the description of individual herbs. It includes 252 botanical substances, 45 mineral substances, and 67 animal-derived substances. For each herb there is a description of its medicinal effect, usually in terms of symptoms. Reference is made to the proper method of preparation, and toxicities are noted.8 Since the writing of the Classic of the Materia Medica almost 2,000 years ago, the traditional Chinese materia medica has been steadily increasing in number. This increase has resulted from the integration into the official tradition of substances from China's folk medicine as well as from 4 Quality Assurance of Herbal Formulations other parts of the world. Many substances now used in traditional Chinese medicine originate in places such as Southeast Asia, India, the Middle East, and the Americas. The most recent compilation of Chinese materia medica was published in 1977. The Encyclopedia of Traditional Chinese Medicine Substances (Zhong Yao da ci dian), the culmination of a 25-year research project conducted by the Jiangsu College of New Medicine, contains 5,767 entries. 5 Quality Assurance of Herbal Formulations Here is a brief history of key dates in the development of herbal medicines: (8) 2800BC First written record of herbal medicines, the Pen Ts'ao by Shen Nung c400BC First Greek herbal written; Hippocrates develops principles of diet, exercise and happiness as the cornerstones of health c100BC First illustrated herbal produced in Greece c50AD Roman Empire spreads herbal medicine and commerce of plants around the Empire c200AD Herbal practitioner, Galen, creates system for classifying illnesses and remedies c500AD Hippocrates' principles followed in Britain by Myddfai practitioners throughout Saxon times c800AD Monks now pioneer herbal medicine with infirmaries and physic gardens at every monastery 1100sAD Arab world now major influence on medicine and healing practices. Physician Avicenna writes the Canon of Medicine 6 Quality Assurance of Herbal Formulations 1200sAD Black Death spreads across Europe; 'qualified' apothecaries try bleeding, purging, mercury and arsenic to stem the epidemic with no more success than traditional herbalists 1500sAD Henry VII promotes herbal medicine in the face of the growing number of untrained apothecaries and other 'medical practitioners' flourishing in London Various Acts of Parliament passed to introduce some regulation of medical practices including protection for 'simple herbalists' to practice without fear of prosecution 1600sAD Society sees the first two-tier health system emerge - herbs for the poor and exotics (plant, animal or mineral extracts) or 'drugs' for the rich Nicholas Culpepper writes his famous herbal: The English Physician, explaining in simple terms the practice of herbal medicine 1700sAD Preacher Charles Wesley advocates a sensible diet, good hygiene and herbal medicine as the keys to a healthy life 1800sAD Herbal medicines begin to be eclipsed by mineral-drug based treatments. With powerful drugs such as calomel (mercury) and laudanum available over the counter serious side effects begin to be documented. Albert Coffin pioneers low-cost herbal remedies using plants from his native America as well as European ones helping hundreds of working class people at his north of England practice. Burgeoning pharmaceuticals industry makes herbal medicine seem outdated. National Association of Medical Herbalists founded to defend the practice. Later to become the National Institute of Medical Herbalists 1900sAD Medicinal herbals used extensively during World War I as drugs are in short 7 Quality Assurance of Herbal Formulations supply. Post war period sees enormous expansion in the international pharmaceuticals industry and the discovery of penicillin A handful of dedicated herbalists keep the tradition alive. A Modern Herbal by Hilda Level is published. Pharmacy & Medicines Act 1941 withdraws herbal practitioner’s rights to supply patients with medicines. Public outcry ensures the Act is never enforced. After much campaigning by the NIMH, the Medicines Act in 1968 reinstates practitioners' rights and the British Herbal Medicine Association is founded. The BHMA produce the British Herbal Pharmacopoeia. Revised edition is published in 1990. Public concern starts to grow over the side effects of the 'wonder drugs' of the 1950s and their impact on the environment. 2000AD EU legislation advocates all herbal medicines should be subject to compulsory clinical testing comparable to that undertaken for conventional drugs. Thus all herbal medicines would be licensed. UK government currently considering the possible impact and public perception of this legislation. Chapter - 2 8 Quality Assurance of Herbal Formulations QUALITY ASSURANCE 9 Quality Assurance of Herbal Formulations Quality Assurance Quality of a product is a very hot topic nowadays, and especially in the pharmaceutical industry.Indeed, the regulatory authorities have paid special attention to quality in this particular industry, due to the high risk of damage of life and health of patients possible, and developed many guidelines to insure a sufficient level of quality. Quality is not any more considered achieve able by strict adherence to, and verification of, specifications of measurable parameters but has to be generated by a systematically planned and guided process. Quality is not any more the sole responsibility of a central quality department but requires the engaged participation of the entire work force.12 It is defined as the fulfillment all the requirements, legal and experience based, connected with all aspects of manufacturing of high quality herbal medicinal products. It starts from the beginning, the specification, processing and procurement of herbal starting material, follows the procedure and quality considerations surrounding the intermediates and ends with devising and monitoring the final production steps towards the final medicinal product.12 Quality assurance is therefore defined as a network. It encompasses the control and documentation mechanism, which insure, that the multitude of regulations pertaining to and used in practice of the pharmaceutical industry. Assurance of product quality depends on more than just proper sampling and adequate testing of various components and the finished dosage form. Prime responsibility of maintaining product quality during production rests with the manufacturing department. Removal of responsibility from manufacturing for producing a quality product can result in imperfect composition, such as ingredients missing , sub potent or super potent addition of ingredients , or mix up of ingredients ; mistakes in packaging or filling , such as product contamination , mislabeling , or deficient package ; and lack of conformance to product registration .Quality assurance personnel must establish control or checkpoints to monitor the quality of the product as it is processed and upon completion of the manufacture .These begin with raw materials and the component testing and include in-process , packaging ,labeling , and finished product testing as well as batch auditing and stability monitoring.13 10 Quality Assurance of Herbal Formulations In context with pharmaceutical industry quality assurance can be represented as: Quality assurance = GAP + GHP + GMP + GLP + other measures GAP = Good agriculture practices. GHP = Good harvesting practices. GMP = Good manufacturing practices. GLP = Good laboratory practices. Good agriculture practices: The guidelines for GAP of medicinal herbs is intended to apply to the growing and primary processing of such plants traded and used in therapy . Hence, it applies to the production of all plants materials used in the food, feed, medicinal, flavoring and perfume industries. The main aim in this GAP is to ensure that the plant raw material meets the demand of the consumer and the standards of the highest quality.11 Good harvesting practices: The starting materials for all phyto medicines are plant drugs. Mostly parts or plant organs of medicinally used species and usually in the dried form. According to WHO, there are 21,000 plant species as being medicinally used as plant drugs. Between 70%-90% of these are commercially obtained by collecting the drugs in the natural habit. Among these only about 50 – 100 species are cultured by PTC technique.11 11 Quality Assurance of Herbal Formulations Good manufacturing practices: To deliver to the public life saving drugs of the highest quality and purity , the pharmaceutical industry traditional has cooperate with FDA, even in recent years, when the regulatory agencies have become increasingly restrictive. This is a wide ranging concept concerning all matter that individually or collectively influence the quality of product. It is the totally of the arrangement made with the object of ensuring that product are of the quality required for their intended use. The system quality assurance appropriate to the manufacturer of the pharmaceutical product shall ensure that: (a) The pharmaceutical are designed and developed in a way that takes account of the requirements of GMP and other associated codes such as those of GLP and GCP. (b) Adequate arrangements are made for manufacturer, supply and use of correct starting and packaging materials. (c) Adequate control on starting materials, intermediate products and bulk products and other in process controls, calibration and validation are carried out. (d) The finished product is correctly processed and checked in accordance with established procedure. (e) The pharmaceutical products are not released for sale product supplied before authorized persons have certified that each production batch have been produced and controlled in accordance with the requirements of the label claim and any other provisions relevant to production, controlled and release of the pharmaceutical products.11,20 12 Quality Assurance of Herbal Formulations Good laboratory practices: It is a quality system for testing laboratory. Implementation of GLP will result in reliable data. Although US FDA GLP are applicable to non clinical laboratory studies that support or are intended to support application for research or marketing permits for Products regulated by the Food and Drug administration, but many elements of these GLP are of universal application. These are two important functions every testing laboratory will involve in via:• • Inspection and test Sampling 12 Quality assurance of herbal drugs:Quality assurance of herbal products may be assured by proper control of the herbal ingredients and by means of GMP.Some herbal products have many herbal ingredients with only small amount of individual herbs being present. Chemical and chromatographic tests are useful for developing finished products specifications. Stability and shelf life of herbal products should be established by the manufacture .There should be no difference in standard set for the quality of different dosage forms; such as tablet and capsule of herbal remedies as well as from those of other pharmaceutical preparation. In UK, for the licensed herbal remedies the European scientific cooperative for phytotherapy (Escop) monographs are an important development. In India the majority of the herbal remedies available are being marketed for a long time, in fact, for many products it may be before D and C act 1948.The condition, in other developing countries for the sale and production of herbal products are similar to UK. Quality, safety and efficacy of herbal drugs have to ensure to provide sound scientific footing to enhance consumer confidence and to improve business prospects for herbal medicines.11 13 Quality Assurance of Herbal Formulations Quality assurance and quality control confusion  Quality Assurance: A set of activities designed to ensure that the development and/or maintenance process is adequate to ensure a system will meet its objectives.  Quality Control: A set of activities designed to evaluate a developed work product. QA activities ensure that the process is defined and appropriate. Methodology and standards development are examples of QA activities. A QA review would focus on the process elements of a project - e.g., are requirements being defined at the proper level of detail. QC activities focus on finding defects in specific deliverables - e.g., are the defined requirements the right requirements Testing is one example of a QC activity, but there are others such as inspectionsThe difference is that QA is process oriented and QC is product oriented. Testing therefore is product oriented and thus is in the QC domain. Testing for quality isn't assuring quality, it's controlling it. Quality Assurance makes sure you are doing the right things, the right way.Quality Control makes sure the results of what you've done are what you expected.  The term “quality assurance” and “quality control” is sometimes used interchangeably, but there is an important difference. Quality control generally refers to testing of raw material, packaging components, and final product for conformance to established requirements .quality assurance is a term that includes quality control, but has broader meaning to include procedures, personnel training, record keeping and facility design and monitoring. The philosophy of a quality assurance program is to build quality into the product, rather than to rely only on final product testingto cull out defective product.16 14 Quality Assurance of Herbal Formulations Chapter – 3 GENERAL CONCEPT 15 Quality Assurance of Herbal Formulations GENERAL CONCEPT The following concepts are important in the development and setting of specifications. They are not universally applicable, but each should be considered in particular circumstances. 1) Characterization:Consistent quality for products of herbal origin can only be assured if the starting plant materials are defined in a rigorous and detailed manner. Characterization of a herbal substance/preparation or herbal medicinal product is therefore essential to allow specifications to be established, which are both comprehensive and relevant. a) Macroscopic/microscopic characterization: Macroscopic characterizations of medicinal plant material are based on the shape, size, color, surface characteristics, texture, fracture and appearance. • Color :- The color is of use in indicating the general origin of the drug, e.g. material derived from the aerial part of the plant is usually green and the underground plant materials is usually devoid of green color. • Size :- The length, width and thickness of the crude materials is of great importance while evaluating a crude drug. A graduate ruler in millimeter is adequate for this measurement. • Odor and Taste :- Odor and taste of a crude material are extremely sensitive criteria based on individual’s perceptions. The strength of the odor like weak, distinct, strong is first determined and then odor sensation like musty, moldy, rancid, fruity, aromatic etc.are determined. 16 Quality Assurance of Herbal Formulations • Surface characteristic – Texture and Fracture:- The texture is best examined by taking a small quantity of material and rubbing it between the thumb and forefinger, it is usually described as ‘smooth’, ‘rough’ , ‘gritty’ . Touch of the material determines its softness or hardness. Bend and rupture caused to the sample provides information of the brittleness and appearance of the fractured plane as fibrous, smooth, rough, granular, etc.All this characteristic are valuable in indicating the general type of material and the presence of more than one component.11 Microscopic characterization is mainly depends on the parts of the plants such as leaves, stems, flowers, fruits, seeds, barks, woods, underground drugs , entire organisms, unorganized drugs.15  Leaf Constants  Palisade Ratio: It is defined as average number of palisade cells beneath each epidermal cell. It can be determined with powdered drugs.  Vein – islet number: It is defined as the number of vein –islets per sq.mm of the leaf surface midway between the midrib and the margin. Levin in 1929 determined vein- islet number of several dicot leaves.  Vein- termination number: It is defined as the number of vein let termination per sq.mm of the leaf surface midway between the midrib and the margin.  Stomatal number: It is average number of stomata per sq.mm of epidermis of the leaf.5 17 Quality Assurance of Herbal Formulations  Stomatal index: It is the percentage which the number of stomata forms to the total number of epidermal cells; each stoma being counted as one cell. It is calculated by using the following equation:- S.I. = S / E + S x 100 Where, S.I. = Stomatal index S = Number of stomata per unit area E = Number of epidermal cells in the same unit area 15  Trichomes: These are another important diagnostic characters helpful in the identification of drugs and detection of adulterants.Trichomes(fig1) are the tubular elongated or glandular outgrowth of the epidermal cell.Trichomes is also called plant hairs. Depending upon the structure and the number of cells present in trichomes, they are classified as:15    Covering trichomes or non-globular trichomes or Clothing trichomes Glandular trichomes Hydathodes or special type of trichomes.15 Fig.1 Trichomes 18 Quality Assurance of Herbal Formulations  Stomata: The primary and most important function of stomata(fig2)is gaseous exchange and the secondary function is transpiration. However, it is generally observed that stomata are abundantly present in dicot leaves. Dicotyledons stomata are classified into following types depending upon the form and arrangement of subsidiary cells.15     Paracytic or rubiaceos or parallel- celled stomata Diacytic or caryophyllaceous or cross-celled stomata Anisocytic or cruciferous or unequal-celled stomata Anomocytic or ranunculaceous or irregular-celled stomata Fig.2 Stomata 19 Quality Assurance of Herbal Formulations  Quantitative Microscopy:  Lycopodium spore method It is an important analytical technique for powered drugs, especially when chemical and other methods of evaluation of crude drugs fail as accurate measure of quality. It is inexpensive technique with official status. Lycopodium spores are very characteristics in shape and appearance and exceptionally uniform in size (25µm). On an average, 94,000 spores per mg of powdered lycopodium are present.15 N x W x 94,000x 100 / S x M x P = % Purity of Drug Where, N = number of characteristic structures (e.g. starch grains)in 26 fields W =weight in mg of lycopodium taken S = no. of lycopodium spores in the same 25 fields M =weight in mg of the sample, calculated on basis of sample dried at 105 °C P =2, 86,000 incase of ginger starch grains powder 20 Quality Assurance of Herbal Formulations b) Phytochemical characterization: Analytical data on constituents including constituents with known therapeutic activity as well as compounds suitable as active markers or analytical markers include chromatographic fingerprinting. A chromatographic finger print profile represents qualitative/ quantitative determination of various components present in a complex plant extract irrespective whether or not their exact identity is known. In view of the enormous progress made during the four decades, standardization of botanical raw material in respect of active ingredient or marker substance poses no problem. The advances made in the separation science have given clear advantage to the chromatographic methods over the conventional trimetric and spetrophotometric methods. Thin layer chromatographic technique, the simplest least expensive, provides of wealth of information on the composition of medicinal plant drugs and its preparations, thus it is the technique of choice for the positive identification of the raw material. In view of the TLC’s limitations in quantitative analysis, liquid chromatography, which provides simultaneous separation and detection, is the technique of choice for quantitative determination of active ingredients or marker substances. The HPTLC technique combines selectivity and sensitivity thus providing stability indicating features. Non-chromatographic Assays (Gravimetric, Titrimetric, and Spectrophotometric) Simpler techniques that give a broader idea of different classes of compounds present in the herb or the polyherbal product being tested. Example: Total Tannins, Total glycosides, etc. Hence some of these assays are non-specific in their results yet valuable quantitative tools in the absence of marker compounds. Chromatographic Techniques (TLC, HPTLC, HPLC, GC): More specific, accurate and versatile techniques for analysing phytoconstituents present in single herbs or mixtures thereof. Usually require "marker compounds" or "reference standards" which can be procured from specialized companies or generated by us. 21 Quality Assurance of Herbal Formulations Depending on the availability of test methods, marker compounds and budget of the client, a suitable technique or mix of methods can be employed to develop a standard testing method for the study sample. This analytical protocol is then transferred to the client so that subsequent tests can be performed at any other competent and equipped lab of their choice.11 c) Impurities: Impurities can be classified as follows: -impurities arising from starting materials (active substances, excipients) and containers -process related impurities arising from the manufacturing process. • Contaminants, which are impurities such as heavy metals, pesticides, mycotoxins, fumigants as well as microbial contamination, including those arising extraneous sources, and radioactive substances, if relevant. • Degradation products, due to the particular nature of herbal medicinal product, should primarily address toxicologically relevant impurities arising from degradation of herbal substances/preparations. • Residual solvents, which are impurities arising from manufacturing processes. 22 Quality Assurance of Herbal Formulations 2) Design and development considerations: The experience and data accumulated during the development of a herbal substance/preparation or herbal medicinal product should form the basis for the setting of specifications. In general, it is only necessary to test the herbal medicinal product for quality attributes uniquely associated with the particular dosage form and the herbal substance or herbal preparation present. For e.g. it may be possible to propose excluding or replacing certain test on this basis some example is: -reduced testing for pesticides residues where a herbal substance is grown under strict organic cultivation without pesticides etc and potential contamination from adjacent plantations has been eliminated, -excluding or reducing tests for microbial limit in herbal preparations such as extracts or tinctures depending on the ethanol content if justified by scientific evidence. 3) Pharmacopoeial tests and acceptance criteria: The Indian Herbal pharmacopoeia contains important requirements pertaining to certain analytical procedures and acceptance criteria that are relevant to herbal substances, herbal preparation and their herbal medicinal products. Whenever they are appropriate, pharmacopoeial methods should be utilized. Drying and Storage of Plant Drugs Drying Drying consists of removal of sufficient moisture content of the crude drug so as to improve its quality and make it resistant to growth of microorganisms. The process adopted for drying is one of the parameters which affect the final quality of the drug. If enzyme action is to be encouraged, slow drying at a moderate temperature is necessary. If enzymatic action is not desired, drying should take place as soon as possible after collection. Drugs containing volatile oils are liable to lose their aroma if not dried or if the oil is not distilled from them immediately and all moist drugs are liable to develop mould. For these reasons, drying apparatus and stills should be3 situated as near to the growing plants as possible. This has the 23 Quality Assurance of Herbal Formulations further advantage that freightage is much reduced, as many fresh drugs contain a considerable amount of water. The duration of the drying process varies from a few hours to many weeks and in the case of open- air drying depends largely on the weather. Drying by artificial heat is more rapid than open-air drying and is often necessary in places where humidity is high. Artificial heating may be done by continuous belt driers or by means of open fires stoves or hot-water pipes. In all drying sheds there must be a space of at least 15 cm between superimposed trays and air must circulate freely17,18,19. Storage:Preservation of the plant drugs needs sound knowledge of their physical and chemical properties. Quality of drugs can be maintained, if these are preserved properly. All the drugs should be preserved in well closed and, preferably in the completely filled containers. The premises which are water-proof, fire-proof and preferably rodent-proof are ideal for storage. Radiation due to direct sun-light also causes destruction of active chemical constituents,e.g. ergot, cod liver oil and digitalis. Squill, when stored in powdered form becomes very much hygroscopic and forms rubbery mass on prolonged exposure to air. Atmospheric oxygen is also destructive to several drugs and hence these are filled completely in well closed containers, or the air in the container is replaced by inert gas like nitrogen; e.g. shark liver oil, papain, etc.17,18,19 24 Quality Assurance of Herbal Formulations PESTICIDE RESIDUES AND MICROBIAL COUNT Pesticide residue The use of pesticide in the agricultural sector has greatly reduced the presence of insects, fungi, and moulds in the plants. However, prolonged or excessive usage of pesticides on the crop ultimately toxicants the entire plant material causing several health hazards. Limits for pesticide residue should be established based on the recommendations of the Food and Agriculture Organization (FAO) and the World Health Organization (WHO). These recommended guidelines for food and animal feed provide the analytical methodology of pesticide residues.17,18 Classification of pesticide A classification based on the chemical composition and structure of the pesticide can be made as follows: • • • • • • • • Chlorinated hydrocarbons and related pesticides: Aldrin, benzene hexachloride(BHC) or hexachlorocyclohesane(HCH) , chlordane, dieldrin, endrin, heptachlor Chlorinated phenoxyalkanoic acid herbicides: 2,4-D; 2,4,5-T Organophosphorus pesticides: Carbophenothion , chlorthion coumaphos, demeton, dichlorvos, dimethoate, ethion, fenchlorphos. Carbamate insecticides: Carbaryl Dithiocarbamate fungicides: Ferbam, maneb, nabam, thiram. Inorganic pesticides: Aluminium phosphide, calcium arsenate, lead arsenate Pesticides of plant origin: Tobacco leaf and nicotine; pyrethrumflower, extract Miscellaneous: Bromopropylate. Chloropicrin, ethylene dibromide. Methods for the determination of pesticide residues Chromatography and other procedures are the most successful when determining pesticide residues. Samples are extracted by a standard procedure, impurities are removed by partition and adsorption and the presence of a moderately-broad spectrum of pesticides is measured in a single determination. However, these techniques are not universally applicable. As a result of limitations in the analytical technique and incomplete knowledge of pesticide interaction 25 Quality Assurance of Herbal Formulations with the environment, it is not yet possible to apply an integrated set of methods which will satisfy all situations.17,18 Maximum limit of residues for medicinal plant materials: The maximum residue limit(MRL) for medicinal plant materials, including their preparations such as tinctures, extracts, oils etc. should be defined within the limits of pesticide residue set by the FAO/WHO Codex. Since the medicinal plant materials are usually taken in much smaller quantities than other food products, the MRL can be calculated based on the maximum acceptable daily intake (ADI) of pesticides for humans and the maximum daily dose (MDD) of the medicinal plant material. Where the nature of the pesticide to which the plant material has been exposed is unknown , it is necessary to determine only the content of total chlorine and to base the calculation on the MRL of the most toxic chlorine- containing pesticide.18,19 4) Periodic/Skip testing: Periodic or skip testing is the performance of specified tests at release on pre-selected batches and/or at predetermined intervals, rather than on batch to batch basis. This represents a less than full schedule of testing and should therefore be justified and presented to the regulatory authority prior to implementation .This concept may be applicable to , for example ,dissolution ,residual solvents, and microbiological testing,e.g.,for solid dosage forms. This concept may therefore sometimes be implemented post approval in accordance with GMP and approval by the regulatory authority. 5) Release versus shelf life acceptance criteria: The concept of different acceptance criteria for release versus shelf life specifications applies to herbal medicinal products. This concept can also apply in exceptional cases to herbal substances and herbal preparations, if justified. It pertains to the establishment of more restrictive criteria for the release of a herbal medicinal product than are applied to the shelf life .Example where they are applicable include assay or impurity (degradation product) levels. 26 Quality Assurance of Herbal Formulations 6) In process tests: In-process tests are tests, which may be performed during the manufacturing of either the herbal preparation or herbal medicinal product, rather than as part of the formal battery of tests which are conducted prior to product release. In-process tests, which are used for the purpose of adjusting process parameters within an operating range, e.g., hardness and friability of tablet cores, which will be coated, are not included in the specification. Certain tests conducted during the manufacturing process, where the acceptance criteria are identical to or tighter than the release requirement, (e.g. of a solution) may be used to satisfy specification requirements when the tests is included in the specification. 7) Reference standard: A reference standard, or reference material, is a substance prepared for use as the standard in an assay, identification, or purity test. In the case of herbal medicinal products, the reference standard may be a botanical sample of the herbal substance, a sample preparation e.g. extract or tincture or a chemically defined substance e.g. a constituents with known therapeutic, an active marker or an analytical marker or a known impurity. The composition of reference standards of herbal substance and herbal preparations intended for use in assays should be adequately controlled and the purity of a standard should be measured by validated quantitative procedures. 27 Quality Assurance of Herbal Formulations • Herbal samples If the herbal substance is not described in the European pharmacopoeia or in another pharmacopoeia of a member state , a herbarium sample of the whole plant or part of the plant, if the whole plant is a tree etc.,must be available. To prepare a pooled sample for herbal drugs is very difficult as most of the foreign matters are adhered to the medicinal plant material, which are intrinsically non-uniform. That is why it is crucial that the size of the sample should be sufficiently large to be considered as a true representative. In the case of whole drug, a weighed quantity (50-500gm) according to the type of the drug is taken as sample.11 28 Quality Assurance of Herbal Formulations Chapter 4 HERBAL SUBSTANCES AND HERBAL PREPARATIONS Herbal Substances These are all mainly whole, fragmented or cut plants, plant parts, algae, fungi, lichen, in an unprocessed, usually dried form but sometimes fresh. Certain exudates that have not been subjected to a specific treatment are also considered to a herbal substance. Herbal substances are precisely defined by the plant part used and the botanical name according to the binomial system (genus, species, variety and author). 29 Quality Assurance of Herbal Formulations Herbal substances are a diverse range of botanical materials including leaves, herbs, roots, flowers, seeds, bark etc.A comprehensive specification must be developed for each herbal substance even if the starting material for the manufacture of the herbal preparation . In the case of fatty or essential oils used as active substances of herbal medicinal products a specification for the herbal is required unless justified. 25  Identification Tests:  Foreign matter: Foreign matter in herbal drugs consists of either parts of medicinal plant other than those named with specified limit; or it may be any organism, part or product or an organism. It may also include mineral admixtures not adhering to the medicinal plant material e.g. soil, stones, dust etc.Medicinal plant material should be entirely free from visible sign of any contamination.11 o Sampling : To prepare a pooled sample for herbal drugs is very difficult as most of the foreign matters are adhered to the medicinal plant material, which are intrinsically non-uniform. That is why it is crucial that the size of the sample should be sufficiently large to be considered as a true representative. In the case of whole drug, a weighed quantity (50-500gm) according to the type of the drug is taken as sample. o o Procedure: The specific quantity of plant material as mentioned below is spread on a thin layer of paper. To sort onto different groups of foreign matter it has to be examined either by visual inspection or by using magnifying lenses (6x or 10x) and the foreign matter are picked out and the percentage is recorded. The remainder of herbal drug sample is passed through 250 mesh sieve to make it free from dust, which is regarded as mineral admixture. The content of each group of foreign matter should be calculated in gram per 100gm of air dried sample. 30 Quality Assurance of Herbal Formulations Unless otherwise specified the quantities of samples to be taken into account to determine the foreign matters of any herbal drugs as specified WHO are as follows: Leaves, flowers, seeds and fruits Roots, rhizomes and barks Cut medicinal plant material 250 gm 500 gm 50 gm  Total ash: Ashing involves an oxidation of the components of the product. A high ash value is indicative of contamination, substitution, adulteration or carelessness in preparing the crude drug for marketing. Total ash is designed to measure the total amount of material produced after complete incineration of the ground drug at as low temperature as possible (about 450°C) to remove all the carbons. At higher temperature, the alkali chloride may be volatile and may be lost by this process. The total ash usually consists of carbonates, phosphates, silicates and silica which include both physiological ash - which is derived from the plant tissue itself and non-physiological ash - which is the residue of the adhering material to the plant surface, e.g., sand and soil.11 While determining the total ash very high temperature (>600°C) may result in the conversion of carbonates to oxides. I n that cases re-carbonation may be done by treatment of the ash with a solution of ammonium carbonate and further re drying to constant weight which give the carbonated ash. The similar treatment with dilute H2SO4 results in sulphated as where the oxides are converted to sulphates. When the same treatment is done by dilute HN03 results in nitrated ash. As recommended by the World Health Organization the total ash can be determined by the following procedure: Place about 2-4 g of ground air dried material, accurately weighed in a previously ignited and tared crucible of platinum or silica. Spread the material in an even layer and ignite it by gradually increasing the heat to 500-600°C until it is white, indicating the absence of carbon.11 Cool in a dessicator and weigh. If carbon free ash cannot be obtained in this manner, cool; the crucible and moisten the residue with about 2 ml of water or a saturated solution of 31 Quality Assurance of Herbal Formulations ammonium nitrate. Dry on a water bath, then on a hot plate and ignite to constant weight. Allow the residue to cool in a suitable dessicator for 30 minutes, and then weigh without delay. Calculate the content of total ash in mg/g of the air dried material. Indian Pharmacopoeia 1996 prescribes suitable method for the determination of ash values. Method I is for crude vegetable drugs and Method II for other substances: o Method I Unless otherwise stated in the individual monographs, weigh accurately 2 to 3 g of the air dried crude drug in the tared platinum or silica dish and incinerate at a temperature not exceeding 450°C 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 until the ash is white or nearly so. Calculate the percentage of ash with reference to the air-dried drug. o Method II Heat a silica or platinum crucible to red heat for 30 minutes; allow cooling in a dessicator and weighing. Unless otherwise specified in the individual monograph, weigh accurately about 1g of the substance being examined and evenly distribute it in the crucible. Dry at 100°C to 105°C for 1 hour and ignite to constant weight in a muffle furnace at 600° + 25°C. Allow the crucible to cool in a dessicator after each ignition. The material should not catch fire at any time during the procedure. If after prolonged ignition a carbon free ash cannot be obtained, proceed as directed in method I. Ignite to constant weight. Calculate the percentage of ash with reference to the air-dried substance.  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 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.11 32 Quality Assurance of Herbal Formulations  Water Soluble Ash Boil the ash for 5 minutes with 25 ml of water, collect insoluble matter in a Gooch crucible or on an ashless filter paper, wash with hot water, and ignite for 15 minutes at a temperature not exceeding 450°c. Subs tract the weight of the insoluble matter from the weight of the ash, the difference in weight represents the water soluble ash. Calculate the percentage of water soluble ash with reference to the air dry.11  Water Soluble Extractive This method determines the amount of active constituents in a given amount of medicinal plant material when extracted with solvents. It is employed for that material for which no chemical or biological assay method exist. As mentioned in different official books (Indian Pharmacopoeia 1996, British Pharmacopoeia 1980, British Herbal Pharmacopoeia.1990 etc.), the determination-of water soluble and alcohol soluble extractives, is used as a means of evaluating crude drugs which are not readily estimated by other means.11 Method I 5 g of the air-dried drug, coarsely powdered have to be macerated with 100 ml of water closed flask for 24 hours, shaking frequently during the first 6 hours and allowing to stand for 18 hours. Thereafter, filter rapidly taking precautions against loss of water, evaporate 25 ml of the filtrate to dryness in a tared flat-bottomed shallow dish, dry at 105°C and weigh. The percentage of water-soluble extractive with reference to the air dried drug has to be calculated. oMethod II Add 5 g of powdered drug to 50 ml of water at 80°C in a stoppered flask. Shake well and allow to stand for 10 minutes. Cool, add 2 g of kieselguhr and filter. Transfer 5 ml of the filtrate to a tared evaporating dish 7.5 cm in diameter. Evaporate the solvent on a water bath, continue drying for 30 minutes, finally dry in a steam oven for 2 hours and weigh the residue. Calculate the percentage of water-soluble extractive with reference to the air-dried drug. 33 Quality Assurance of Herbal Formulations  Alcohol Soluble Extractive The alcohol soluble extractive value is also indicative for the same purpose as water soluble extractive value. The solvent strength of alcohol varies from 20 - 95 % v/v. The solvent strength has to be chosen depending on the nature of drugs to be extracted. The extractive value varies depending on the strength of alcohol used for extraction, e.g. ginger when extracted with 90% alcohol gives an alcohol soluble extractive value of approximately 4.5% vlv, which-includes the oil and resins present in it. In case of rhubarb, 45% vlv alcohol strength is suitable to extract the anthraquinone present in it. With reference to Indian Pharmacopoeia 1996, the ethanol soluble extractive can be determined as follows: Macerate 5 g of the air dried drug, coarsely powdered, with 100 ml of alcohol of the specified strength in a closed flask for 24 hours, shaking frequently during six hours and allowing standing for eighteen hours. Filter rapidly, taking precautions against loss of solvent, evaporate 25 ml of filtrate to dryness in a tared flat bottomed shallow dish, and dry at 105°, to constant weight and weigh. Calculate the percentage of alcohol soluble extractive with reference to the air dried drug.11,25  Water Soluble Extractive Proceed as directed for the determination of Alcohol soluble extractive, using chloroform water instead of ethanol.  Ether Soluble Extractive (Fixed Oil Content) Transfer the suitably weighed quantity of the air dried, crushed drug to an extraction thimble, extract with solvent ether (or petroleum ether, b.p. 40° to 60°) in a continuous extraction apparatus (Soxhlet) for 6 hours. Filter the extract quantitatively into a tared evaporating dish 34 Quality Assurance of Herbal Formulations and evaporate off the solvent on a water bath. Dry the residue at 105° to constant weight. Calculate the percentage of ether soluble extractive with reference to the air dried drug. In the determination of all extractive values, the percentage has to be determined with respect to the air-dried material where the determination of moisture content is important. This is in contrast to some of the assay procedures on plant drugs, where, if the drug has to be dried before extraction apparatus is made for the loss on drying and the constituent content is calculated with reference to the un-dried or fresh drug. 11  Moisture content: Moisture is an inevitable component of crude drugs, which must be eliminated as far as practicable. The preparation of crude drug from the harvested drug plants involves cleaning or garbling to remove soil or other extraneous material followed by drying which plays a very important role in the quality as well as purity of the material. The objective of drying fresh material is: ℘ To aid in their preservation ℘ To "fix" their constituents, i.e., to check enzymatic or hydrolytic reactions that might alter the chemical composition of the drug ℘ To facilitate subsequent comminution (grinding into a powder) and ℘ To reduce their weight and bulk Insufficient drying favors spoilage by molds and bacteria and makes possible the enzymatic destruction of active principles. The moisture requirements for the active growth of some of the common molds and bacteria that may be found in or on drugs are relatively low. Therefore, the drying process should reduce the moisture content of the drug below this critical, or threshold level. Since the moisture requirements for enzymatic activity and that which microorganisms demand, vary not only with the species, but also with other environmental factors (e.g., temperature, oxygen and carbon dioxide tension, light etc.). It is difficult to state a precise upper limit of moisture that can be permitted in crude drugs. The USP and the NF make no commitment in this regard in most cases. However, most drugs 35 Quality Assurance of Herbal Formulations may be stored safely if the moisture content is reduced to 6 per cent or less. A notable exception is agar, for which USP permits as much as 20 per cent moisture. Not only is the ultimate dryness of the drug is important, equally important is the rate at which the moisture is removed and the conditions under which it is removed. If the rate is too slow, much spoilage may occur before the drying process is completed. Therefore, in general, drying should be accomplished as rapidly as is possible with good practice. The duration of the drying process varies from a few hours to several weeks, depending on the water content and other features of the drugs. Consideration of the time during which an elevated drying temperature is maintained is important, because destructive enzymatic reactions are accelerated by increasing the temperature, although the net effect of most such reactions commonly encountered in the preparation of crude drugs is accelerated only up to approximately 45°C. But higher temperatures shorten the time required for drying, and thus the time during which destructive reaction can occur also. The methods employed for drying are variable in detail, but they may be classified as spontaneous or as artificial. Artificial methods may be physical, which involves the use of elevated temperature and/or decreased pressure (vacuum), or the use of radiation of infrared or radio-frequency wave lengths; or they may be chemical, which involve use of the desiccants. The extractive values as described under section 9.2 determine the presence of specific component or group of specific components or contaminants in a plant drug. The moisture content of many crude drugs may be ascertained by the simple physical process of evaporation. A weighed sample of the crude herb is dried at 100°C and weighed periodical~ until no more than 0.25 percent is lost in one hour's drying. The total weight lost is expressed as a percentage of the initial weight of the sample; this figure is the moisture content of the sample. The residual moisture (if any) which cannot be driven off in this way is called "bound" water. A high-speed mill or other devices that are likely to create excessive heat I from friction should not be used for grinding or cutting samples intended for moisture determination. It is doubtful whether moisture can truly be classified with these types of constituents an contaminants, but as the procedure for the determination of moisture is analogous with the extraction methods described, it is included in this section. Excess moisture in a sample 36 Quality Assurance of Herbal Formulations suggests not only that the purchaser could be paying a high price for unwanted water, but also that the drug has been incorrectly prepared, or, subsequent to preparation, has been incorrectly stored. Excess moisture can result in the breakdown of important constituents by enzymatic activity and may encourage the growth of -yeast and fungi during storage. The results in either case will be the eventual rejection of the drug as an unsuitable material. Moisture content limits are stated in the pharmacopoeial monographs on many drugs. Other drugs have no limit statement expressed, the limit, by inference, being that the drug should be air-dried. This only requires that the drug has reached equilibrium with the surrounding humidity, which will naturally vary according to the location of the material. The significance of this standard is demonstrated in the exercises on the moisture content of starch and digitalis. Methods of determination of moisture content include the loss on drying, the volumetric azeotropic distillation method and the titrimetric Karl Fischer method. Which method is to be applied depends on the nature of the drug and its constituents. The test for loss on drying determines both water and volatile matter in the crude drug. It can be carried out either by heating at 1 00°C-1 05°C or in a desiccator’s over phosphorous pent oxide under atmospheric or reduced pressure at room temperature for specific period of time. The second method is especially useful for those materials that melt to a sticky mass at elevated temperature.11  Particle size: For some herbal substances intended for use in herbal teas or solid herbal medicinal products, particle size can have a significant effect dissolution rates, bioavailability, and/or stability. In such instances, testing for particle size distribution should be carried out using an appropriate procedure, and acceptances criteria should be provided. Particle size can also be affected the disintegration time of solid dosage forms. 37 Quality Assurance of Herbal Formulations  Contaminants: i) Inorganic impurities /Toxic metals: The need for inclusion of tests and acceptance criteria for inorganic impurities should be studied during development and based on knowledge of the plant species, its cultivation and the manufacturing process.11 ii) Determination of arsenic: The medicinal plant materials can be contaminated with arsenic and heavy metals which can be attributed to many causes including environmental pollution and traces of pesticides. As these components even in trace amounts are dangerous, they have to be removed from the herbal drugs. Limit tests for these materials have been prescribed in almost all the Pharmacopoeia through out the world. As prescribed by WHO the following procedures have been recommended for their respective limit tests 11 Limit Test for Arsenic: The amount of arsenic in the medicinal plant material is estimated by matching the depth of color with that of a standard stain. The limit test can be accomplished by using the following procedures. 38 Quality Assurance of Herbal Formulations Preparation of the sample by acid digestion: Place 3S-70 g of coarsely ground material, accurately weighed, in a kjeldahl flask, capacity 800-1000 ml. Add 10-25 ml of water and 25-50 ml of nitric acid (-1000 g/l) and then carefully add 20 ml of sulfuric acid (-1760g/l), drop by drop, until all the organic matter is destroyed. This is achieved when no further darkening of the solution is observed with continued heating, and a clear solution with copious vapors of sulfur trioxide is obtained. Cool, and add 75 ml of water and 25 ml of ammonium oxalate (25g/l). Heat again until sulfur trioxide vapors develop. Cool, transfer with the help of water to a 250 ml volumetric flask, and dilute to volume with water(1). Apparatus A suitable type of apparatus is constructed as follows. A wide mouthed bottle of about 120ml capacity is fitted with a rubber bung through which passes a glass tube. The latter, made from ordinary glass tubing, has a total length of about 200 mm and an internal diameter 01 exactly 6.5 mm (external diameter about 8 mm). The lower end of -the tube is drawn out to an internal diameter of about 1 mm, and there is a hole not less than 2 mm in diameter below the side of the tube, near the constricted part. The tube is positioned so that when the bottle contains 70 ml of liquid the constricted end is above the surface of the liquid and the hole in the side is below the bottom of the tube, with slightly rounded-off edges. One of two rubber bungs (about 25 mm x 25 mm), each with a central hole of exactly 6.5 mm diameter, is fitted at the upper end of the tube. The other bung is fitted with a piece of glass tube about 3 mm long and with an internal diameter of exactly 6.5 mm and with a similar ground surface. One end of each of the tubes is flush with the larger end of the bungs, so that when these ends are held tightly together with a rubber band or a spring clip, the openings of the two tubes meet to form a true tube. Alternatively, the two bungs may be replaced by any suitable construction satisfying the conditions described in the test.11 39 Quality Assurance of Herbal Formulations Method Moisten some cotton-wool with lead acetate (80 g/I), allow drying, and lightly packing into the tube which fits into wide-mouthed bottle to not less than 25 mm from the top. Between the flat surfaces of the tubes, place a piece of mercuric bromide paper that is large enough to cover their openings \ -15 mm x 15 mm). The mercuric bromide paper can be fitted byan1 other means provided that: The whole of the evolved gas passes through the paper The portion of the paper in contact with the gas is a circle 6.5 mm in diameter The paper is protected from sunlight during the test Place an aliquot (25-50 ml) of the solution being tested, prepared as described above, in the wide-mouthed bottle. Add 1 g of potassium iodide ASR and 10 g of granulated zinc ASR and place the prepared glass tube assembly quickly in position. Allow the reaction to proceed for 40 minutes. Compare any yellow stain that is in a similar manner with a known quantity of dilute arsenic As TS. Examine the test and standard stains without delay in daylight; the stains fade with time. The most suitable temperature for carrying out the test is generally about 40°C but, as the rate of evolution of the gas varies somewhat with different batches of granulated zinc ASR, the temperature may have to be adjusted to obtain an even evaluation of gas. The reaction may be accelerated by placing the apparatus on a warm surface, care being taken to ensure that the mercuric bromide paper remains dry throughout. Between successive tests, the tube must be washed with hydrochloric acid (- 250g/l) As TS, rinsed with water and dried.(1) 40 Quality Assurance of Herbal Formulations Limit Test for Cadmium and Lead The method of determination is left to the analyst. Nevertheless the determination must be consistent and sensitive enough to allow comparison with a reference material. The procedure for the determination of the same as recommended by the WHO is as follows:11 Apparatus The equipment consists of a digestion vessel, consisting of a vitreous silica crucible (DIN 12904), "tall form", height 62 mm, diameter 50 mm, capacity 75 ml, with a vitreous silica cover. Materials used are: Digestion mixture: 2 parts by weight of nitric acid (-1 000g/1) TS and 1 part by weight of perchloric acid (-1170g/I). Reference materials: olive leaves (Olea europaea) and hay powder. Clean scrupulously with nitric acid (-1000g/l) the digestion vessel and all other equipment to be used for the determination, rinse thoroughly several times with water and dry at 120°C. Preparation of the sample: For the wet digestion method in an open system, place 200-250 mg of air-dried plant material, accurately weighed, finally cut and homogeneously mixed, into a cleaned silica crucible. Add 1 ml of the digestion mixture, cover the crucible without exerting pressure and place it in an oven with a controlled temperature and time regulator (computer-controlled, if available). Heat slowly to 1OQoC and maintain at this temperature for up to 3 hours, then heat to 120°C and maintain at this temperature for 2 hours. Raise the temperature very slowly to 240°C, avoiding losses due to possible violent reactions especially in the temperature range of 160-200°C, and maintain at this temperature for 4 hours. Dissolve the remaining dry inorganic residue in 2.5 ml of nitric acid (-1000g/l) and use for the determination of heavy metals. Every sample should be tested in parallel with a blank. 41 Quality Assurance of Herbal Formulations Method The contents of lead and cadmium may be determined by inverse voltametry or by atomic absorption spectrophotometry. The following maximum amounts in dried plant materials, which are based on the ADI values, are proposed: Lead, 10 mg/kg Cadmium, 0.3 mg/kg Microbial limits: There may be a need to specify the total count of aerobic micro-organisms, the total count of yeasts and moulds, and the absence of specific objectionable bacteria. The section contains tests for the determination of the number of replicable microorganisms and for the demonstration of the absence of certain species of microbes in all types of pharmaceutical products and minerals, from crude starting materials to finished products. Strict adherence to aseptic working conditions must be ensured in the preparation and execution of the tests. Unless stated otherwise, ‘incubation’ means the storage of the material in an incubator at 3035 C for 24-48 hours. The word ‘growth’ is used in the sense of expressing the presence and expected multiplication of live micro - organisms.11 42 Quality Assurance of Herbal Formulations Limits of Microbial load: Microbes Aerobic Bacteria Yeast and moulds Escherichia coli Other enterobacteria Salmonella Plant material for topical Plant material for internal use Per gram 107 104 102 104 None use per gram 105 103 10 103 None Mycotoxins The potential for mycotoxins contamination should be fully considered. Where necessary suitable validated methods should be used to control potential mycotoxins and the acceptance criteria should be justified. Pesticides, fumigation agents etc Microbiological contamination and foreign materials are important quality criteria in the testing of medicinal plants. As with any other product from agricultural or wild sources, medicinal plants can be contaminated by organic substances of natural or synthetic origin , such as insects, micro-organisms, e.g. fungi and their mycotoxins , radioactive materials , fumigation residues and plant protection substances, i.e. pesticides. 43 Quality Assurance of Herbal Formulations Pesticides are simple substances or mixtures used to eliminate undesirable vegetable and animal life in agricultural and urban ecosystems. Owing to the great variability in plant chemical composition that results from factors to which plants are exposed during their growth, storage and the different stages of manipulation, characterization and /or standardization of phytopharmaceuticals are necessary. Standardization of herbal preparations should allow the knowledge of their composition and prevent, or at least make less likely, the consumption of drugs of questionable quality. Depending on the type of preparation, organoleptic features, moisture and ash content, physical properties and adulterants are check to confirm identity and determine purity.11 Other appropriate tests: Swelling Index: The dried ripe seeds of Plantago ovata, P. psylium, P. arenaria, P. indica etc. contain mucilage in the epidermis of the testa. The seeds of such types of plant may be evaluated by measuring the volume of mucilage produced in 24 hours from 1 g of the seeds. This evaluation procedure is termed as swelling factor. The swelling index is the volume in ml taken up by the swelling of 1 g of plant material under specified conditions. Its determination is based on the addition of water or a swelling agent as specified in the test procedure for each individual plant material (either whole, cut or pulverized). Using a glass-stoppered measuring cylinder, the material is shaken repeatedly for 1 hour and then allowed to stand for a required period of time. The volume of the mixture (in ml) is then read. The mixture of whole plant material with the swelling agent is easy to achieve, but cut or pulverized material requires vigorous shaking at specified intervals to, ensure even distribution of the material in the swelling agent. As this constitutes a valid parameter for the evaluation of the mucilaginous plant, the W!-IO has prescribed the following method for the determination of the swelling index.11 44 Quality Assurance of Herbal Formulations Recommended Procedure for the Determination of Swelling Index No fewer than three determinations for any given material have to be carried out simultaneously. The specified quantity of the plant material concerned is introduced, previously reduced to the required fineness and accurately weighed, into a 25 ml glass stoppered measuring cylinder. The internal diameter of the cylinder should be about 16 mm, the length of the graduated portion about 125 mm, marked in 0.2 ml divisions from 0 to 25 ml in an upwards direction. Unless otherwise indicated in the test procedure, 25 ml of water is to be added. The mixture is shaken thoroughly every 10 minutes for 1 hour. Allow to stand for 3 hours at room temperature or as specified. The volume (in ml) has to be measured which is occupied by the plant material, including any sticky mucilage. The mean value of the individual determinations is calculated relating to 1 g of plant material.11 Assay: In the case of herbal substances with constituents of known therapeutic activity or with active markers, assays of their content are required with details of the analytical procedures. Where possible, a specific, stability- indicating procedure should include determining the content of the herbal substance. In cases where use of non –specific assays justified, other supporting analytical procedures may be used to achieve overall specificity, if required. Foaming Index:The saponins are high molecular weight containing phytoconstituents having the detergent activity. Saponins are mostly characterized based on their frothing property. Medicinal plants of different groups, especially those derived from the families Caryophyllaceae, Araliaceae, Sapindaceae, Primulaceae, and Dioscoreaceae contain saponins. 11 Recommended procedures for foaming index determination Reduce about 1gm of the plant material to a coarse powder (sieve no. 1250) , weigh accurately and transfer to a 500ml conical flask containing 100ml of boiling water. Maintain 45 Quality Assurance of Herbal Formulations at moderate boiling for 30min. Cool and filter into a 100ml volumetric flask and add sufficient water through the filter to dilute to volume. Pour the decoction in to ten stoppered test tubes (height 16cm , diameter 16mm) in successive portion in 1ml ,2ml, 3ml and adjust the volume of the liquid in each tube with water to 10ml. Stopper the tubes and shake them in a length wise motion for 15sec, two shakes per second. Allow to stand for 15min and measure the height of the foam. Calculate the foaming index by using the following formula: 1000 / A Where, A= the volume in ml of the decoction used for pre paring the dilution in the tube where foaming to a height of 1cms is observed. 46 Quality Assurance of Herbal Formulations Herbal preparations They are obtained by subjecting herbal substances to treatments such as extraction, distillation, expression, fractionation, purification, concentration or fermentation. These include comminuted or powdered herbal substances, tinctures, extracts, essential oils, expressed juices and processed excudes. Identification tests: o Water content o Residual Solvents o Inorganic impurities, toxic metals o Microbial Limits o Mycotoxins o Pesticides, Fumigation agents,etc o Assay 47 Quality Assurance of Herbal Formulations Herbal medicinal products These are any medicinal product, exclusively containing as active substances one or more herbal substances or one or more herbal preparation, or one or more herbal substances in combination with one or more such herbal preparations. The tests and acceptance should be included for particular herbal medicinal products. The specific dosage forms addressed include solid oral herbal medicinal products. Tablet (Coated and uncoated) and hard capsules: One or more of these tests may also be applicable to soft capsule and granules. a) Disintegration : The first important step toward solution is break down of the tablet into smaller particles, a process known as disintegration. Disintegration Test For Tablets Apparatus (a) A rigid basket – rack assembly supporting six cylindrical glass tubes, 77.5+ -2.5 mm long, 21.5mm in internal diameter and with a wall thickness of about 2mm. (b) The tubes are held vertically by two superimposed transparent plastic plates, 90mm in diameter and 6m thick, perforated by six holes having the same diameters as the tubes. The holes are equidistant from the centre of the plate and are equally spaced from one another. Attached to the under side of the lower plate is a piece of woven gauze made from stainless steel wire 635micro meter in diameter and having nominal mesh apertures of 2.00mm The upper plate is covered with a stainless steel disc perforated by six holes, each about 22mm in diameter, which fits over the tubes and holds them between the plastic plates. The holes coincide with those of the upper plastic plate and the upper open ends of the glass tubes. 48 Quality Assurance of Herbal Formulations (c) The plates are held rigidly in position and 7705mm apart by vertical metal rods at the periphery and a metal enable the assembly to be attached to a mechanical device capable of raising and lowering it smoothly at a constant frequency of between 28 &32 cycles per minute through a distance of 50&60 mm. (d) A cylindrical disc for each tube, each 20.7+- o.15mm in diameter & 9.5+-0.15mm thick, made of transparent plastic with relative density of 1.18 to 1.20, & pierced with 5holes, each of 2mm in diameter, 1 in the centre & the other 4 spaced equally on the circle of radius 6mm from the centre of the disc. (e) The assembly is suspended in the liquid medium in a suitable vessel, preferably a 1000ml beaker. (f) A thermostatic arrangement for heating the liquid & maintaining the temperature at 37o +-2o. Method Unless otherwise stated in the individual monograph, introduced 1tab into each tube, add a disc to each tube. Suspend the assembly in the beaker containing the specified liquid and operate the apparatus for the specified time. Remove the assembly from the liquid. The tablet pass the test if all of them are disintegrated. If 1or2 tab. failed to disintegrate repeat the test on 12 additional tab; not than 16 of the total of 18 tab. Tested disintegrate less b) Dissolution: The original rationale for using tablet disintegration tests was the fact that as the tablet breaks down into smaller particles, it offers a greater surface area to the dissolving media and therefore must be related to the availability of the drug to the body. Dissolution Test For Tablets Apparatus 1 An assembly consisting of the following: 49 Quality Assurance of Herbal Formulations (a) A cylindrical vessel, A, made of borosilicate glass or any other suitable transparent material, with a hemispherical bottom and with a nominal capacity of 1000 ml . The vessel has a flanged upper rim and is fitted with a lid that has a number of openings, one of which is central. (b) A motor with a speed regulator capable of maintaining the speed of rotation of the paddle with in 4% of that specified in the individual monograph. The motor is fitted with a stirring element which consists of a drive shaft and blade forming a paddle, B (c) A water-bath set to maintain the dissolution medium at 36.5o to 37.5o . The bath liquid is kept in constant and smooth motion during the test. The vessel is securely clamped in the water-Bath in such equipment. Method :Introduce the stated volume of the dissolution medium, free from dissolved air, in to the vessel of the apparatus. Warm the dissolution medium to between 36.5o and 37.5o.Unless otherwise stated use one tablet . When Apparatus 1 is used, allow the tablet to sink to the bottom of the vessel prior to the rotation of the paddle. A suitable device such as a wire or glass helix may be used to keep horizontal at the bottom of the vessel tablets that would otherwise float. Care should be taken to ensure that air bubbles are excluded from the surface of the tablet . Perform the analysis as directed in the individual monograph. Repeat the whole operation five times. Where two or more tablets are directed to be placed together in the apparatus, carry out six replicate tests. For each of the tablet tested, calculate the amount of dissolved active ingredient in the solution as a percentage of the stated amount. a way that the displacement vibration from other c) Hardness: Tablets require a certain amount of strength, or hardness and resistance to friability, to withstand mechanical shocks of handling in manufacture, packaging, and shipping. Historically, the strength of a tablet was determined by breaking it between the second and third fingers with the thumb acting as a fulcrum. If there was a sharp snap, the tablet was deemed to have acceptable strength. Nowadays diametric compression tests are applied by using following instruments:14 50 Quality Assurance of Herbal Formulations Monsanto tester, Strong-Cobb tester, Pfizer tester, Erweka tester etc. The Monsanto hardness tester consists of a barrel containing a compressible spring held between two plungers. The lower plunger is placed in contact with the tablet, and a zero reading is taken. The upper plunger is then forced against a spring by turning a treaded bolt until the tablet fractures. The force of fracture is recorded, and the zero force reading is deducted from it. The Strong –Cobb tester was developed about 20 years later. The original design employed a plunger activated by pumping a lever arm, which forces an anvil against a stationary platform by hydraulic pressure. The force required to fracture the tablet is read from a hydraulic gauge. The Pfizer tester was developed and made available to the industry. This tester operates on the same mechanical principle as a pair of pliers. As the plier’s handles are squeezed, the tablet is compressed between a holding anvil and a piston connected to a direct force reading gauge. 14 d) Friability: The tablet friability can be determined by Roche friabilator. The tablets are subjected to the combined effects of abrasion and shock by utilizing a plastic chamber that revolves at 25 rpm, dropping the tablets at a distance of 6 inches with each revolution. Normally, a preweighed tablet sample is placed in the friabilator, which is then operated for 100 revolutions. The tablets are then dusted and reweighed. Loss of tablet material less than 0.5 to 1.0% of the tablet weight is accepted.14 e) Weight Variation: As per USP weight variation test, 20 tablets are weighed individually; average weight is calculated, followed by comparing the individual tablet weights to the average. The tablets 51 Quality Assurance of Herbal Formulations meet the USP test if not more than 2 tablets are outside the percentage limit and if no tablet differs by more than 2 times the percentage limit.14 Table: Weight variation tolerances for uncoated tablets: S.No. 1. 2. 3. Average weight of tablets(mg) 130 or less 130- 324 More than 324 Max. % difference allowed 10 7.5 5 52 Quality Assurance of Herbal Formulations f) Moisture content: Moisture is an inevitable component of crude drugs, which must be eliminated as far as practicable. The preparation of crude drug from the harvested drug plants involves cleaning or garbling to remove soil or other extraneous material followed by drying which plays a very important role in the quality as well as purity of the material. The objective of drying freshmaterial is: 11 ℘ To aid in their preservation ℘ To "fix" their constituents, i.e., to check enzymatic or hydrolytic reactions that might alter the chemical composition of the drug ℘ To facilitate subsequent comminution (grinding into a powder) and ℘ To reduce their weight and bulk g) Uniformity of dosage units: This term include both uniformity of content and uniformity of mass. h) Microbial limits Microbial limit testing is seen as an attribute of good manufacturing practice, as well as of quality assurance. It is advisable to test the herbal product unless its components are tested before manufacture and the manufacturing process is known, through validation studies, not to carry a significant risk of microbial contamination. Where appropriate, acceptance criteria should be set for the total count of aerobic microorganisms, the total count of yeasts and moulds, and the absence of specific objectionable bacteria (e.g., Staphylococcus aureus, Escherichia coli, Salmonella, pseudomonas).Counts should be determined using phamacopoeial or other validated procedures, and a sampling frequency or time point in manufacture which is justified by data and experience. With acceptable scientific justification, it may be possible to omit microbial limit testing for solid dosage forms.11 53 Quality Assurance of Herbal Formulations Oral liquids: One or more of the following specific test will normally be applicable to oral liquids and to powders intended for reconstitution as oral liquids.22 i) Uniformity of dosage units: This term include both uniformity of content and uniformity of mass. Generally, acceptance criteria should be set for weight variation, fill volume, and/or uniformity of fill. Pharmacopoeial procedures should be used If appropriate, tests may be performed as in-process controls; however, the acceptance criteria should be included in the specification. This concept may be applied to both singledose and multiple dose packages. The dosage unit is considered to be the typical dose taken by the patient. If the actual unit as taken by patient, is controlled, it may either be measured directly or calculated, based on the total measured weight or volume of drug, divided by the total number of doses expected. If dispensing equipment (such as medicine droppers or droppers tips for bottles) is an integral part of packaging, this equipment should be used to measure the dose. Otherwise, a standard volume measure should be used. The dispensing equipment to be used is normally determined during development. For powders for reconstitution, uniformity of mass testing is generally considered acceptable.11,14 ii) pH: Acceptance criteria for pH should be provided where applicable and the proposed range justified. 54 Quality Assurance of Herbal Formulations iii) Microbial limits: Microbial limit testing is seen as an attribute of good manufacturing practice, as well as of quality assurance. It is advisable to test the herbal product unless its components are tested before manufacture and the manufacturing process is known, through validation studies, not to carry a significant risk of microbial contamination. Where appropriate, acceptance criteria should be set for the total count of aerobic microorganisms, the total count of yeasts and moulds, and the absence of specific objectionable bacteria (e.g., Staphylococcus aureus, Escherichia coli, Salmonella, pseudomonas).Counts should be determined using phamacopoeial or other validated procedures, and a sampling frequency or time point in manufacture which is justified by data and experience. With acceptable scientific justification, it may be possible to omit microbial limit testing for solid dosage forms. iv) Antimicrobial Preservative Content: For oral liquids needing an antimicrobial preservative, acceptance criteria for preservative content must be stated. These criteria should be based on the levels necessary to maintain microbiological product quality throughout the shelf-life. Releasing testing for antimicrobial preservative content should normally performed. Under certain circumstances, in-process testing may suffice in lieu of release testing. When acceptance criteria hen antimicrobial preservative content testing is performed as an inprocess test, the acceptance criteria should remain part of the specification. Antimicrobial preservative effectiveness should be demonstrated during development, during scale-up, and through shelf-life.23,24 55 Quality Assurance of Herbal Formulations v) Antioxidant preservative content: Releasing testing for antioxidant content should normally performed. Under certain circumstances, where justified by development and stability data, shelf –life testing may be unnecessary, and in-process testing may suffice in lieu of release testing. When accept antioxidant content testing is performed as an in-process test, the acceptance criteria should remain part of the specification. If only release testing is performed, this decision should be reinvestigated whenever either the manufacturing procedure or the container/closure system changes. vi) Extractables: Generally, where development and stability data show no significant evidence of extractable from the container/closure, elimination of this test may be proposed. This should be reinvestigated if the container/closure system changes. Where data demonstrate the need , tests and acceptance criteria for extractable from the container-closure system components ( e.g. , rubber stopper , cap liner , plastic bottles, etc.) are considered appropriate for oral solutions packaged in non-glass system, or in glass containers with non-glass closure. The container/closure components should be listed, and data collected for these components as early in the development process as possible. 23,24 vii) Alcohol content: Where it is declared quantitatively on the label in accordance with pertinent regulations, the alcohol content should be specified11 56 Quality Assurance of Herbal Formulations . viii) Dissolution: In addition to the attributes recommended immediately above it may be appropriate (e.g. where constituents of the herbal substance or herbal preparation are sparingly soluble) to include dissolution testing and acceptance criteria for oral suspensions and dry powder products for resuspension. Dissolution testing may be performed as in-process test, or as a release test, depending on its relevance to product performance. The discussion of dissolution for solid dosage forms, and of particle size distribution, should also be considered.14 ix) Particle Size Distribution: Quantitative acceptance criteria and a procedure fort determination of particle Size Distribution may be appropriate for oral suspensions Particle size distribution testing may be performed as in-process test, or as a release test, depending on its relevance to product performance. If these products have been demonstrated during development to have consistently rapid drug release characteristics, exclusion of a particle size distribution test from the specification may be proposed. Particle size distribution testing may also be proposed in place of dissolution testing; justification should be provided. The acceptance criteria should include acceptable Particle size distribution in terms of the percentage of total particle size ranges. The mean, upper, and/or lower particle size limits should be well defined.14 57 Quality Assurance of Herbal Formulations x) Redispersibility: For oral suspensions, which settle on storage (produce sediment) acceptance criteria for redispersibility may be appropriate. Shaking may be an appropriate test. The procedure (mechanical or manual) should be indicated. Time required to achieve resuspension by the indicated procedures should be clearly defined. Data generated during product development may be sufficient to justify skip lot testing, or elimination of this attribute from the specification.14 xi) Rheological Properties: For relatively viscous solutions or suspensions, it may be appropriate to include rheological Properties (viscosity) in the specification. The test and acceptance criteria should be stated. Data generated during product development may be sufficient to justify skip lot testing, or elimination of this attribute from the specification.14 xii) Specific Gravity: The specific gravity of a liquid is the weight of a given volume of the liquid at 25° Compared with the weight of an equal volume of water at the same temperature, all Weighing being taken in the air.11 Method: Proceed as described under Wt. per ml. Obtain the specific gravity of the liquid by Dividing the weight of the liquid contained in the pyknometer by the weight of the water Contained, both determined at 25° unless otherwise directed in the individual Monograph.(1) 58 Quality Assurance of Herbal Formulations Xiii) Reconstitution Time: Acceptance criteria for reconstitution Time be provided for dry powder products, which require reconstitution. The choice of diluents should be justified. Data generated during product development may be sufficient to justify skip lot testing, or elimination of this attribute from the specification. xiii) Water Content: For oral products requiring reconstitution, a test and acceptance criterion for water content should be proposed when an appropriate. Loss on drying is generally considered sufficient if the effect of absorbed moisture vs. water of hydration has been adequately characterized during the development of the product. In certain cases (e.g. essential-oil containing preparations) a more specific procedure (e.g., Karl Fischer titration) is required.11 59 Quality Assurance of Herbal Formulations PHYSICAL QUALITY ASSURANCE Quality assurance of phyto-pharmceuticals products has so far been discussed solely from the chemical and physiological point of view. The physical quality however plays an equally important role for the manufacturer and processor of plant extracts. Without the addition of suitable adjuvant substances, many plant extracts occur in a form, which makes further processing considerably more difficult, often even impossible. Hence , extracts of Crataegus fruits, Curcuma extracts and many others cannot be processed to more manageable dry products either by roller, belt or spray drying. One particular example is the male fern extract, which is produced as a solvent free thin extract. In all such cases the manufacturer cannot handle this without consideration addition adjuvant substances (Newall, 1996) . Before drying therefore, a proportion of Aerosil, lactose, maltodextrin, glucose syrup or starch constituting up to 50% of the end product is added to such plant extracts. As the ratio of active substances to accompanying plant substances remain unaltered here, the manufacturer has only to declare the measures he has taken.11 Quality Assurance by Cultivation and Breeding Although medicinal plant cultivation and breeding are not in the province of pharmaceuticals technology, but they may have great influence on the use of phyto -pharmaceuticals as useful medicines.GAP has a major role to play in QA based on the following parameters.  The fact that that many medicinal substances of natural origin cannot be synthesized Only with unacceptably great effort, necessitates creation of the natural starting material i.e., cultivation of the medicinal plant. 60 Quality Assurance of Herbal Formulations The unreliability of supply of drug plants gathered from the wild shows the need for their cultivation. The qualities available in the often widely scattered gathering areas are limited. Expert collectors are becoming increasingly difficult to find. All this results in the increasing occurrence of mistaken identity and adulteration of drug plant materials.  The increased demands for safety of medicines in general have also led to increasing demands for the purity and quality of phytopharmaceuticals and of the plant drug materials from which they are gathered.  Legal regulations such as the 1973 Washington protection of species agreement and the more recent 1980West German nature protection order will considerably hinder the trade and processing of wild plants.11  61 Quality Assurance of Herbal Formulations Medicinal plants must be cultivated with phytochemical aspects in view, as the success of such cultivation depends less on the quality of plants produced and much more on their quality. The active substance content of a cultivated medicinal plant can be affected by various factors:   Genetic variation and hereditary transmission of the secondary substances Morpho and ontogenic variability, i.e. differences in the active substances contents in various parts of the plant and during its growth.  Environmental influences 21: - Temperature - Rainfall - Day-length and radiation characteristics - Altitude - Atmospheric composition 62 Quality Assurance of Herbal Formulations SUMMARY 63 Quality Assurance of Herbal Formulations . SUMMARY The present work is an attempt to brief out the various quality assurances of herbal formulation and their parameters. The quality assurance parameters have been briefly accounted which are to be strictly followed for the herbal formulations, during and after the manufacturing process, for the finished products in according to ensure their efficacy, stability and safety till the shelf life of the products. A description regarding the introduction and history of herbal formulation usage is presented followed by the difference between quality assurance and quality control and general concept of quality assurance of herbal formulations like macroscopic and microscopic characterization. The identification tests for various herbal substances, herbal preparations and herbal medicinal products are also described and in brief about physical quality assurance and quality assurance by cultivation and breeding. 64 Quality Assurance of Herbal Formulations CONCLUSION 65 Quality Assurance of Herbal Formulations CONCLUSION Quality assurance of herbal products may be assured by proper control of the herbal ingredients and by means of GMP. Quality Assurance makes sure you are doing the right things, the right way. Quality Assurance is a process oriented.. It encompasses the control and documentation mechanism, which insure, that the multitude of regulations pertaining to and used in practice of the pharmaceutical industry. one can not rely upon the quality and efficacy because almost all of the formulations do not pass through appropriate quality control procedure. The main reason behind this is the unavailability of systematic quality control procedures for herbal formulations. Therefore, the pharmaceutical formulations with combinations of drugs have shown an increasing trend to counteract other symptoms specific to one drug n formulation, and hence analytical chemist will have to accept the challenge of developing reliable methods for analysis of drugs in such formulation. 66 Quality Assurance of Herbal Formulations REFRENCES 67 Quality Assurance of Herbal Formulations REFRENCES 1) Calixto J.B, “Efficacy, Safety, Quality control, marketing and regulatory guidelines for herbal medicines; Brazillian Journal of medical and Biological research, Volume 33 (2000), page no.179-189. 2) Edwin E., Sheeza E, Vaibhav J.and Shweta D., “Toxicology of herbs pharma times, Volume 37, page no. 27-29. 3) Agarwal A., “Critical Issues In quality Control Of Natural Products”, Pharma times, Volume 37, page no. 9-11. 4) Solescki R.S, A.Neanberthal Flower Berial Of Northern Iraq Science, 4th Edition 1975. 5) Bensky D., Chinese Herbal Materia Madica (Revised edition) 1993. 6) Sarsworth N.R, Higher Plants- The sleeping gained Of Drug Development, American General of Pharmaceutical Education March, April 46. 7) Ethackerknecht, Therapeutic-From the Primetives to the 20th Century New York 1973. 8) Baby K.L, U Zutschi, C.L Chopra, N.V Amla, “Picrorhiza an Ayurvedic Herb May Potentiate Photo Chemotherapy in Vitiliva. 9) Journal Of Ethicno Pharmacol 1989. 10) Oxford’s Advanced Learner’s Dictionary”, 7th Edition 729 (2005) 68 Quality Assurance of Herbal Formulations 11) Dr. Mukherjee k. pulok, “Quality Control Of Herbal Drugs”, first edition 2002, published by Business Horizons, Page no. 124, 129, 132,186, 189, 193, 214, 217, 219, 653, 658, 679, 680. 12) World Health Organization, “Quality assurance of pharmaceuticals” , 1st edition reprint 2002 , volume I , published by pharma book syndicate, Hyderabad, page no.15. 13) World Health Organization, “Quality assurance of pharmaceuticals” , 1st edition reprint 2002 , volume II , published by pharma book syndicate, Hyderabad 14) Lachman Leon, Lieberman A. Herbert, Kanig L. Joseph, “The theory and Practice of industrial pharmacy”, 3rd edition , published by Varghese Publishing House, Page no. 297, 298, 299, 300, 301, 302, 303, 804. 15) Kokate C.K., Purohit A.P., Gokhale S.B., 25th edition, December 2003, published by Nirali prakashan, Pune, Page no. 100, 101, 102, 103, 104. 16) Mosaic, Inc., Software Risk Management Services, 205 N. Michigan Ave., Suite 2211 17) Siverajan V.V and Belachandra I., “Ayurvedic Drugs And Their Plant Sources”, Oxford And IBH, publishing Co. Pvt. Ltd, New Delhi 50 (1994). 18) Maurice MIWU, Hand Book of African Medicinal Plants, CRC, press Tokyo, 263 (1963). 19) Iyergor M.A. and Nayak S.G.K, Anatomy of Crude Drugs, 8th Edition, 26 (2004). 20) Malik Vijay, “Drugs and Cosmetic Act”, 1940, 7th Edition, published by Eastern Book company, page no.407. 21) Treas and Evans W.C , “Pharmacognosy”, 14th Edition , published by Harcourt Brace and Company Asia PTE Ltd , page no. 59,60,61. 69 Quality Assurance of Herbal Formulations 22) Monomancy T. Labels’ potency claim often inaccurate, analysis finds, Los Angeles Times 1998 August31;A10 23) Herbal Roulette, consumer Reports 1995;60 (Nov):698-705 24) De Smet PAGM,Toxicological Outlook on the quality assurance of herbal remedies, Volume I , Berlin:Springer; 1992 , page no. 1-72 25) United States, “USP 25/ Nf20 2002”, First Asian Edition, published by U.S pharmacopoeia Convention, INC. 26) India, “Indian Pharmacopoeia 1996”, Volume I and II , published by the controller of publications, Delhi. 27) Eisenberg DM, Davis RB, Ettner SL, et.al Trends in alternative medicine use it the United States, 1990-1997. JAMA 1998. 28) Brevoort p. The blooming U.S botanical market :a new overview.HerbalGram 1998 Fall;44:33-48 29) John H. Book, Organic Medicinal and Pharmaceutical Chemistry, 11 thEdition, 901 (2004). 30) Chaudhri N.C, Gurbani N.K., “Pharmaceutical Chemistry”, 1st Edition, Vallabh Prakashan, page no. 187-188(1995). 31) Goyal R.K., “Quality Control of herbal and herbal mineral products”, An emerging Trend, Pharma Times 37. 32) Harnischfeger G., “Quality Assurance of herbal preparations and herbal medicinal products, Pharma city, Panama (11). 33) Huxtable Rj. The harmful potential of herbal and other plant products. Drug Safety 1990;5 (suppl 1):126-136 70 Quality Assurance of Herbal Formulations Declaration We hereby declare that the work presented below entitled “Market Survey Of Antihypertensive Drugs” is full of truth in my knowledge and belief. We are kinely abide by the rules and regulation of our institute and university. There is no parallel work is going on this topic in our institute. Amit D.Khanvilkar Aman Preet Duggal 71 Quality Assurance of Herbal Formulations 72 Quality Assurance of Herbal Formulations 73