eChapters

ABSTRACT

In vitro culture and cryopreservation techniques are complementary to ex situ conservation of plant genetic resources that are vegetatively propagated or having recalcitrant seeds. An in vitro germplasm conservation study has been conducted on Banana passionfruit (Passiflora mollissima), a plant used in rural medicine. Micropropagated plants survived up to 12 weeks in Murashige and Skoog medium supplemented with 0.5% sucrose. Cryopreservation experiments were carried out on the explants (apical and/or axillary buds), pre-treated with vitrification solutions, dimethyl sulphoxide (DMSO) and PVS2 (Plant vitrification solution-2) with or without desiccation treatment and frozen in liquid nitrogen (LN). The survival and regeneration of apical buds were higher than axillary buds when pre-treated with 15% DMSO. Treatment with 20 % PVS2 for 1 h followed by 100 % PVS2 for 20 min gave maximum shoot recovery (40%) from cryopreserved apical buds. Desiccation alone was not successful but combination of pre-treatment of PVS2 (20% for 1h and 100% for 20min) with 3 h desiccation gave 30% plant survival from frozen apical buds. Regenerated shoots were rooted on MS medium with 1.0 mgl-1 naphthalene acetic acid (NAA) and 0.2 % activated charcoal.

1. Introduction

Phytochemists and pharmacologists from all over the world obtain several patents for herbal drugs developed on the basis of the indigenous knowledge systems of different ethnic groups. Many countries rich in traditional medicinal knowledge have been conducting ethnobotanical survey of medicinal plants (Patil and Bhaskar, 2006). In India the native people have exploited a variety of herbal medicines for effective curing of various diseases. The plants used, preparation and administration of drug varies from area to area (Rajaram, 2006). The ethnobotanical research still plays its evident scientific role in stimulating further phytochemical and pharmacological studies (Leporatti and Corradi, 2001).

The objective of this study is to interact with Kani traditional healers and document their traditional knowledge. The Kani-tribals are one of the most primitive hill tribes of Western Ghats. They occupy the slopes of the Agasthiarmalai in Tirunelveli, Tamil Nadu, India. Some ethnobotanical studies have been done in this area (Anonymous, 1948-76, Chopara et al., 1956, Satyavati, 1976, Jain, 1981, Nagendra Prasad et al., 1996). The impact of deforestation, urbanization and modernization makes the tribals shifting from their natural habitats to urban areas. Documenting the traditional knowledge through ethnobotanical studies is important for the conservation of medicinal plants and traditional medicines.

ABSTRACT

Traditionally, the rural communities of Trivandrum have been preserving small tracts of virgin forests or small groves since time immemorial due to strong religious beliefs anointed with the groves. These forest patches are biodiversity rich communities and a large number of medicinal plants of the region are confined to these groves only. The indigenous people use the plants for medicinal as well as religious purposes. Over the years, such forests have been attributed as a treasure trove of threatened species and dispensary of medicinal plants. During the investigation, 132 medicinal plants belonging to 63 families and 119 genera were recorded from 25 miniature sacred forests of Trivandrum district. This study was based on medicinal plants used in traditional health care by local and indigenous people. About 80% local people use different parts of plants for various ailments. The present finding revealed that most of the plants are used for skin diseases, cold, cough and fever, etc. Leaves are predominantly used than the other plant parts. Important families within the groves were Apocynaceae, Papilionaceae, Urticaceae, Combretaceae and Euphorbiaceae. Presence of a large number of monospecific families indicates immediate need of a conservation measure to secure sustainable utilization of medicinal plants of sacred groves of Trivandrum district.

ABSTRACT

Free radical scavenging property and antimicrobial activity of Triphala, the herbal product made of equal proportion of Terminalia chebula, Terminalia bellerica and Emblica officinalis were evaluated. Ethanol extracts of the formulation were tested for its invitro antioxidant activity by using improved ABTS radical cation decolorizing assay and invitro antimicrobial activity against Streptococcus mutans (predominantly involved in bio-film formation) by both well-diffusion and turbidity method to determine its minimal inhibitory concentration. An effort was also made to correlate its antiplaque activity using an invitro assay (where conditions kept similar to oral cavity) with Triphala and two commercial toothpastes (Colgate and Pepsodent). The ethanol extract showed significant antimicrobial and antioxidant activity. Triphala showed effective antiplaque activity when compared with the other tooth pastes. The herbal extract effectively inhibited the bio-film formation and more antioxidant activity exhibited by the extract might protect the gum cells effectively from free radicals than the commercial toothpastes, having less antioxidant activity.

1. Introduction

India is an exquisite example of biodiversity. From time immemorial scholars like Charaka, Barathwaj, Athreyan, Agnivesha, Dhanyandhari, Shushruthan, Wakbadan and Bharathduja etc. have studied and explored the possibility of such a diversity for human welfare and the most conspicuous exploration in this field has lead to the discovery of so many indigenous medicinal plants that were scripted mainly in Vedas (1500 BC) that contain rich materials on herbal lore of that time. Charaka in his medical treatise – Charaka samhita delt with innumerable medicinal plants, their usage and descriptions. Besides the two above said texts, treatise like Sushruth samhita and Ashtangha hridaya samhita are a few among other precious ancient texts. Thus the gradual accumulation of practical and systematic medical knowledge lead to the formation of Ayurveda.

Ayurveda - the science of life, prevention and longevity, is the oldest and most holistic or comprehensive medical system available. It was placed in written form over 2000 years ago. Ayurveda is said to have been first complied as a text by Agnivesha (Agniveshtantra). This text was later revised by Charaka and renamed as Charaka samhita (completed by Dhridhabala). The concept of medicines envisaged in the Ayurveda comes from the monumental scripture called Ashtanga hridaya and Sahasrayoga.

ABSTRACT

Purine Nucleoside Phosphorylase (PNP) is an enzyme that functions in the purine salvage pathway by catalyzing the phosphorolysis of purine nucleoside thereby releasing the purine bases and phosphorylated sugar. Interest in PNP as a drug target arises from its role in T-cell proliferation and T-cell mediated autoimmune diseases. Recently, we have isolated the compound, coixol, from the plant Scoparia dulcis L. and its three-dimensional structure has been elucidated. As the structure of the coixol mimics the structure of PNP inhibitors, we have undertaken the molecular modeling studies of this compound with Human PNP, as a screening test for further in vivo studies. Generally PNP inhibitors have been synthesized with a ribose sugar moiety attached to them. As the plant Scoparia dulcis L. contains glucose and rhamnose (six membered ring), these sugar moieties are also included along with ribose in the presen study. In order to understand the binding mode and interactions of the isolated compound attached with different sugar groups, we have carried out the docking studies with Human PNP. The interactions between the ligand molecules and active site residues of Human PNP are similar to those of solved complexes and the energy values are also comparable, which clearly indicate that these ligands may act as inhibitors.

ABSTRACT

Phospholipase A₂ catalyzes the hydrolysis of glycerophospholipids at the sn-2 position and releases fatty acids and lysophospholipids. When the fatty acid is the arachidonic acid, a complementary metabolism leads to pro-inflammatory mediators such as prostaglandins, leukotrienes, thromboxanes and platelet activating factors. Thus, modulating pro-inflammatory lipid mediator production by inhibiting PLA₂ activity remains a potential target for the development of new drugs for the treatment of inflammatory diseases. Medicinal plants have been explored therapeutically in traditional medicines and are a valuable source for drug discovery. Molecular docking studies were carried out for anti-inflammatory compounds from various plant sources with human non-pancreatic secretory phospholipase A₂.

ABSTRACT

Diabetes is a chronic endocrinologic disorder characterized by high blood levels of glucose due to insufficient secretion of insulin by the pancreas or improper utilization of insulin by target cells. More than 120 million people from all over the globe are estimated to suffer from this malady. Many plants and their constituents have become chief ingredients of a number of pharmaceutical preparations used against diabetes. An intensive survey undertaken for over 4 years in the tribal belts of Orissa to identify the plants used in folk medicine and the methodologies employed by the herbal healers have revealed that 35 species are being employed to cure diabetes. Out of 313 patients (185 males and 128 females) interviewed, 211 (67.5%) use Gymnema sylvestre, Momordica charantia, Trigonella foenum-graecum, Tinospora cordifolia, Pterocarpus marsupium, Andrographis paniculata and Rauwolfia serpentina in one form or the other regularly. Multi-plant preparations gave 80% results whereas single plant administration gave less than 50% results. Polyherbal treatment is advocated as multi-pronged attack to achieve desired results.

ABSTRACT

Colchicine, one of the most widely used alkaloids is synthesized in Gloriosa species, a plant of great medicinal importance. The seeds, tubers and leaves of six different species of Gloriosa were phytochemically investigated for alkaloids and were found to contain colchicine and colchicoside. These alkaloids were isolated and quantified by HPLC analysis. This is the first report of alkaloids in Gloriosa plantii, Gloriosa lutea, and Gloriosa vuchuria. The colchicine and colchicoside content of the Gloriosa species was found to be organ dependent as there was statistically significant differences (P<0.05) between different organs but not among different species under study. The total alkaloids are organ as well as species dependent as there were statistically significant differences (P<0.05) between all the species as well as organs of different species of Gloriosa under study and its organs.

A new buzz-word is rapidly spreading through the manufacturing and analytical environment. Discussions on the application of Non Invasive Research namely the Near Infrared (NIR) Spectroscopy are often found on the agenda of production, project, and technical and even board meetings. Engineers and chemists are confronted more frequently with the challenge to decide whether a NIR instrument will be the solution to their production quality control and other analytical problems. Are there really amazing machines that can determine protein, fat, moisture, fibre, starch, nicotine, alcohol, sugar, amino acids and whatever analyses you require on any product from foodstuff to fertilizer and animal feeds in less than a minute and so simple to operate that even the unskilled labourer can handle it? In this article we will endeavor to answer the questions that often crop up in the evaluation of NIR as an analytical technique towards Natural Products.

ABSTRACT

Senna (Cassia angustifolia), a traditional medicinal crop in Tamil Nadu warrants the modern quality guidelines viz. Good Agricultural Practices (GAP) for marketing overseas. Buyers of developed countries insist on GAP and organic certification for medicinal plants. With this in view, an experiment was conducted at Medicinal Plants Unit, Botanic Garden, Tamil Nadu Agricultural University, Coimbatore to study the effect of various biostimulants on yield and yield attributing characters to standardize the biostimulant(s) for higher yield. The treatments included different concentrations of panchakavya (2% & 4%), moringa leaf extract (2 & 4%) and humic acid (0.2 & 0.4%). The field was laid out in randomized block design. Vermicompost @ 4 t ha^-1 was applied commonly to all the plots. Spraying with the above said chemicals was done at 15 days interval from 30 days after transplanting for four times. The observations were recorded for growth a yield characters and were statistically analyzed. The treatment 2% panchakavya + 2% moringa leaf extract + 0.2% humic acid recorded the highest value of growth and yield characters like plant height (44.58 cm), number of branches (6.1), number of pods (53.0), dry leaf yield (560.8 kg / acre) and dry pod yield (6. 72 g / plant).

ABSTRACT

A plant grows in a highly competitive environment. It is continually being threatened by other plants encroaching upon the space from which it draws its sustenance, by microorganisms, by insects, avian, reptilian or mammalian herbivores. In order to survive, each plant must draw upon a complex of defenses, which may be physical, such as Spines or Leathery Leaves or Chemical. These defenses are passive, coming into action only when the plant is attacked which repel insects or the herbivores that attempt to feed on them. Plants protect themselves from insect attack by providing a multitude of repellent chemicals including toxic and bitter alkaloids and tannins. Non-specialist insects and other animals that attempt to feed on the plant may be repelled, injured through severe gastrointestinal malfunction or even killed. The cabbage, spinach, mustard etc. are well known for their mustard pungent flavor that deters most animals from consuming it. Even most medicinal plants like Cynanchum collialatum of the family Asclepediaceae have tannins or alkaloids like “cardenolides” that repel most animals and insects. All these plants have a basic enzyme “Myrosinase” which react with these specific tannins to produce the toxic metabolite, which repels insects and animals. But caterpillars of few insects are known to tolerate these toxic alkaloids (ex: caterpillar of Striped Tiger Butterfly-Danaus genutia) and in turn sequester the toxic alkaloids into their tissues making themselves non-palatable to their enemies. We hypothesize the analysis of possible roles of toxic alkaloids isolated from these butterflies as bio-insecticides

1. Introduction

During the last decade, considerable attention has been focused on the involvement of oxygen free radical (OFR) in various diseases. Molecular oxygen, while providing an efficient energy from ingested food, results in free radical and peroxide by-products, which have high intrinsic toxicity. Numerous physiological processes in living organisms occasionally produce free radicals, reactive nitrogen species (RNS) and reactive oxygen species (ROS) as by-products. Their broad ranges of effects in biological systems have drawn the attention of many experimental works. ROS have been implicated in the pathophysiology of several clinical disorders, including ischemia, reperfusion injury, myocardial infarction, rheumatoid arthritis, neurodegenerative, atherosclerosis, acute hypertension, hemorrhagic shock and diabetes mellitus (Hemnani and Parihar, 1998).

Oxidative stress occurs in a cell or tissue when the concentration of reactive oxygen species (ROS) generated exceeds the antioxidant capability of the cell (Simic and Jovanovic, 1986). Although humans and other organisms possess antioxidant defense and repair systems to protect against oxidative damage, these systems are insufficient under certain conditions to prevent the damage totally (Chang and Miles, 1982). Hence, the oxygen free radicals mediated damage to protein, DNA and other biomolecules accumulates during the lifetime of organisms. Many synthetic antioxidant components exhibited toxic or mutagenic effects that have directed the attention to the naturally occurring antioxidants in recent years. A multitude of natural antioxidants have already been isolated from different plant materials such as oilseeds, cereal crops, vegetables, fruits, leaves, roots, spices and herbs (Ramarathnam et al., 1995).

Looking forward, it is clear that traditional and western medicine have the potential for rich collaboration that benefits both disciplines. There is growing interest in this partnership, as shown by the number of international journals now dedicated to such research, such as Journal of Ethnopharmacology, Complementary Therapies in Medicine, eCAM, Alternative Therapies in Health and Medicine, and the Journal of Alternative and Complementary Medicine. Even the WHO has taken note of the rich possibilities of the future of TM, having put out such documents as ‘WHO General Guidelines for Methodologies on Research and Evaluation of Traditional Medicine’ and ‘WHO Guidelines on Good Agricultural and Collection Practices (GACP) for Medicinal Plants’. Despite this enthusiasm, there are several areas where the international community will need to focus their attention if the goal of integrating botanical medicines for the good of human health is to be reached.

The traditional medicines are actually used in human for number of years and therefore its clinical existence comes as a presumption. However, for bringing more objectivity and also to confirm traditional claims, systematic clinical trials are necessary. In Ayurvedic medicine research, clinical experiences, observations or available data becomes a starting point. In conventional drug research it comes at the last. Thus, the drug discovery based on Ayurveda follows a ‘Reverse Pharmacology’ path546.

Nevertheless, all the critical Pharmacopoeial tests such as dissolution time, microbial, pesticide and heavy metals contamination etc. must be in accordance with global standards. It is important to ensure that all the Ayurvedic medicine manufacture is in accordance with Current Good Manufacturing Procedures for herbal products547,548. There have been concerns about quality standards and safety issues of herbal medicines549. The need for new regulations for botanical medicines has also been frequently stressed and some such regulations are coming in force in different parts of the world550,551.

Very few traditional systems have a fair level of documentations in place. Most of the little traditions, the flow of knowledge is mainly through word of mouth, however, the great traditions such as Ayurveda and Traditional Chinese Medicine do have a good level of documentation. For instance, Ayurveda means science of life in Sanskrit and aims at holistic management of health and disease. It remains one of the most ancient medical systems widely practiced in Indian subcontinent and has a sound philosophical, experiential and experimental basis541. Charak Samhita and Sushrut Samhita (100-500 B.C.) are main Ayurvedic classics, which describe of over 700 botanicals along with their classification, pharmacological and therapeutic properties542,543. A classic diagnostic text Madhav Nidana includes over 5000 signs and symptoms. During the course of evolution and years actual practice, the interpretation of such documents developed different schools of thoughts and resulted in wide variations in understandings. Similar situation also exists in other systems including TCM. These differences often raise issues related to authenticity and correctness of Traditional Knowledge (TK). Thus systematic documentation, interpretations and harmonization of concepts and practices remain a major challenge in most of the systems of TM. Many governments, professional and community organizations have undertaken such documentation and harmonization exercises. Projects like Traditional Medicine Knowledge Digital Library, AyuSoft, Triskandha Kosha and Medicinal Plants Database of FRLHT are attempting systematic documentation of Ayurveda in India544. Other databases such as The NAPRALERT (NAtural PRoducts ALERT) developed by Professor Norman Farnsworth contain bibliographic and factual data on natural products, including information on the pharmacology, biological activity, taxonomic distribution, ethno-medicine and chemistry of plant, microbial, and animal (including marine) extracts. Another project relates to the Global IP, Benefit Sharing and Traditional Medicine Database. It is being developed as a component of the Global Information Hub on Integrated Medicine from the work of the Commonwealth Working Group (CWG) on Traditional and Complementary Health Systems, now directed by the Malaysian Ministry of Health. This Information Hub is planning to create a network of legal centers, scholars and NGOs working together to develop a comprehensive legislative and policy review, information resource and exchange on issues pertaining to IP rights over traditional medical knowledge and the use of medicinal plants545. Other than such efforts, there is limited systematic documentation of TM knowledge and practices.

Genetics forms the basis of human life. All individuals have a physical and physiological self, determined by the interactions of genes and environment. Pharmacogenetics is the connotation used to study how heritability affects an individual’s response to drugs.494 The term ‘Pharmacogenomics’ represents systematic identification of all human genes, their products, interindividual variation and intraindividual variation in expression and function over time. It focuses on candidate genes and may include transcriptome and proteome information that affect drug metabolism, pharmacokinetics, and pharmacodynamics. It helps in predicting the patient’s response to a drug and to design new drugs495. Pharmacogenetics is study of single genes and the effects on drug response while pharmacogenomics is related to overall variation in genome. A gene is considered polymorphic when a mutation appears in more than 1% population, which may be present in the form of single nucleotide polymorphism (SNP), simple sequence length polymorphism (SSLP) and nucleotide addition or deletion. After the Human Genome Project, study in this field is accelerated dramatically. It was found that out of 1.4 million SNPs identified in the human genome, over 60,000 are present in the coding region496. These SNPs lead to miscoding of the various proteins that plays crucial role in the biological processes. If a point mutation is present in the gene coding region for various drug-metabolizing enzymes, drug target and drug transporters, it can lead to many non-therapeutic and unpredictable drug responses. In United States, over two million people suffer from severe adverse drug reactions per year and about 100,000 cases are life threatening497.

In the area of pharmaceutical discovery, screening methods continue to play a major role in drug discovery. The screening process is simply testing molecular entities (compounds/natural products) for activity to advance those that exhibit the desired activity. This process has evolved over time in both the testing methods and in the degree of testing. Originally, the screening was done entirely by dosing animals and studying their responses. Then came test tube experiments and now, testing is conducted either with cell lines or with purified proteins in miniaturized test tubes called microplate wells. Screening by microplate has allowed the degree of testing to increase from a very small selected set of compounds to libraries, which contain thousands of test substances. Technological improvements in many areas of science and information technology have led to the evolution of screening methods from time consuming, animal dosing studies to high throughput screening (HTS) methods that have the potential to generate very large datasets of information in a very short time. Today, developments in bioinformatics have opened up the possibility of virtual (in silico) screens for targets where there is sufficient knowledge of the target or of its ligand. High throughput screening has become a pivotal part of drug discovery for identifying potential lead molecules for specific disease targets. The success of screening is measured by several factors including the throughput and costs of the assays, but ultimately by how many of the HTS hits progress forward through the drug discovery pipeline. Miniaturization of assays by conversion from 96- to 384- to 1536- wells to well-less platforms, such as microarrays and the utilization of automated equipments are methodologies implemented to increase the throughput and reduce the cost of the screen.

The main purpose of herbal drug technology is to convert botanical materials into medicines with a focus on standardization and quality control with proper integration of modern scientific techniques and traditional knowledge. The use of chromatographic techniques and marker compounds to standardize botanical preparations has limitations because of their variable sources and chemical complexity. DNA-based molecular markers have utility in fields like taxonomy, physiology, embryology, genetics etc. DNA based techniques have widely been used for authentication of plant species of medicinal importance. Pharmacognosy mainly addresses the quality related issues using routine botanical and organoleptic parameters of crude drugs, and chemoprofiling assisted characterization with chromatographic and spectroscopic techniques. The new pharmacognosy includes all the aspects of drug development and discovery where the biotechnology driven applications play an important role. Current focus on chemotype-driven fingerprinting and related techniques require integration with genotype-driven molecular techniques so that an optimal characterization of botanical materials is possible. This review provides a brief account of various DNA based technologies that are useful in genotyping and quick identification of botanicals with suitable examples.

Review of Health Claims

Regulators of different countries have different procedures and views to review health claims of traditional medicine, herbal medicines, botanicals, functional foods, nutraceuticals, health products, diatory supplements and such other natural forms of materials intended to use for health promotion. However, if such products carry any therapeutic or health benefit claims then the regulators may provide general guidance and demand for sufficient evidence base in support of such claims. For instance, U. S. Food and Drug Administration, Center for Food Safety and Applied Nutrition (CFSAN) provides general guidance to review of health claims in functional foods and dietary supplements. Ethnopharmacological preparations involving herbal medicine and traditional medicines prepared from natural ingredients also will need similar process. The scientific review process for health claims of such products is comprehensive and mainly focuses on review of individual studies³²⁴