A New Look At Medicinal And Aromatic Plants*

kos Mth University of West Hungary, Faculty of Agriculture and Food Industry Vr 2. 9200 Mosonmagyarvr Hungary Keywords: medicinal plants, collection, cultivation, standards, safety, efficacy, and quality, trends, modern technologies, sustainability

Abstract Herbs, Medicinal and Aromatic Plants (MAPs) have maintained their traditional basic curative role while new trends seek natural alternatives with lesser side effects to using conventional drugs. Besides their similarly traditional culinary and food industry uses, MAPs are intensively consumed as food supplements (food additives). In animal husbandry, feed-additives are used to replace synthetic chemicals and production increasing hormones. Nearly unlimited and increasing huge demand have lead to the overexploitation of natural resources, thus endangering not only plant species but incomes, even livelihoods ,especially in developing countries. A New Look, a different holistic focus and R+D action is needed to sustain an energetic and socio-economically sound MAPs sector. Guided by international standards (e.g.: ISSC-MAP, GA(C)P, FairWild), the sustainable exploitation and management of MAP natural resources have become an imperative from both environment protection and socio-economic points of view. The raw material supplies should be secured by conserving and improving the germplasm of cultivated species, and using both in situ and/or ex-situ technologies. There is a strong demand on the domestication/introduction into cultivation of presently wildcrafted species. Sophisticated in vitro propagation and breeding (selection) technologies aided by advanced phytochemical and molecular biological analytical techniques can farther assist this progress There is also a need to ensure the quality of medicinal plant products by using modern control techniques and applying suitable standards. Based on the already available modern sample preparation techniques (e.g.: SPME, SFE, PLE, MAE and SME), the study of the plant metabolome has already yielded successes. Advances in plant genomics and metabolite profiling, also seem to offer unprecedented possibilities in exploring the extraordinary complexity of plant biochemical capacity. State-of the art genomics tools can be used to enhance the production of known target metabolites and/or to synthesize entire novel compounds in cultivated plant cells by the so-called combinatorial biochemistry. Ultimate goal of these efforts should be to help improve the traceability and safety (reliability) of natural products, as well as the appropriate policies and legal frameworks to guide the protection, production (including organic production), trade, and applications of medicinal and aromatic plant materials. * Invited lecture: at 28. International Horticultural Congress, Lisbon, August 22 27, 2010 to be published in Acta Horticulturae

INTRODUCTION Herbs, Medicinal and Aromatic Plants (MAPs) have been utilized in various forms, since the very early periods of mankind. Early forms as complemented by the up to-date areas of utilization constitute powerful drivers for the exploitation of these natural resources that for a long-long time had been wasted in an unrestricted manner. Todays huge demands counterpointed with the already rather limited availability and exhaustion of natural resources brings up the necessity to make a renewed approach to MAPs, to reconsider our ideas regarding R + D, production and utilization, especially from the viewpoint of sustainability. THE ROLE OF MAPS IN MODERN SOCIETIES The initial basic curative role of MAPs has been maintained due to traditional, economic and other practical reasons. With almost half of the worlds population (i.e.: three billion people) living in poverty, earning less than two US dollars per day, even to date, ca. 80 % of the worlds population still relies primarily on traditional medicines. For many, i.e. ca. 900 million people living in rural areas, deriving their income mainly from agriculture and other related activities, MAP collection and/cultivation plays an important role as an income generating factor. MAPs have maintained their importance also for the modern pharmaceutical industry. According to WHO (1998) statistics, currently one-fourth of all prescribed pharmaceuticals in industrialized countries contain compounds that take their origin, directly or indirectly (via semi-synthesis), from plants. 11% of the 252 drugs considered as basic and essential by WHO are exclusively derived from flowering plants. In the developed parts of the world, a new trend has emerged, i.e.: to seek natural alternatives to using conventional drugs, food and cosmetics, etc. with lesser side effects. As a result, MAPs are intensively consumed in the form of food supplements and food additives. Recently, the use of phytogenic feed-additives (i.e.: in animal welfare and nutrition) is also gaining on popularity (Franz et al., 2005). In view of the controversial, constantly rising, demand on raw materials and limited resources, a new approach to MAP production and exploitation is desirable. Recent achievements in (medicinal) plant research (e.g.: biology, chemical analytics), have opened up new horizons to our knowledge on MAPs, their genetic/physiological traits, etc. As a result, new methods, technologies have emerged that ultimately may modernize both production and utilization including the standardization and quality assurance of MAP derived produces. Present review is aimed at highlighting some of the main elements of MAP research, production and utilization that have been greatly affected by changes and deserve, therefore, more attention ORIGIN OF MAP RAW MATERIAL RESOURCES Wild crafted MAPs vs. biodiversity conservation The primary resource for raw materials is wild-crafting. Some over nine tenth of MAPs used are traditionally gathered in the so called Third World countries. The natural resources of MAPs are, however, limited and this has already lead to the overexploitation of natural resources endangering both the survival of an increasing number of species and valuable incomes, especially for rural households in developing countries.

The number of endangered species is increasing menacingly According to IUCN (The World Conservation Union) the greatest risk of extinction occurs in those regions of the world (e.g.: Asia, Africa, Indonesia), where medicinal plants are still widely used. A global estimate of endangered plant species (Graham, 2002) indicates that 13 % of global flora is at the risk of extinction with 22 - 47 % of the worlds plants endangered. The belated growth in international awareness about the declining supplying capacity of the world's medicinal plants, the over-harvesting of natural resources, the destructive harvesting practices accompanied by habitat loss, forest degradation of habitats, etc. have brought about alarming problems to biodiversity. The sustainable use of natural resources The imperative for the sustainable use of natural resources was first recognized by the Chiang Mai Declaration (1988) that had expressed alarm over the consequences in the loss of plant diversity. It highlighted "the urgent need for international cooperation and coordination to establish programs for the conservation of medicinal plants to ensure that adequate quantities are available for future generations. It has also called for a need to coordinate conservation actions based on both in situ and ex situ strategies. The subsequent two decades have been marked by several farther declarations and sets of recommendations calling for the Conservation and Sustainable use of biodiversity including also medicinal plants. The Convention of Biological Diversity (CBD), an international, legally binding treaty reached at the Earth Summit, in Rio de Janeiro (1992), established the following main goals: the conservation of biological diversity, the sustainable use of its components, and the fair and equitable sharing of the benefits from the use of genetic resources. The CBD coming into force in 1993, secures rights to control access to genetic resources for the countries in which those resources are located. In view of the objective to enable lesser-developed countries better benefit from their resources and traditional knowledge, the CBD rules that bio-prospectors are required to obtain informed consent to access such resources, and must share any benefits with the biodiversity-rich country. Despite of several positive changes, the target agreed by the worlds Governments in 2002, to achieve by 2010 a significant reduction of the current rate of biodiversity loss at the global, regional and national level as a contribution to poverty alleviation and to the benefit of all life on Earth, has not been met, yet (Figure 1.). METHODS/STANDARDS FOR THE SUSTAINABLE MANAGEMENT OF MAP RESOURCES Several field-based methods have been developed for the sustainable harvest, assessment and monitoring of MAPs. Limiting the harvest to a sustainable level requires an effective management system including annual harvest quotas, seasonal or geographical restrictions and restriction of harvest to particular plant parts or size classes. In many cases harvesting techniques need to be improved, since collecting methods are often crude and wasteful, resulting in loss of quality and reduction in price (Iqbal, 1993; Vantomme in Anon. 2002). In addition, clarification of the access and user rights to the resources providing MAPs is part of the essential baseline information (FAO 1995; Leaman et al. 1997; Prescott-Allen and Prescott-Allen, 1996; Schippmann, 1997; WHO, IUCN and WWF, 1993). a. International Standard on Sustainable Wild Collection of MAPs (ISSC-MAP) To date, the most widely accepted collection standard, with a focus on the species. Its mission is to ensure the long term survival of medicinal and aromatic plant populations in their natural habitats, while respecting the traditions, cultures and livelihoods of all

stakeholders if the process. Through its holistic approach, the ISSC-MAP is aimed at bridging the gap between the existing broad conservation guidelines and management plans for local conditions. It is meant to provide guidance for sustainable wild collection of MAPs and also a basis for audit and certification in wild collection (incl. the organic sector). Ultimately, the standard is expected to have several benefits: e.g.: substantially contribute to the livelihood improvement of those involved in wildcrafting of MAPs, serve as a communication tool for the industry, serve as a guideline for MAP protection, harvest, and monitoring. For collectors, it can offer both insurance against resource and market failures, and for consumers, reliability of claims about ecological and social sustainability. Most importantly, at the level of species and habitats, it can contribute to maintaining biodiversity. b. Guidelines for Good Agricultural (and Collection) Practice of MAPs (GA(C)P Good Agricultural and/or Collection Practices (e.g.: EUROPAM, 2006) for medicinal plants are the first step in quality assurance, on which the safety and efficacy of herbal medicinal products directly depend. Harvesters of wild plants must apply collection practices that address not only their need to gain economic benefits from the sale of wild-harvested plants, but also make sure that each of the collected species survives. In addition to preserving plant populations, harvest practices must also minimize damage to local habitats. These practices are also expected to play an important role in the protection of natural resources of medicinal plants for sustainable use. c. FairWild Initiated by the Swiss Import Promotion Organization (SIPPO), FairWild is a verification system that offers a comprehensive guidance framework and certification option for all sustainably collected wild plant, fungi and lichen species worldwide. FairWild Standard covers both ecological sustainability (based on the ISSC-MAP1) and aspects of fair trade as well as social sustainability. The FairWild Standard is implemented either by the FairWild Foundation (2010) and/or its partners. Although specifically designed for wild collection situations, FairWild also includes the collection of plants, lichens or fungi or parts or products thereof on cultivated land, if the target species for collection are only a by-product and not the target of cultivation. SIPPO supports small and medium-sized enterprises (SMEs) from emerging markets and markets in transition to access the Swiss and European markets. d. Domestication/Introduction into cultivation of endangered species In view of the complex and manifold possible implications (e.g.: biodiversity conservation, management and quality assurance), as well as sustainability issues, to date, MAP domestication/introduction into cultivation are increasing considered as methods that could secure the reliable raw material supplies. As a consequence, in addition to conserving and farther improving the germplasm of traditionally cultivated species, the domestication/introduction of farther crops is needed. In this process, in situ and/or ex-situ production technologies play a decisive role. Sophisticated in vitro micropropagation and breeding (selection) technologies aided by advanced phytochemical and molecular biological analytical techniques can farther assist this progress. It is a special feature of MAPs selection and breeding that besides the usual breeding goals - e.g. to increase plant yields - the quality of active principle content plays an important role. The amount and quality (composition) of these metabolites is influenced by ecological factors, it can undergo ontogenetic and diurnal changes, it can be accompanied by the

accumulation / translocation of metabolites between various organs (intra individual variability of active principles), etc. Due to the high costs of the relevant chemical analyses, the breeding of MAPs can be rather costly while frequently also the effect of influencing factors must be investigated in the course of breeding. These and also farther (here not discussed) considerations have led to the elaboration of new analytical methods with the advantage, that they can be used in the subsequent chains of quality assurance. It is well known that even to-date, quality related problems (e.g.: lack of consistency, safety and efficacy) seem to be overshadowing the potential genuine health benefits of several herbal products. One of the major causes for this can be related to the lack of simple and reliable analytical techniques and methodologies of quality assessment (Huie, 2002). TRENDS IN THE QUALITY ASSESSMENT OF MAPS According to certain authors (Nyiredy, 2002), the major goals of medicinal plant research include: a/ qualitative and quantitative assessment and analysis of plant chemical components, b/ isolation of plant derived fractions and molecules c/ optimization of the production of plant bioactive principles. In order to achieve these, the plant materials need to be made suitable for analysis by using appropriate sample-preparation techniques. Earlier, the chemical analysis of medicinal plants meant mainly the assessment of the total amount of certain groups of compounds, (e.g.: total-flavonoid content), whereas the recent results in the development of modern separation techniques, especially those of the high throughput gas-, and liquid-chromatography, have rendered possible the more accurate separation and subsequent quantitative measurement of chemical components (Bisset and Wichtl, 2001). Sample preparation is regarded as the crucial first step in the chemical study of medicinal plants, either for basic scientific (e.g. plant breeding, physiological topics) or applied (e.g.: raw materials for the pharmaceutical industry) purposes. In addition, the modern methods are also indispensible in order to improve the acceptance of MAPs, i.e.: facilitate their reliable consumption, either as home remedies (e.g.: in the form of over-the-counter drug products) or their utilization as raw materials by the pharmaceutical industry, etc. The need to ensure the quality of medicinal plant products by using modern control techniques and applying suitable standards (WHO, 1998) has been worded also by the World Health Assembly - in resolutions WHA31.33 (1978), WHA40.33 (1987) and WHA42.43 (1989). RECENT DEVELOPMENTS (METHODS) OF NATURAL PRODUCT CHEMISTRY Sample preparation techniques The elaboration of modern sample-preparation techniques, as a precondition of plant extraction and analysis, plays an important role in the efforts of ensuring and providing highquality herbal products. The modern techniques include solid phase micro extraction (SPME), supercritical-fluid extraction (SFE), pressurized-liquid extraction (PLE), microwave-assisted extraction (MAE) and surfactant-mediated extraction (SME) (Huie, 2002). Over the past 50 years, spectroscopic methods coupled with good extraction techniques have significantly contributed to the success of natural product chemistry. E.g.: in the analysis of essential oil containing herbs (where medicinal properties can be related to the volatile constituents in the plant matrix), gas chromatography (GC) is frequently used for determination of the volatile composition. The inclusion of a fractionation step before GC analysis can help avoid the disadvantages of commonly used sample-preparation techniques (e.g. distillation and liquid solvent extraction that generally require large amounts of organic

solvents and man labor). Headspace (HS) sampling, though destructive in nature, is regarded as well suited for the fractionation of volatile compounds from complex solid matrices such as plant materials. As another promising method, supercritical fluid extraction (SFE) seems to offer high selectivity, minimum degradation of thermally labile analytes and the elimination of the use of hazardous organic solvents. SFE has been used for many years for the extraction of volatile components, like essential oils and aroma compounds from plant materials, on an industrial scale. Recently, the application of this technique has started to attract wide interest for sample preparation before chromatographic analysis. Pressurized liquid extraction (PLE) using subcritical water extractant seems to be promising especially in the case of analytes with a wide range of polarity. Its main advantages are: that water is inexpensive and non-toxic, environment friendly. These techniques are not restricted to plant sources but they are also applied to microbial and even fungal sources of metabolites. Analytical vs. Rapid Analytical Methods Hardly less than a decade ago, a new array of analytical methodologies and technologies evolved and were introduced to the analysis of microbial, plant and animal metabolomes. (Metabolome = complete collections of all low molecular weight compounds in a cell). The size of plant metabolome varies greatly. It is estimated that the plant kingdom has approximately 200,000 primary and secondary metabolites (Fiehn, 2002). As plant secondary metabolites occur usually in low concentrations, these sophisticated technologies look promising and have already been successfully applied in their analytics. A rough overview of available analytical methods for the study of MAPs is given in Table 1. Above methods have several relevant advantages, among them the most important being rapidity, the ability to select high-quality single plants from populations, progenies (e.g.: crossing experiments), as well as industrial uses for quality checks and supervision, etc. Recently, Schulz (2009) gave an overview of the applications of the so called rapid analytical methods (RAM) with a focus on the efficient evaluation of plant genetic resources and breeding material of MAPs. According to this, Rapid High-Throughput (RHP) methods are capable of simultaneously characterizing several quality parameters and as such reducing sample preparation to a minimum. Vibrational spectroscopy methods (attenuated total reflectance/Fourier-transform-infrared (ATR/FT-IR), FT-Raman and near infrared (NIR) spectroscopy) have a potential for the identification and quantification of valuable, as well as carcinogenic substances (e.g.: in different basil chemotypes). The main volatile components occurring in the plant samples can be reliably determined in both isolated essential oils or solvent extracts and the air-dried herbs. MIR (Mid_Infrared Spectroscopy), NIR (Near Infrared Spectroscopy) as well as Raman sectroscopy are reported to be suitable for the rapid and reliable non-destructive discrimination of plant species or chemotypes. High throughput screening (HTS) methods. By using molecular targets, a large number of samples (up to 100,000 in 24 h) can be screened for a single activity. HTS offers the following new possibilities for natural products: rapid screening of large number of extracts (a), it is very suitable for bioassay-guided fractionation (in the past, the major bottleneck in studies of active compounds in plant extracts) (b), it has significantly increased throughput and reduced assay volumes (c). Key advances over the past few years include: new fluorescence methods, detection platforms and liquid-handling technologies.

From Metabolomics to Metabolic Engineering Cutting edge research in MAPs, nowadays, is also frequently engaged in the study of the plant metabolome. We are seeing a shift in gene studies, going from single-gene studies towards metabolic pathways, ultimately to the study of the whole-genome (OksmanCaldentey and Inz, 2004). With other words the qualitative and quantitative analysis of all metabolites combined with new and powerful tools of functional genomics can be used to elucidate biosynthetic pathways of natural products. As a precondition for Metabolic Engineering, the details involved in the biosynthesis of a natural product need to be identified at both enzyme and regulation levels. According to the general concept of metabolic engineering certain pathways within a biosynthesis network could be stimulated/favored over others (e.g. by over-expressing the crucial enzyme squalene synthase gene in Panax ginseng, a stimulating a higher biosynthesis of triterpenes and phytosterols). Production of designer secondary metabolites The chemical synthesis of many compounds produced by plants in the form of secondary metabolites is already available, still due to their relatively high costs, even to-date, most of the pharmaceutically important secondary metabolites are isolated from plants. Biotechnological production in plant cell cultures could be an attractive alternative, but even currently this has had only limited success because of a limited understanding of how these metabolites are synthesized. Recent advances in plant genomics and metabolite profiling, however, seem to offer unprecedented possibilities to explore the extraordinary complexity of the plant biochemical capacity. State-of the art genomics tools can be used to enhance the production of known target metabolites or to synthesize entire novel compounds by so-called combinatorial biochemistry in cultivated plant cells (Oksman-Caldentey and Inze, 2004). Ultimate goal of these efforts should be to assist the recent upsurge in improving the traceability and safety (reliability) of natural products, as well as the appropriate policies and legal frameworks to guide the protection, production (including organic production), trade, and applications of medicinal and aromatic plant materials (GWP, GAP, GCP, GMP, etc.). Literature Cited Bisset N. G. and Wichtl M. 2001. Herbal Drugs and Phytopharmaceuticals. Medpharm, Stuttgart Chiang Mai Declaration 1988. In: Akerele O, Heywood V, Synge H, editors. The conservation of medicinal plants. Proceedings of an International Consultation; 1988 March 21-27; Chiang Mai, Thailand. Cambridge: Cambridge University Press; 1991. Convention on Biological Diversity 1992. In: Secretariat of the Convention on Biological Diversity (2010) Global Biodiversity Outlook 3. Montral, 94 pages. EUROPAM 2006. Guidelines for Good Agricultural and Wild Collection Practice (GACP) of Medicinal and Aromatic Plants. Brussels, 3rd April, 2006; EUROPAM GACP Working Copy No. 7.3 FairWild Foundation 2010. FairWild Standard, Version 2.0. Weinfelden, Switzerland FAO, 1995. Non-wood forest products for rural income and sustainable development. Rome (Non-Wood Forest Products 7). Fiehn, O. 2002. Plant Molecular Biology 48: 155171, Kluwer Academic Publishers. The Netherlands. Franz C, Bauer R, Carle R, Tedesco, D, Tubaro A, and Zitterl-Eglseer K 2005. Study on the assessment of plants/herb extracts and their naturally or synthetically produced

components as additives for use in animal production. CFT/EFSA/FEEDAP/2005/01. Graham, S. 2002. Global estimates of endangered plant species triples. Scientific American. November 01, 2002 Huie, C.W. 2002. A review of modern sample-preparation techniques for the extraction and analysis of medicinal plants. Anal. Bioanal. Chem. (2002) 373 :2330 DOI 10.1007/s00216-002-1265-3 Iqbal, M. 1993. International trade in non-wood forest products: An overview. FAO, Rome, November 1993. FO: Misc/93/11 - Working Paper. Leaman, D.J., Schippmann, U. and Glowka, L. 1997. Environmental protection concerns of prospecting and producing plant-based drugs. In: Wozniak, D.A., Yuen, S., Garrett, M. and Schuman, T.M. (eds.): International symposium on herbal medicine. A holistic approach. Documents, proceedings and recommendations. 14 June 1997. Honolulu. pp. 352378, San Diego, USA, International Institute Human Resources Development. Nyiredy, Sz. 2002. Gygynvnykutats: A Gygyszerkutats Termszetes Rsze. Magyar Tudomny, 12/1200. Oksman-Caldentey, K-M. and Inze, D. 2004. Plant cell factories in the post-genomic era: new ways to produce designer secondary metabolites. TRENDS in Plant Science Vol.9 No.9 September 20004., p. 433-440. Prescott-Allen, R. and Prescott-Allen, C. 1996. Assessing the sustainability of uses of wild species. Case studies and initial assessment procedure. Gland & Cambridge, IUCN (Occasional Paper of the IUCN Species Survival Commission 12). Schippmann, U. 1997. Plant uses and species risk. From horticultural to medicinal plant trade. In: Newton, J., ed., Planta Europaea. Proceedings of the first European Conference on the conservation of wild plants, Hyres, France, 28 September 1995. pp. 161165, London, Plantlife. Schulz, H. 2010. Application Of Rapid Analytical Methods For Efficient Evaluation Of Plant Genetic Resources And Breeding Material. Acta Hort. (ISHS) 860:225-234 Vantomme, P. 2002. In: Anon., 2002. Conservation impacts of commercial captive breeding workshop. Briefing notes II. 79.12.2001, Jacksonville. Cambridge, UK, IUCN/SSC Wildlife Trade Programme. WHO, IUCN & WWF 1993. Guidelines on the conservation of medicinal plants. Gland & Geneva, Switzerland. WHO 1998. Quality control methods for medicinal plant materials World Health Organization Geneva. Pp. 122. WHO 2002. Traditional medicine strategy 20022005. Retrieved from WHO website, http://whqlibdoc.who.int/hq/2002/who_edm_trm_2002.1.pdf (viewed: 27.02.2011)

Figure 1

Conservation status of medicinal plant species in different geographic regions

Table 1 -

Analytical methods for the study of medicinal and aromatic plants

Chromatographic separation methods Non-volatile compounds: TLC = thin layer chromatography HPTLC = high performance thin layer chromatography HPLC = high performance liquid chromatography Volatile compounds: GC = Gas chromatography applying different detectors FID = flame ionization detector NPD = nitrogen-phosphorous detector Modern sample preparation techniques linked to fast chromatographic methods SPE = Solid Phase Extraction, SPME = Solid Phase Microextraction SBSE = Stir-Bar Sorptive Extraction ASE = Accelerated Solvent Extraction SFE = Simultaneous Distillation Extraction