However, starting the screening with enriched fractions is based on a higher number of starting test samples since multiple fractions are generated from a single plant extract. In cases when the pharmacological assays allow HTS, this approach is advantageous, since many HTS approaches that do not work well with plant extracts due to interferences with ubiquitous perturbing compounds, could be successfully applied to enriched fractions Eldridge et al.
In addition to HTS with libraries of randomly obtained plant-derived fractions, also in this occasion different knowledge-based strategies can be applied for the assembly of focused libraries containing just fractions with certain characteristics see Section 3. For example, an approach for selection of extracts and fractions based on their drug-like physicochemical properties was recently developed Camp et al. Since the bioactivity of a plant extract may be the result of synergistic interactions of several components, and in this case bioactivity-guided fractionation might fail, a synergy-directed fractionation strategy was recently developed Junio et al.
This approach combines bioactivity testing of the generated fractions for synergistic interactions, with MS-profiling and natural products isolation, aiming to identify synergistic interactions of extract constituents which could be missed using the traditional bioactivity-guided fractionation. Noteworthy, in addition to the identification of single bioactive plant constituents, there is also an increasing interest in the use of standardized plant extracts as herbal medicinal products or dietary supplements.
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Especially if there are synergistic or additive effects, if bioactive compounds are more stable in the extract, or if the bioavailability is better, an extract might possibly have higher therapeutic efficacy compared to the isolated pure compounds. However, more research is needed in this direction Gertsch, ; Liu and Wang, ; Liu, ; Schmidt et al.
Another important aspect related to the identification of plant-based bioactive molecules, is that it is now increasingly recognized that some natural products are prodrugs that need to be metabolized in vivo by the intestinal microorganisms or by the mammalian organism in order to yield pharmacologically active molecules Akao et al.
This is posing serious challenges to some of the established approaches related to bioactivity investigations of traditionally used medicinal plants, since in such occasions the effector compounds are not present in the starting plant material and bioactivity cannot be detected with many conventionally used assays and approaches [e. Many potent plant-derived natural products such as paclitaxel, podophyllotoxin, or vinblastine share severe difficulties to meet the market demands, as their natural sources are often slow-growing or even endangered species that tend to accumulate these compounds at very low quantities over long growth periods Miralpeix et al.
Natural populations of many medicinal plants have been increasingly facing pressure from mostly anthropogenic factors like environmental damage, deforestation and industrialization due to extensive population growth, fires and other natural disasters, development of land for agriculture, climate change, and last but not least, extensive collection of plant material from the wild Arora et al.
Two thirds of the estimated 50, medicinal plant species currently used are collected from the wild, raising concerns about issues such as diminishing populations, loss of genetic diversity, local extinctions, and habitat degradation Canter et al. While isolation of natural products directly from the plant species in which they occur is often acceptable if small to moderate amounts of the respective compounds are needed for example for the supply of sufficient quantities for small-scale experimental laboratory work , additional routes for resupply need to be considered once there are increased market demands for the respective natural product.
In these cases natural products can be resupplied by the application of plant cell and tissue culture, heterologous production, total chemical synthesis, or semi-synthesis from isolated precursors occurring more abundantly in nature, as discussed in greater detail in the following subsections.
Cultivation of endangered medicinal plants under controlled conditions represents a promising protective approach Tasheva and Kosturkova, For some of the stages which make up the process of domestication of wild-growing plant species, in vitro techniques can be applied, for example in selection breeding for genetic improvement or storage of plant materials Bohr, ; Franz, Since the fundamental report by Haberlandt on the in vitro cultivation of isolated plant cells Haberlandt, , plant cell and tissue culture has developed into an important and well established discipline, with impact on both basic research and applied technologies.
Out of these, the in vitro propagation micropropagation of plants and the in vitro culture of plant organs typically roots can provide means for the supply of plant material as source of natural products. Plant cell culture i. Micropropagation is defined as the in vitro plant regeneration from excised plant parts through either shoot organogenesis or somatic embryogenesis Siahsar et al.
On a commercial scale it has developed into a multi-billion-dollar industry and offers important advantages over the conventional propagation of plants, like production of large numbers of genetically homogenous plants year-round in a comparatively short time, disease-free propagules, and greatly enhanced multiplication rates Debnath et al. Micropropagation has become a standard method of plant production for many species of economic value, such as plants of agricultural, ornamental, vegetable, and forestry importance Debnath et al.
In vitro propagation certainly also holds significant potential for the mass production of medicinal plants and numerous protocols for micropropagation of a huge number of medicinally used species have been developed up to date Afolayan and Adebola, ; Chaturvedi et al. However, the higher costs of in vitro techniques as compared to conventional propagation are a major disadvantage and have limited its use at a commercial level Mehrotra et al. Although medicinal plants are increasingly introduced into field culture Wiesrum et al. Plant cell cultures dedifferentiated cells suspended in liquid nutrient medium have been aptly denominated as chemical factories of secondary metabolites Rao and Ravishankar, They offer a number of advantages over the conventional use of plants as sources of phytochemicals, like for example independence of geographical, seasonal, and environmental variations; continuous reliable production in uniform quality and yield; avoidance of pesticide and herbicide application; and comparatively short growth cycles Bonfill et al.
Technologies which have been developed for other cell culture systems like mammalian cells or yeast can be adapted for large scale industrial applications with plant cells Kolewe et al.
But, despite of many decades of research in the field, at present only fourteen substances or products are produced commercially from plant cell cultures Frense, ; Kolewe et al. This is due to several limitations of plant cell cultures in comparison to microbial production sources, including slow growth rates and low and variable yields of metabolites — in fact, many metabolites do not accumulate in plant cell cultures because of lacking differentiation and compartmentalization Kirakosyan et al. While still very little is known about the plant secondary metabolites biosynthesis and its regulation, it remains questionable whether in a foreseeable time plant cell cultures will play a significant role in the industrial production of natural products.
Nevertheless, in order to get a broad overview of the existing resupply approaches, some aspects of the methodology will be briefly addressed here. Noteworthy, in many cases the formation of desired compounds in plant cell cultures can be distinctly enhanced by different measures. In this regard, one important approach is the modification of the nutrient medium with the aim of increasing biomass and, often in a subsequent step, optimizing metabolite production Bonfill et al.
Herein, key factors include the ionic strength of the basal medium, phosphate and nitrate level, the level and type of sugars, growth regulators, and the feeding with precursors Murthy et al. Furthermore, a number of physical factors can influence the properties of plant cell cultures. These are the inoculum density, temperature, light quality and intensity, pH of the nutrient medium, and agitation and aeration of the culture batch Kanokwaree and Doran, ; Murthy et al.
Dedifferentiated cells, typically constituting plant cell cultures are usually prone to high variability in product biosynthesis, formation of large cell aggregates, and they are sensitive to shear stress — with the latter two features being particularly important obstacles in industrial-scale cultivation Yun et al.
Recent research indicates e. Cambial meristematic cells are considered to represent a key platform technology for the large-scale production of plant natural products Roberts and Kolewe, ; Yun et al. Elicitation is a further strategy to improve the accumulation of secondary metabolites in plant cell and organ cultures. The major function of plant secondary metabolites is the protection from pathogens and insects, or from other biotic or abiotic stresses Ramakrishna and Ravishankar, ; Zhao et al. Biotic elicitors include substances such as pectin, chitosan, methyl jasmonate, or yeast extract, while salicylic acid, heavy metals, or electromagnetic treatment are typical abiotic elicitors Shilpa et al.
Consequently, elicitation has been successfully utilized for the enhancement of secondary product formation in cell and organ cultures of many plants Namdeo, ; Sharma et al. The progressive understanding of signal transduction pathways in the elicitor-induced production of secondary metabolites will be important for the further optimization of plant tissue culture-based production systems Zhao et al.
Metabolic engineering holds a high potential for the enhancement of biomolecule formation in plant cell cultures. A range of techniques are nowadays available for the identification of rate-limiting gene products within biosynthetic pathways, and the subsequent transient or stable transformation of plant cells with the aim of up- or down-regulating relevant gene products Wilson and Roberts, Herein, major progress has been achieved in the manipulation of the biosynthesis of various terpenoids, flavonoids, and alkaloids in a range of plants Glenn et al.
In general, the productivity of a plant in vitro culture is related to the degree of differentiation, and this is one of the reasons why in many cases only low yields of desired secondary metabolites can be found in cell cultures Kolewe et al. In recent years hairy root cultures have received increasing interest. The hairy root disease is the result of the infection of wounded plants by the soil bacterium, Agrobacterium rhizogenes. The molecular mechanisms underlying this disease, which can cause significant losses in agriculture, have been largely elucidated and reveal a form of natural genetic engineering Georgiev et al.
Comprehensive Bioactive Natural Products Vol 1 : Potential & Challenges
Specifically, the so-called T-DNA containing rol genes and harbored by the bacteria's root inducing Ri plasmid is transferred to the plant cell and integrated into the nuclear genome. As a result, prolific growth of neoplastic roots occurs, and these hairy root systems can be maintained in in vitro culture Ron et al. Hairy root cultures have been established from many plant species and have been shown to be a valuable biological system with a wide range of applications such as studies in root physiology, phytoremediation, elucidation of biosynthetic pathways, molecular breeding, and production of secondary metabolites Ono and Tian, Hairy roots exhibit high growth rates without the need of phytohormones.
Unlike cell cultures they are genetically and biochemically stable, and compared to the respective mother plant they often show the same or even greater biosynthetic capabilities for secondary metabolite production Georgiev et al. As with cell cultures, secondary metabolite production can often be enhanced through optimized growth conditions or by using elicitors, and through metabolic engineering by transferring heterologous genes with engineered A.
Recent research on the production through hairy roots of lignans Wawrosch et al. Examples of hairy root cultures fromplants producing important pharmacologically active molecules developed from ROOTec bioactives Ltd a. The heterologous biosynthesis of plant-derived natural compounds, i. This approach has originally been developed in order to transfer the biosynthesis of valuable microbial secondary metabolites from their original producers that are often poorly characterized and difficult to cultivate into microbial hosts that are more amenable to fermentation processes Ongley et al.
Predominantly, microbes have served as host organisms, however, heterologous biosynthesis of plant-derived secondary metabolites also has been implemented in plant species genetically more amenable than the native sources, such as tobacco or Arabidopsis ssp. Howat et al. Heterologous biosynthesis has the advantage of being more environmentally friendly than chemical synthesis, since it avoids the use of organic solvents, heavy metals, and strong acids or bases Marienhagen and Bott, Furthermore, especially in case of microbial hosts, processes are comparably cost-efficient, as microbial growth is based on inexpensive renewable feedstocks and the fast doubling rates of microorganisms allow short production times.
In contrast to synthetic chemistry-based routes, microbial fermentations are readily scalable from the lab bench to industrial-sized fermenters Chemler and Koffas, ; Marienhagen and Bott, Furthermore, recombinant microorganisms usually do not possess pathways competing to the heterologously expressed one. Therefore, the desired products are chemically distinct and can be rather easily purified without the need to remove closely related constituents Chemler and Koffas, ; Marienhagen and Bott, The successful reconstitution of a biosynthetic pathway in a heterologous host requires in-depth knowledge about the involved enzymes and the genes encoding them, its regulation, and its compartmentalization Miralpeix et al.
Therefore, the main obstacles in the establishment of heterologous biosynthesis routes are that for many medicinally important natural compounds, the plant biosynthetic pathways are not fully elucidated and the required pathway genes are not available Miralpeix et al. The required genes need to be isolated from their native sources and mobilized into the heterologous host via an appropriate vector construct.
Expression strategies depend on the host organism, but will either involve the introduction of plasmids that remain episomal or of constructs that integrate into the genome and become a new genetic locus Miralpeix et al. The choice of the appropriate host system is crucial for success of heterologous production.
Concerning microbial hosts, Escherichia coli and Saccharomyces cerevisiae have been most widely applied. Both are non-pathogenic microorganisms that have been extensively used for industrial fermentation and have served as model organisms for fundamental molecular biology research, resulting in a high level of knowledge on their physiology and genetic manipulation Chen et al. However, E. Moreover, as a prokaryote, E. This complicates the implementation of certain eukaryotic enzymes such as the cytochrome Ps, that include several key enzymes of flavonoid biosynthesis, and that are usually attached to the eukaryotic cell's endoplasmic reticulum Miralpeix et al.
In this respect, S. Also plant-based expression platforms, including transgenic cell suspension cultures, hairy root cultures, and whole plants, have been used for the heterologous biosynthesis of secondary plant metabolites Miralpeix et al. For example, a recent work described the discovery of the last missing steps of the secoiridoid biosynthesis pathway in C.
Don and the expression of the complete pathway in the heterologous host Nicotiana benthamiana Domin. While the production of recombinant proteins through transgenic plant cell cultures is not the topic of this review, it should be noted that a number of proteins are currently in clinical stages of development or, as is the case for human glucocerebrosidase, already on the market Obembe et al. As an example, it is possible to produce artemisinin in transgenic tobacco as discussed in more detail below Farhi et al. Once a biosynthetic pathway has been successfully reconstituted in a heterologous host, it is still not guaranteed that the desired metabolite is produced at satisfactory amounts.
High titre production might be hampered by the limited availability of one or more precursors needed for biosynthesis thus creating bottlenecks in the pathway, by insufficient stability of the heterologously expressed enzymes against proteolytic activity, by pathway imbalance due to redistribution of metabolic fluxes, by consumption of the produced metabolites in an endogenous pathway of the heterologous host, or by negative or toxic effects of the heterologously produced metabolites to the host organism Lo et al.
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Therefore, in order to successfully design a heterologous production process, a number of challenges have to be addressed. This is mainly done by metabolic engineering, i. In order to allow efficient metabolic engineering, scientists increasingly make use of tools from the emerging field of synthetic biology in order to enable balanced expression of pathway genes, to allow spatio-temporal separation of the molecular pathway components, and to optimize genes by directed evolution approaches Jensen and Keasling, ; Li and Pfeifer, The rational application of these tools is increasingly supported by in silico tools for modeling of genome-scale metabolism, pathway search and enumeration, metabolic flux analysis, and pathway ranking Fernandez-Castane et al.
Based on their biogenetic origin, plant secondary metabolites can be roughly grouped into three major classes: phenylpropanoids, alkaloids, and terpenoids Peters and Crouteau, Heterologous production of plant secondary metabolites from all three classes was recently reviewed elsewhere Marienhagen and Bott, ; Miralpeix et al.
Since the currently most successful heterologous production process has been developed for an isoprenoid, namely for a precursor of the sesquiterpene artemisinin, the class of terpenoids will be discussed in more detail in the following paragraphs. With at least 40, structures known to date, many of them being plant-derived, isoprenoids also known as terpenoids represent the largest class of secondary plant metabolites scheme of the isoprenoid biosynthetic pathway with a focus on the biosynthesis of paclitaxel and artemisinin is presented on Fig.
Biosynthesis of plant-derived terpenoids originates from the two isomeric C5-building blocks isopentenyl-pyrophosphate IPP and dimethylallyl-pyrophosphate DMPP. They are formed either in the mevalonate pathway that is compartmentalized in higher plants in the cytosol early steps , the endoplasmic reticulum 3-hydroxy-3 methylglutaryl-CoA reductase and peroxisomes late steps , or from the 2C-methyl- d -erythriolphosphate MEP pathway also known as non-mevalonate pathway that is present in most prokaryotes, all eukaryotes, and the chloroplasts of higher plants Kuzuyama and Seto, ; Lange and Ahkami, Condensation of these two C5-pyrophosphates and of larger building blocks derived from them leads to the biosynthesis of monoterpenes C10, comprising many volatile oil constituents , sesquiterpenes C15, e.
Due to their high medicinal relevance and their restricted availability from traditional production routes, very big efforts have been invested in order to facilitate the heterologous production of artemisinin and paclitaxel Howat et al.
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Scheme of the isoprenoid biosynthetic pathway with a focus on the biosynthesis of paclitaxel and artemisinin. Dotted arrows indicate multiple enzymatic steps. The content of the scheme is adapted from: Lange and Ahkami, ; Malik et al.
www.esport.cz/scripts/tattnall/wof-floert-grup-ueyeleri.php Artemisinin is a sesquiterpene lactone endoperoxide naturally occurring in sweet wormwood A. Artemisinin is effective against severe malarial forms and is also currently studied for activity against certain cancer types and viral diseases. However, its availability is limited due to its low abundance in the natural source and due to the low yield of chemical total synthesis, thus resulting in a high price that is unaffordable for people in developing countries where malaria frequently occurs Kong et al.
Therefore, there has been an intensive exploration of alternative strategies for artemisinin production in order to enhance supply and to reduce costs, among them the development of a heterologous production process. As the artemisinin biosynthetic pathway downstream of artemisinic acid or dihydroartemisinic acid is not yet fully understood, the goal of the hitherto strategies was the heterologous production of artemisinin precursors such as amorpha-4,diene, artemisinic acid or dihydroartemisinic acid, followed by their semi-synthetic derivatization to artemisinin Kong et al.
These efforts have led to the development of an economically feasible production process consisting of heterologous production of artemisinin from inexpensive carbon sources glucose, ethanol in a S. Sanofi is now using this technology for large-scale production of artemisinin.
Alternatively to this very successful development, efforts have been made to directly produce artemisinin in heterologous plant hosts, since microbial hosts only allowed the heterologous production of artemisinin precursors, while the heterologous production of artemisinin itself could only be realized in plants so far Farhi et al. Indeed, Farhi et al. The gene products were targeted to diverse cellular compartments to increase precursor availability.
Paclitaxel, a complex natural product with a diterpenoid core, was isolated and structurally elucidated for the first time in the late s from the stem bark of western yew T. Wani et al. In the late , it was discovered as the first antimitotic agent that acted by promoting the irreversible assembly of tubulin into microtubules Schiff et al.
Currently, paclitaxel is approved for the treatment of different kinds of cancers, and its fields of application are expected to expand as the compound is currently also studied for the treatment of other, non-cancer related, diseases. As extraction from natural sources and total synthesis have proven infeasible to meet the market demands, the compound is currently produced by two different approaches: first, paclitaxel and analogs thereof are produced by semi-synthesis using taxanes that occur more abundantly than paclitaxel in various yew species such as bacchatin III or deacetylbacchatin III as starting materials; second, Taxus plant cell cultures are used for paclitaxel production Howat et al.
Also heterologous paclitaxel production has been intensively investigated. Indeed, the early-stage paclitaxel intermediate taxadiene could be produced in several microbial and plant heterologous hosts: while production in Arabidopsis thaliana L. As both transfected Arabidopsis and tomato plants were found to grow more slowly than the wild type plants, Anterola et al. Concerning microbial heterologous hosts, taxadiene levels of 8. However, one has to keep in mind that taxadiene is only a very early-stage intermediate of the paclitaxel biosynthetic pathway Fig.
As an example, it would be necessary to engineer six more hydroxylation reactions and other steps including several yet unidentified ones in order to achieve baccatin III, a feasible precursor for paclitaxel semi-synthesis. In addition, several steps in the overall pathway have not been conclusively identified or confirmed. Therefore, the total heterologous production of paclitaxel is not within reach to date. As can be seen from the examples discussed above, heterologous production has the potential to serve as an alternative production route, especially for plant-derived compounds that are difficult to access due to restricted abundance in their natural sources or due to structural constraints that make a synthetic approach unfeasible.
However, it is evident that the technique is currently far from being a commonly applicable alternative to traditional production routes of natural compounds. The major obstacle in the development of heterologous production processes is the current lack of in depth-knowledge concerning the biosynthetic pathways of many medicinally important plant constituents. Therefore, significant efforts will be necessary to achieve complete elucidation of key target pathways in medicinal plants, e. Don Miralpeix et al. As from the 25 plants with completely sequenced genomes published to date, only a few are medicinal plants, more or less all metabolic engineering approaches involving genes from medicinal plants have started with an incomplete datasets.
The complete sequencing of representative medicinal plant species would therefore allow more rationally designed metabolic engineering strategies Miralpeix et al. This task might be within reach, considering the revolutionary progresses sequencing techniques are currently facing Shendure and Ji, In cases where the total heterologous production of a complex natural compound is not yet possible, the production of decorated skeletons that are easily accessible to subsequent semi-synthetic derivatization could also be an option, as has been shown in the case of artemisinin.
A further problem is the low yield of many heterologous production processes, that makes them economically unfeasible or not competitive to other, maybe less environmentally-friendly or sustainable approaches. There are many issues to be considered in order to finally allow the heterologous production of an increasing number of plant-derived secondary metabolites in economically feasible processes, including the modification of pathway enzymes to improve their compatibility to the host organisms; the design of more stable expression systems that allow tens of heterologous enzymes to be expressed simultaneously; enhancement of the tolerance of host microorganisms to the heterologously produced metabolites, e.
Over nearly years, the synthetic chemistry community has achieved a vast and diverse arsenal of transformations for bond connection and functionalization of organic molecules at an ever increasing pace. Total synthesis of a plethora of intricate naturally occurring compounds, many originating from plants, could be achieved Nicolaou, , very often as a result of arduous work and sometimes in a race between academic research groups. Yet, these synthetic endeavors are not reflected in the broad industrial manufacture of natural products.
Costs and feasibility of reagents and catalysts, exothermic and cryogenic reaction characteristics, chromatographic purification, solvents, spent reagents and byproducts handling are among the aspects that chemical engineering is concerned with during process development of therapeutic compounds Ager, Specifically, the safety, health and environment SHE considerations led to the formulation of a framework now known as Green Chemistry.
It attempts to achieve sustainability at the molecular level and applies to all stages of the chemical life cycle Anastas and Eghbali, Key to this are the Twelve Principles of Green Chemistry, meant to be a cohesive set of guidelines for the design of products and processes. Outlined very briefly, they are: Prevention of waste, rather than treating it , Atom Economy vide infra , Less Hazardous Chemical Synthesis applies to methodology , Designing Safer Chemicals while preserving their intended function , Safer Solvents and Auxiliaries if they cannot be avoided altogether , Design for Energy Efficiency methodologies should allow ambient temperature and pressure , Use of Renewable Feedstock rather than depleting limited raw materials , Reduce Derivatives in concession steps such as non-strategic redox reactions and functional group interconversions, protective group manipulations , Catalysis as selective as possible , Design for Degradation applies to products which should break down to nonhazardous materials at the end of their function , Real-Time Analysis for Pollution Prevention before hazardous substances are formed , and Inherently Safer Chemistry for Accident Prevention minimizing the hazard factor in the risk function.
These principles are discussed in more detail with examples elsewhere Anastas and Eghbali, Meaningful statements about the impact on health and environment also require a quantitative assessment to describe the various shades of green of a product or process. For this reason, different metrics have been proposed, gradually marking a paradigm shift from the overall yield as the most important criterion toward a more integrated view Dicks and Hent, With regard to a synthetic plan chemical reactions on paper , the concept of atom economy AE was proposed early on Trost, as a measure of how much of the reactants, in terms of their molecular weight M , remains in the desired final product Eq.
When molecules are transformed, waste material is produced in most cases, depending on the type of reaction mechanism. For example, addition and rearrangement reactions tend to be more atom economic while substitution and elimination reactions do not. When extending the concept to a reaction sequence composed of a number of individual steps, the intermediate compounds do not appear in Eq. A reactant is defined as any component which contributes atoms to intermediate or final products, even if none of the reactant eventually remains in the final product as it is the case in protection—reaction—deprotection sequences.
Catalytically active species, solvents and other auxiliary components are neglected unless they are partially incorporated into intermediate or final products. The equivalents of reaction components used to calculate AE are taken as demanded by the stoichiometry of the transformation, i. The advantage of this approach is that chemists can evaluate different synthetic strategies before real experiments are attempted Dicks and Hent, , where subsequent reaction optimization then very likely calls for reactant equivalents, catalysts, solvents, auxiliaries, and other conditions to be varied considerably.
For the same reasons, however, atom economy is also quite remote from real-world chemistry.
A metric which actually accounts for experimental details, called the reaction mass efficiency Curzons et al. It was shown that Curzons RME could be used to compare costs for drug manufacture in general Constable et al. A related quantity is the maximum or kernel RME Andraos, , which applies to a hypothetical scenario where all excess reagent is recovered i. The most critical view of a process is the generalized RME Andraos, because it is calculated as the mass of product divided by the total mass of all input materials including catalysts, auxiliaries, solvents and work-up and purification materials, Eq.
In order to organize all this information, a visual aid called the radial pentagon was proposed Andraos and Sayed, Contrarily to the green metrics discussed so far, the most prominent counter-green metric the ideal value of which is zero is the environmental impact factor, or E factor, as introduced by Sheldon Sheldon, , which focuses on waste production irrespective of where it originates from Eq. Notably, E factors associated with pharmaceutical production are generally higher by three to four orders of magnitude when compared with the oil refining petrochemical industry Sheldon, , and non-optimized laboratory preparations of complex natural products have E factors which are even far higher.
It is important to understand that no single metric will completely reflect all aspects of efficiency and safety. Moreover, calculating various metric parameters for multistep especially convergent synthetic plans by hand is rather impractical, since it is easy to lose track and make mistakes.
To address this, a spreadsheet based, step-by-step algorithm was created for practical application to any complex synthesis, and the method was applied to exhaustively compare the greenness of 18 synthetic plans both industrial and academic of oseltamivir Andraos, , the initial preparations of which were based on the elaboration of quinic acid obtained from e. This revealed that the industrial semi-synthetic shikimic acid route Harrington et al. The industrial route produced the least amount of total waste It was also noted that academia continues to improve existing synthesis as well.
Generally, semi-synthesis can provide access to large quantities of a desired drug if a suitable precursor is available. As mentioned in Section 4. While more than metric tons of T. Pacific yew bark had to be collected in and Cragg, to supply the compound for clinical trials only, thereby killing the slow-growing trees, the tetracyclic diterpene deacetylbaccatin III representing the structurally most demanding moiety of paclitaxel is readily obtainable from the needles of Taxus baccata L. This harvesting method leaves the trees intact, and deacetylbaccatin III is converted chemically to paclitaxel in high yield Denis et al.
Thereafter, a chemoenzymatic process was developed in which the chiral intermediate necessary to elaborate deacetybaccatin III to paclitaxel was produced via kinetic resolution employing reusable, immobilized lipases, and the process was scaled to up to l, resulting in excellent yield, enantiomeric excess and purity Patel et al. Strategies for natural product synthesis are not readily delineated by type of chemical reaction, although a methodology-oriented central step may sometimes be identified.
On the other hand, a synthetic strategy may be built upon procuring important intermediates or starting materials. An example for this from chemoenzymatic methodology is the enantioselective dihydroxylation of variously substituted benzenes by toluene dioxygenase Hudlicky and Reed, This transformation is remarkable since it accomplishes dearomatization of benzene derivatives to give arene dihydrodiols, allowing for synthetic routes, which would otherwise not be feasible.
It is also a whole-cell fermentation in E. Compounds of this sort, the production of which by chemical means would be a lengthy procedure, are highly useful building blocks because chemical synthesis might be next used to elaborate them further to plant compounds such as pancratistatin and morphine Hudlicky, ; Reed and Hudlicky, , or analogs and derivatives thereof, and also oseltamivir Werner et al.
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