Higher plants produce a wide range of chemicals. More than 25,000 terpenoids, about 12,000 alkaloids, and 8000 phenolic substances have been identified thus far (Croteau et al., 2000). These chemicals serve in a variety of functions in plants. They defend against herbivores and pathogens, aid in interplant competition, attract beneficial organisms such as pollinators, and have protective effects with regard to abiotic stresses such as UV exposure, temperature changes, water status, and mineral nutrients. In addition, many secondary metabolites produced in plants are used by humans as spices, dyes, fragrances, flavoring agents, or pharmaceuticals. Many of these chemicals also promote human health and enrich our lives in many different ways.

The use of plant metabolites as natural medicines has a long history that can be traced back more than 3500 years, when Egyptian, Sumerian, Ayurvedic, and Chinese medicines were developed (Askin et al., 2002). Morphine was isolated as the principal active ingredient in opium just 200 years ago (1806), and the pain-killing and fever-reducing aspirin (acetyl salicylic acid) became available via chemical synthesis, instead of from willow bark extract, only about 100 years ago (1897). Following rapid progress in chemical synthesis, the microbial production of antibiotics, as well as biotransformation approaches, has increased the supply of modern medicines. Today, however, a large proportion (more than 25%) of products beneficial to humans are still derived from natural sources, especially from plants (Askin et al., 2002; Briskin, 2000; Fig. 11.1).

Many medicinal plants are still harvested in the wild due to the technical difficulties of cultivation, as well as for economic reasons. This harvesting of medicinal plants along with human disturbance of the natural environment increasingly raises concerns about diminishing biodiversity. For example, Taxus brevifolia (yew), which is used for the production of paclitaxel (TaxolTM), a potent antineoplastic agent, is an endangered species on the west coast of the United States. The increased demand and drastic reduction in plant availability increase the pressure to produce medicinal compounds in alternative ways, especially via cell/tissue cultures and transgenic plants since plant cells have a high potential for totipotency. Furthermore, the demand for quality materials has also increased since variations in medicinal plant quality and incorrect plant identification occasionally cause tragic consequences. Thus, the application of biotechnological approaches to produce secondary metabolites is an attractive alternative for their production, particularly in transgenic cell cultures.

In vitro cell culture systems have several advantages, whose benefits have been discussed in depth previously (Dougall, 1981). In summary, the desired metabolites are produced in a controlled environment, independent
FIGURE 11.1 Chemical structures of some important medicinal compounds.
FIGURE 11.1 Chemical structures of some
important medicinal compounds.
of climatic changes and soil conditions, and free of microbes and insects. In addition, materials of definitive quality can be produced, and whose productivity can be improved through cell selection and optimization of culture conditions. The established cell culture systems are also useful for understanding the synthesis of natural products since the results obtained are generally uniform and reproducible. The products are obtained under controlled conditions and are available in large quantities from cells that are less structurally organized than those of higher plants. On the other hand, cell culture systems also have several disadvantages; for example, the labor required to maintain cell culture systems by serial passages, and the possibility that cell properties may change during serial cultures over long periods of time or that cultures may be lost due to contamination.

Before 1970, the reported yields from cell cultures were generally lower than those in plants. However, as shown in Table 11.1, several cell culture systems can have considerable productivity, and in some cases their production exceeds than that found in the intact plant. Also some plant cell cultures still produce little, if any, of the desired compounds. These facts highlight the difficulties faced in producing useful metabolites in economically viable amounts. Accordingly, the current advancements in the molecular and cellular biology of secondary metabolism provide a basis for optimism regarding the commercial production of secondary products in cell/organ cultures and/or transgenic plants (Facchini and St-Pierre, 2005; Kutchan, 2005a,b; Zhao et al., 2005).

Since the biochemistry and cell biology of secondary metabolism represent the foundation of attempts directed toward biotechnological improvement, we first review studies that focus on alkaloids for illustrative purposes only. Simplest applications are biochemical conversions of chemicals that are readily available using isolated native, or recombinant, enzymes and cells as biocatalysts. More complicated metabolic engineering attempts require additional biochemical and cell biological information to optimize the conditions for production. The first section thus addresses the biochemistry and cell biology and then is directly related to the application of metabolic engineering.