Recent Advances in Polyphenol Research, Volume 3

Chapter 2

Polyphenols: From Plant Adaptation to Useful Chemical Resources

Alain-Michel Boudet

Abstract: Plants produce a large range of natural substances called secondary metabolites. Although these compounds do not appear to be directly involved in the basic activities of plant cells, they play an essential role in plant development and plant/environment interactions. Among these, an immense variety of phenolic compounds has been progressively synthesized by plants during the course of evolution. All of them are derived from primary metabolites that are fed via the shikimate pathway. Many of these compounds are of medical or socio-economic value and there is growing interest in research and industry in polyphenols. In this chapter, we will consider essentially three different aspects:

1. The emergence of phenolic metabolism during evolution and the adaptation of plants to a terrestrial environment.

2. The role and the specificities of the shikimate pathway in higher plants.

3. The diversified uses of phenolics in different human activities.

Keywords: plant evolution; vessels and conducting éléments; lignins, shikimate pathway; phenylpropanoid pathway; arogenate pathway; quinic acid; comparative genomics; polyphenols and health; lignocellulosics bioconversion

2.1 The emergence of phenolic metabolism and the adaptation of plants to a terrestrial environment

Fossil data indicate that the first land plants appeared around 500 million years ago (Kenrick & Crane, 1997). As emphasized by these authors “the origin and early diversification of land plants mark an interval of unparalleled innovation in the history of plant life with an extraordinary array of complex organs and specialized tissue systems.” Independent evidence from morphological, ultrastructural, biochemical and molecular data have shown that land plants (embryophytes) consisting of liverworts, hornworts, mosses and tracheophytes originated from charophycean green algae. This small group of predominantly freshwater green algae possesses several biosynthetic attributes that are expressed more fully among land plants including the capacity to produce sporopollenin, cutin, and some phenolic compounds (Kroken et al., 1996). However, they lack other biochemical and morphological characteristics that likely evolved during the complete transition to land.

The fossil record of spores combined with phylogenetic studies indicates that groups related to living bryophytes were early colonizers of the land. Vascular plants arose later in the late Silurian period (∼400 million years ago). They diversified rapidly, when an early split in the history of land plant evolution gave rise to two major lineages: the lycophytes and euphyllophytes (Kenrick & Crane, 1997) (Fig. 2.1). These lineages contain specialized water-conducting tracheary elements reinforced by lignins. Together, these changes resulted in more highly differentiated plants with different tissue systems including impermeable exterior surfaces and water conducting cells.

Fig. 2.1 Simplified phylogenetic tree showing the origin of land plants and the occurrence of lignins.

The appearance of phenolic compounds in large amount in plants is clearly related to their adaptation to terrestrial environments. The first plants that moved from an aquatic environment to emerged land had to face important stresses, including desiccation, varying temperatures, UV radiation and the attack of microorganisms. They had also to compete between each other for light and nutrients. The necessary biochemical adaptations to these new challenges were in large part issued from the phenyl propanoid pathway. This pathway provided different precursors at the origin of lignins for stem rigidity and vascularization, flavonoids for reproductive biology (fruit and flowers colors). Pathway products also provided protection against UV, microbial defense and also symbiotic plant-microbe interactions (e.g., leguminosae/rhyzobium). Other simple phenolics were also involved as attractants or deterrents in different adaptative strategies. Among these adaptations, the deposition of lignins in plant cell walls was likely a crucial mechanism that allowed the development of upright plants of large size adapted to a terrestrial habitat. Lignin provided structural rigidity for tracheophytes to stand upright and strengthened the cell wall of their water-conducting tracheary elements allowing them to withstand the negative pressure generated during transpiration.

The problem is to identify at which stage of evolution these different compounds appeared and how the enzymes/genes responsible for their synthesis emerged. As far as phenolic compounds are concerned, qualitative and quantitative differences occur in the biochemistry of embryophytes relative to charophytes, and between bryophytes and tracheophytes in the embryophytes. Classically, flavonoids seem to be restricted among extant plants to embryophytes, while true lignins are limited to (eu)tracheophytes.

Nevertheless, lignin-like polymers have been identified in the cell walls of the charalean alga Nitella and in primitive green algae (Delwiche et al., 1989). Lignans (Raven, 2000; Suzuki & Umezawa, 2007) and other phenolic cell wall material different from lignins (Erickson & Miksche, 1974; Miksche & Yasuda, 1978) have been characterized in mosses and liverworts that do not harbor a vascular system. Interestingly, lignans, which are dimers derived from hydroxycinnamyl alcohols, allyl phenols or hydroxycinnamic acids, are widely distributed within embryophytes, but whereas hydroxycinnamyl alcohols are involved in their synthesis in tracheophytes, the lignans from bryophytes are derived from hydroxycinnamic acids (Scher et al., 2003; Umezawa, 2003). All these different phenolic compounds have been associated to defense mechanisms against microorganisms or UV radiation (Ligrone et al., 2008).

From the point of view of the biosynthetic pathways, the shikimate pathway, which provides phenylalanine as a protein amino acid, is present in procaryotes and has been conserved in unicellular and pluricellular eucaryotes including higher plants. However, the phenylpropanoid nucleus, from which most of phenolic compounds are derived, is also a product from phenylalanine via phenylalanine ammonia-lyase (PAL) and the presence of this enzyme is a prerequisite for an active phenolic synthesis. Although PAL enzymes have been extensively characterized in all land plant lineages, including bryophytes, their distribution in lower organisms is limited. PAL is present in some fungi where it participates in the catabolism of phenylalanine (MacDonald & D’Cunha, 2007) and has also been characterized from a few bacteria (Xiang and Moore, 2005; Moffit et al., 2007). Emiliani and coworkers (Emiliani et al., 2009) have performed an extensive analysis of the taxonomic distribution and phylogeny of PAL. The 160 representative sequences chosen for final tree construction were obtained from different prokaryotes and eukaryotes (plants and fungi). The authors conclude that PAL emerged in bacteria, likely with an antimicrobial role, and that a member of an early fungal lineage obtained a PAL via horizontal gene transfer from a bacterium. This fungal PAL was transferred to an ancestor of land plants via an ancient arbuscular mycorrhizal symbiosis where it paved the way for the development of the phenylpropanoid pathway. It is also possible that the ancestor of land plants and the ancestor of fungi independently acquired their PAL from two different but related bacteria. Even though they are quite speculative, these hypotheses are supported by phylogenetic analyses and can explain the emergence of PAL in land plants.

Despite the absence of data about the other genes involved in the phenyl propanoid pathway, it can be proposed that the assembly of the whole pathway likely occurred stepwise by the recruitment of pre-existing enzymes from other metabolic routes (Dixon & Steele, 1999; Lehfeldt et al., 2000). As underlined by Weng and Chapple (2010), the enzymes of phenolic metabolism were gradually acquired from ancestors of primary metabolism through mutations and selection. As an example, PAL is homologous to histidine ammonia-lyase (HAL), an enzyme involved in histidine degradation in intermediary metabolism. Horizontal gene transfer from the few bacteria and fungi that harbor homologues of some enzymes of the pathway or mutations of some “enzyme precursors” of phenolic metabolism could have represented additional possibilities. As an example, we have found interesting homologies at the protein sequence level but also at the level of the structure of the genes (identical relative locations of introns and exons) between cinnamoyl-CoA reductase (CCR) and dihydroflavonol 4-reductase (DFR) (Lacombe et al., 1997). These observations suggest that either these closely related genes, with different functions, derive from a common ancestor gene or alternatively that the DFR that is involved in the synthesis of anthocyanins (assumed to be anterior or concomitant to lignins during evolution) was a precursor of CCR.

As stressed by Emiliani et al. (2009), it is generally assumed that there is no evidence for the presence of a full phenylpropanoid metabolism in organisms other than land plants, although some bacteria and fungi harbor homologues of a few enzymes of the pathway (Moore et al., 2002; Seshime et al., 2005). However, recent results lead to reconsider this general dogma. Secondary...