Salicylates of Intact <em>Salix myrsinifolia</em> Plantlets Do Not Undergo Rapid Metabolic Turnover^{<a href="#FN1" rid="FN1" class=" fn">1</a>}

Abstract

The phenolic glucosides of the Salicaceae play an important role in plant-herbivore interactions, which have been studied intensively by many research groups. These glucosides may protect plants against many generalist insect herbivores (Tahvanainen et al., 1985b; Lindroth et al., 1988; Lindroth and Peterson, 1988; Clausen et al., 1989; Denno et al., 1990). But on the other hand, they may act as positive cues or feeding stimulants for specialist herbivores (Matsuda and Matsuo, 1985; Smiley et al., 1985; Tahvanainen et al., 1985b; Rowell-Rahier and Pasteels, 1986; Denno et al., 1990; Rank, 1992). Some specialists may use phenolic glucosides for their own defense against predators (Pasteels et al., 1983; Smiley et al., 1985; Rowell-Rahier and Pasteels, 1986; Denno et al., 1990). Phenolic glucosides also affect the food selection of mammalian herbivores (Palo, 1984; Smiley et al., 1985; Tahvanainen et al., 1985a; Reichardt et al., 1990; Bryant et al., 1992) and are considered to be constitutive defenses because they are present in plant tissues all the time (Feeney, 1976).

The ability to synthesize a variety of simple phenolic glucosides is characteristic for willows (Salix spp.) and other members of the Salicaceae family (Thieme, 1965c; Egloff, 1982; Palo, 1984; Julkunen-Tiitto, 1989). Salicylates, a group of chemically related phenolic glucosides based on the structure of salicin (Fig. ​(Fig.1),1), are the most common phenolic glucosides found in willows (Palo, 1984; Julkunen-Tiitto, 1989). Salicin and salicortin are the most widespread salicylates (Palo, 1984; Julkunen-Tiitto, 1989), while acetylsalicortin is found in only a few willow species (Julkunen-Tiitto, 1989).

Figure 1

Chemical structures of salicylates of S. myrsinifolia and their interrelationships. Salicortin degrades to salicin and 2′-O-acetylsalicortin to 2′-O-acetylsalicin and further to salicin and in both cases a toxic 6-HCH-moiety will be released. 2′-O-acetylsalicin was not detected in this study. Salicin will degrade to saligenin (salicyl alcohol) by the hydrolytic removal of a Glc molecule. Saligenin can be converted back to salicin and salicin to diglucoside of salicin by glucosylation. The dashed line from diglucoside of salicin to salicin indicates a hypothetical step.

Salicortin and other salicylates such as tremulacin and acetylsalicortin, which contain a 1-hydroxy-6-oxo-2-cyclohexen-1-carbonyl moiety, are very labile in vitro and easily degrade to salicin (Thieme, 1965b; Pearl and Darling, 1971; Lindroth and Pajutee, 1987; Meier, 1988) and then to saligenin (Julkunen-Tiitto and Tahvanainen, 1989) during the isolation procedure (Fig. ​(Fig.1).1). At the same time, the labile 1-hydroxy-6-oxo-2-cyclohexen-1-carbonyl moiety is released and converted to 6-hydroxy-2-cyclohexenone (6-HCH) and catechol (Julkunen-Tiitto and Meier, 1992b), which are effective components of salicortin and its derivatives in defense against herbivores (Clausen et al., 1989; Reichardt et al., 1990). Improper preservation of samples (Julkunen-Tiitto, 1985; Lindroth and Pajutee, 1987; Julkunen-Tiitto and Gebhardt, 1992; Orians, 1995), long extraction times (Julkunen-Tiitto, 1985), and freezing (Julkunen-Tiitto, 1989) may all cause the degradation of salicortin.

As carbon-based compounds that do not contain nitrogen, the salicylates are thought to be quite cheap defenses for plants, especially when nitrogen is the growth-limiting factor (Bryant et al., 1983). On the other hand, in certain willow species the concentrations of salicylates may be very high (Julkunen-Tiitto, 1989), and if they also undergo rapid metabolic turnover (Reichardt et al., 1991), their maintenance at a given concentration may demand large amounts of resources from the plants (Gershenzon, 1994).

Phenolic glucosides of Salicaceae are considered to be “dynamic” substances that are subjected to metabolic turnover (Reichardt et al., 1991), but no earlier studies have determined the rate of metabolic turnover. According to models used to estimate the metabolic costs of the chemical defense of plants (Fagerström, 1989; Skogsmyr and Fagerström, 1992), an increase in concentration of the defense substance or its metabolic turnover rate raises the cost of defense.

The aim of this work was to ascertain whether willow salicylates are exposed to metabolic turnover. Because of the high inter-individual variation in the concentrations of salicylates (Julkunen-Tiitto, 1985, 1989; Meier, 1988; Julkunen-Tiitto and Meier, 1992a), clonal material was used in the experiment. Salix myrsinifolia was chosen because it contains a high level of salicylates, especially salicortin, and is easy to culture in vitro. In addition, S. myrsinifolia is one of the most promising willow species for the production of herbal drugs (Julkunen-Tiitto and Meier, 1992a), so it is important to ascertain to what extent the maintenance of a high level of salicylates requires plants resources

Changes in the concentrations of salicylates of control and AIP-treated plants during the turnover study were tested by linear regression after ln-transformation. Differences between regression coefficients of control and AIP-treated lines were tested by Student's t test (n = 29–30).

Regression coefficients were determined by linear regression after the data were ln-transformed, and half-lives of salicylates were calculated using the equation of first-order kinetics.

ACKNOWLEDGMENTS

Footnotes

LITERATURE CITED