Differentiation between de novo synthesized and constitutively released terpenoids from Fagus sylvatica

doi:10.1016/S0031-9422(98)00765-1

Phytochemistry

Volume 51, Issue 3, June 1999, Pages 383-388

https://doi.org/10.1016/S0031-9422(98)00765-1 Get rights and content

Abstract

Plants can use different carbon sources for the biosynthesis of individual C₁₀-volatiles. Feeding $^{13} CO$ ₂ to Fagus sylvatica plants, we monitored the incorporation of $^{13} C$ into the emitted compounds by mass spectrometry. By analyzing the rate and the distribution of labelling in the different fragments of the released compounds, we distinguished the instant emission within minutes after CO₂ assimilation from the delayed emission from storage compartments. The experiments provide evidence that the carbon skeleton of the emitted monoterpenes derived from two different carbon sources and that the contribution of the sources can be completely different for individual monoterpenes.

Introduction

Plants emit a wide range of volatile hydrocarbons into the atmosphere (Fehsenfeld et al., 1992). Numerous studies implicate that monoterpenes, next to isoprene, are the dominant class of released compounds (Guenther et al., 1995). Although the role of volatile terpenoids in secondary metabolism still represents a largely debated issue, models have been proposed describing for example the isoprene release as a means of controlling energy flow in plants (Sharkey, & Singsaas, 1995). The important role of volatile terpenoids in tritrophic plant–insect interactions is well established Dicke, & Sabelis, 1988, Turlings, Tumlinson, & Lewis, 1990. Pare and Tumlinson showed that, in cotton, the volatiles induced by insect herbivore damage are synthesized de novo with little or no release from storage, an observation that was interpreted as an active channelling of energy towards the release of volatiles as a defensive response (Pare, & Tumlinson, 1997a). The induced terpenoids showed significant higher incorporation levels of $^{13} C$ than those which were released constitutively. Recently, Loreto et al. gave evidence for the photosynthetic origin of released monoterpenes from Quercus ilex by $^{13} C$ labelling (Loreto et al., 1996b). Terpenoids released from Picea abies plants were partly enriched with isotope label whereas analysis of endogenous terpenoids in the needles indicate that even after a 24 h exposure to $^{13} CO$ ₂ there was no incorporation of $^{13} C$ (Schürmann, Ziegler, Kotzias, Schönwitz, & Steinbrecher, 1993).

In this study we used $^{13} C$ labelling to distinguish whether the volatile terpenoids from Fagus sylvatica are emitted directly after CO₂ assimilation depending on photosynthetic activity (connected with a rapid turnover of carbon) or are emitted from preformed intermediates which are stored in different compartments of the plant (McGarvey, & Croteau, 1995). Recently, we have shown that the rate of emission of certain terpenes from Fagus sylvatica is closely coupled to the rate of biosynthesis (Schuh et al., 1997). Emissions of sabinene, the main compound emitted from Fagus sylvatica, was shown to be light-depended and decreased below the detection limit in darkness whereas emission of another monoterpene, limonene, did not change in darkness (Table 1).

Here we present results, focussing on the monoterpenes α-pinene, sabinene and limonene, which give evidence that different carbon sources are used for the synthesis of the released terpenes and that the contribution of the sources vary depending on light condition and the particular terpenoid structure.

Section snippets

Results

After 66 min of exposure to $^{13} CO$ ₂ a fraction of the fragments in the mass spectrum of the emitted sabinene showed complete incorporation of $^{13} C$ into the carbon skeleton (Fig. 1). The high intensity of m/z 100 indicates that all carbons in the seven-carbon-fragment (C₇H₉⁺) are replaced by $^{13} C$ . Similarly, m/z 130 as well as m/z 146 could only result if all carbons from the nine-carbon-fragment and the ten-carbon fragment were $^{13} C$ . The rapid shift of all fragments to higher masses shown in Fig. 1

Discussion

Two different carbon sources exist for the de novo biosynthesis of monoterpenes in the leaves of Fagus sylvatica. For sabinene the de novo synthesized intermediates supply all of the carbon inside the monoterpene-skeleton (whereby 90% originates from recently assimilated CO₂, 10% from preformed intermediates) whereas 100% of the limonene molecules seem to come from a monoterpene pool. This is the first demonstration for intact plants, which gives a strong evidence that plant volatile terpenoid

Plant material

6 year old Fagus sylvatica plants were from commercial supplier. Plants were grown outside in 5 l pots in commercial soil under natural light conditions. Plants were watered the last time 12 h before the experiment.

Gas exchange and $^{13} CO$ ₂ administration

Three 6 year old Fagus sylvatica plants were placed 48 h before the experiment in a controlled plant enclosure chamber. The chamber allows the simulation of typical ambient conditions. The wall materials (PFA and a specific glass material (SANALUX©)) transmit the PAR and partly UV

Acknowledgements

The authors thank the Bundesministerium für Bildung, Wissenschaft, Forschung und Technologie for financial support within in the research program: “cycles of trace constituents”.

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Citation Excerpt :
Previously, Kahl et al. (1999) concluded that the contribution of the different sources of intermediates supplying the carbon for the terpene-skeleton varies not only between plant species for a selected molecule, but also within one class of compounds emitted from one plant species. A light dependency experiment carried out on a F. sylvatica L. under controlled growth room environmental conditions (varying PPFD at a fixed temperature of 27 ± 1 °C) showed supportive results for Kahl’s (Kahl et al., 1999) investigation. One could conclude from Fig. 2 that the synthesis of sabinene in the leaves of F. sylvatica L. is strongly linked to light intensity, but the release of α-farnesene was not influenced by PPFD above ∼40 μmol m−2 s−1.
Induced volatiles have been a focus of recent research, as not much is known of their emission behavior or atmospheric contribution. BVOC emissions were measured from Pseudotsuga menziesii saplings under natural environmental conditions, using a dynamic branch enclosure system and GC–MS for their analysis. We determined temperature and light dependency of the individual compounds, studied seasonality of the emissions and discuss the effect of heat stress in comparison with two specific biotic stresses that occurred naturally on the trees.
A standardized emission rate of 6.8 μg g_(dw)⁻¹ h⁻¹ for monoterpenes under stressed conditions was almost a magnitude higher than that obtained for healthy trees (0.8 ± 0.2 μg g_(dw)⁻¹ h⁻¹), with higher beta factors characterizing the stressed trees. The response of the emissions to light intensity was different for the individual compounds, suggesting a distinct minimum light intensity to reach saturation. Heat stress changed the relative contribution of specific volatiles, with larger extent of increase of sesquiterpenes, methyl salicylate and linalool emissions compared to monoterpenes. Biotic stress kept low the emissions of sesquiterpenes, (E)-4,8-dimethyl-1,3,7-nonatriene and methylbutenol isomers, and increased the level of methyl salicylate and monoterpenes. The ratio of β-pinene/α-pinene was also found to be significantly enhanced from 1.3 to 2.4 and 3.2 for non-stressed, heat stressed and combined biotic and heat stressed, respectively.
History effect of light and temperature on monoterpenoid emissions from Fagus sylvatica L.
2010, Atmospheric Environment
Citation Excerpt :
In the following discussion, the corrected, daytime emission data are compared with literature data and tested against existing emission algorithms for newly synthesized monoterpenoids. The values of the daily standard emission factors (ɛG97, see Eq. (2)) inferred from the measured monoterpenoid emission rates of the second tree are found to vary between 100 and 1550 μg m−2 h−1 or 2 and 32 μg gDW−1 h−1, in good agreement with literature data reported for F. sylvatica L. in recent years (Schuh et al., 1997; Kahl et al., 1999; Spirig et al., 2005; Moukhtar et al., 2005; Dindorf et al., 2006), which have been recently compiled in Table 3 of Dindorf et al. (2006), and with the values adopted in the two most recent European plant-specific BVOC inventories, 22.1 μg gDW−1 h−1 (Karl et al., 2009) and 10.0 μg gDW−1 h−1 (Schurgers et al., 2009). The temporal variation of the standard emission factors ɛG97 for the second tree is shown in Fig. 4.
Monoterpenoid emissions from Fagus sylvatica L. trees have been measured at light- and temperature-controlled conditions in a growth chamber, using Proton Transfer Reaction Mass Spectrometry (PTR-MS) and the dynamic branch enclosure technique.
De novo synthesized monoterpenoid Standard Emission Factors, obtained by applying the G97 algorithm (Guenther, 1997), varied between 2 and 32 μg g_DW⁻¹ h⁻¹ and showed a strong decline in late August and September, probably due to senescence.
The response of monoterpenoid emissions to temperature variations at a constant daily light pattern could be well reproduced with a modified version of the MEGAN algorithm (Guenther et al., 2006), with a typical dependence on the average temperature over the past five days.
The diurnal emissions at constant temperature showed a typical hysteretic behaviour, which could also be adequately described with the modified MEGAN algorithm by taking into account a dependence on the average light levels experienced by the trees during the past 10–13 h.
The impact of the past light and temperature conditions on the monoterpenoid emissions from F. sylvatica L. was found to be much stronger than assumed in previous algorithms.
Since our experiments were conducted under low light intensity, future studies should aim at confirming and completing the proposed algorithm updates in sunny conditions and natural environments.
Variation in biogenic volatile organic compound emission pattern of Fagus sylvatica L. due to aphid infection
2010, Atmospheric Environment
Citation Excerpt :
Since some compounds or their specific isomers were not available as standards, their retention indices were compared to literature data. F. sylvatica L. has been characterized as an MT emitter (König et al., 1995; Schuh et al., 1997; Kahl et al., 1999; Dindorf et al., 2005; Moukhtar et al., 2005; Holzke et al., 2006a). Sabinene, α-pinene, β-pinene and limonene are the most frequently noted compounds from beech.
Volatile organic compounds (VOCs) have been the focus of interest to understand atmospheric processes and their consequences in formation of ozone or aerosol particles; therefore, VOCs contribute to climate change. In this study, biogenic VOCs (BVOCs) emitted from Fagus sylvatica L. trees were measured in a dynamic enclosure system. In total 18 compounds were identified: 11 monoterpenes (MT), an oxygenated MT, a homoterpene (C₁₄H₁₈), 3 sesquiterpenes (SQT), isoprene and methyl salicylate. The frequency distribution of the compounds was tested to determine a relation with the presence of the aphid Phyllaphis fagi L. It was found that linalool, (E)-β-ocimene, α-farnesene and a homoterpene identified as (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), were present in significantly more samples when infection was present on the trees. The observed emission spectrum from F. sylvatica L. shifted from MT to linalool, α-farnesene, (E)-β-ocimene and DMNT due to the aphid infection. Sabinene was quantitatively the most prevalent compound in both, non-infected and infected samples. In the presence of aphids α-farnesene and linalool became the second and third most important BVOC emitted. According to our investigation, the emission fingerprint is expected to be more complex than commonly presumed.
Chemotaxonomical investigations of fossil and extant beeches. I. Leaf lipids from the extant Fagus sylvatica L.
2007, Comptes Rendus - Palevol
The chemistry of fossil plants can help in precising their taxonomic affinities as well as their taphonomic history. The lipids from mature leaves of Fagus sylvatica L. were thus characterised to identify the components potentially informative that might be preserved in fossil beeches. Gas chromatography–mass spectrometry analyses of the lipids revealed a complex mixture comprising more than 60 identified constituents, belonging to diverse chemical families (e.g., sterols, fatty lipids, acyclic isoprenoids, triterpenes, glycerols, esters). Although most of the identified components are reported here for the first time in the European beech, they predominantly correspond to rather common compounds in Angiosperms. Nevertheless, their co-occurrence may constitute a useful fingerprint in further chemotaxonomic investigations of beeches. The newly reported compounds also include degradation products of plant lipids and fungal markers, showing that these degradation markers are to be considered as molecules related to the leaves in further taphonomic studies.
La chimie des plantes fossiles peut aider à préciser leurs affinités taxinomiques et leur histoire taphonomique. Les lipides de feuilles matures de Fagus sylvatica L. ont donc été caractérisés, afin d’identifier les composés potentiellement informatifs qui pourraient être préservés dans les hêtres fossiles. L’analyse de ces lipides par spectrométrie de masse a révélé une mixture complexe de plus de 60 molécules identifiées, appartenant à des familles chimiques variées. La plupart des composés identifiés sont décrits ici pour la première fois chez F. sylvatica. Ils correspondent majoritairement à des constituants communs chez les Angiospermes, mais leur présence concomitante pourrait constituer une empreinte chimique pertinente pour les futures études chimiotaxonomiques sur le hêtre. Certains des composés nouvellement identifiés correspondent à des produits de dégradation de lipides végétaux et à des marqueurs fongiques, montrant que de tels marqueurs de dégradation peuvent contribuer significativement aux lipides des fossiles.
Techniques for gas chromatography of volatile terpenoids from a range of matrices
2001, Journal of Chromatography A
The commercial importance of the volatile mono- and sesqui-terpenoids has resulted in a wide range of techiques being used for extraction, concentration, chromatography, and characterisation of constituents. The major chromatographic technique is gas chromatography, and tandem techniques of chromatography linked to further chromatography and spectroscopy, allow much increased resolution, and greater ease of characterisation of terpenes. A wide range of extraction techniques are discussed, and suitability for particular matrices and sample sizes outlined. Chromatography operating conditions and stationary phases, and techniques for solute identification are laid out. A number of applications of terpene analysis in many different matrices are discussed.
Seasonal variation in biogenic volatile organic compound (BVOC) emissions from Norway spruce in a Swedish boreal forest
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Differentiation between de novo synthesized and constitutively released terpenoids from Fagus sylvatica

Abstract

Introduction

Section snippets

Results

Discussion

Plant material

Gas exchange and 13CO2 administration

Acknowledgements

FEBS Letters

Plant Cell Environ.

Netherland J. of Zoology

Global Biogeochemical Cycles

J. Geophys. Res.

Fres. J. Anal. Chem.

Proceedings of the EUROTRAC Symposium

Proceedings of the National Academy of Science of the USA

J. Plant Physiol.

Gas exchange and $^{13} CO$ ₂ administration