Carbon-based Secondary Metabolites Research Articles

Nitrogen addition modifies the relative gene expression level and accumulation of carbon-based bioactive substances in Cyclocarya paliurus

In China, lots of Cyclocarya paliurus plantations have been established for tea and functional food production on nitrogen (N)-limited land. The optimum N levels require for biosynthesis and accumulation of carbon-based bioactive substances vary among plant species. This study integrated field trial with hydroponic culture to assess impact of nitrogen addition on accumulation and relative gene expression level of carbon-based secondary metabolites in C. paliurus. N addition significantly influenced not only contents of polyphenols, flavonoids and triterpenoids and relative gene expression levels of their biosynthetic pathway in C. paliurus leaves but also leaf biomass production and the bioactive substance accumulations. An intermediate N addition induced the highest contents of polyphenols, flavonoids and triterpenoids in leaves, but the optimized accumulation of these bioactive substances in the leaves was the trade-off between their contents and leaf biomass production. Correlation analysis showed that related gene expression levels were closely correlated with contents of their leaf corresponding secondary metabolites. Compared with ratios of carbon/N (C/N) and carbon/phosphorus (C/P) in the soil, ratios of C/N and C/P in the leaves were more strongly related to the contents and accumulations of polyphenols, flavonoids and triterpenoids. To obtain higher yields of targeted phytochemicals, the threshold ratios of C/N and C/P in the leaves are recommended for N and P fertilization at similar sites. Overall, our findings would provide the theoretical basis and technical support for manipulating N fertilization in C. paliurus plantations to obtain higher accumulations of targeted bioactive substances.

Effects of phytochemical crusts formed under two desert shrubs on physical properties of soils in arid ecosystems

Xerophytic shrubs are usually dominant in arid environments. Because of environmental pressures both biotic and abiotic, many plants have developed a huge diversity of carbon based secondary metabolites (CBSM). Several functions have been found for these CBSM, although their fate after being released is not clear. One of the hypothesis, tested here, is that CBSM accumulate in the soil under the plant's canopy. Particularly, we tested the hypothesis that terpene and polyphenolic resins present in Grindelia chiloensis (Asteraceae) and Larrea divaricata (Zigophyllaceae) would accumulate in the soil under their canopies and change its physical properties. Our results show that resins accumulate in the soil, and increase hardiness and hydrophobicity, while they reduce water infiltration. These changes in soil properties, compared to non-CBSM producing shrubs and bare soil, were more evident close to the plants' center and decreased towards the edge and surrounding soil. As they alter water dynamics, chemical crusts may have consequences both to the individual plant and to the ecosystem, and should be present in other arid lands, outside Patagonia.

Effect of crop load on fruit nutritional and nutraceutical traits of greenhouse tomato

ABSTRACT This study aimed to evaluate the effect of crop load on the fruit quality traits especially nutritional and nutraceutical properties of two cultivars of greenhouse tomato (Solanum lycopersicum L.) i.e. ‘Grandella’ and ‘Isabella’. The crop load treatments were: saving one fruit per truss (1F), two fruits per truss (2F), three fruits per truss (3F), and no fruit thinning (control). The results showed that decreasing crop load increased the dry matter content, glucose, fructose, and sucrose concentrations of fruits because of less competition between fruits of each truss and higher relative expression level of tomato hexose transporter genes (LeHTs). Higher transport of sugars toward fruits of 1F resulted in high secondary metabolites concentration. The high secondary metabolites concentration with high fruit load in control that has low soluble sugar availability may result from the opportunity to take advantage of the excess pool of carbon to build cost-free carbon-based secondary metabolites. A reduced crop load would be a recommended management strategy to improve greenhouse tomato quality.

Metabolite Profiling Reveals Distinct Modulation of Complex Metabolic Networks in Non-Pigmented, Black, and Red Rice (Oryza sativa L.) Cultivars.

Comprehensive profiling of primary and secondary metabolites was performed to understand metabolic differences associated with color formation in pigmented rice (Oryza sativa L.). Overall, 110 metabolites from non-pigmented, black, and red rice cultivars were identified. Black and red rice contained high levels of flavonoids associated with plant color. Black rice also contained high levels of terpenoids (carotenoids, tocopherols, phytosterols, and monoterpenes). The non-pigmented rice contained relatively low levels of secondary metabolites. Multivariate and pathway analyses were performed to data-mine the metabolite profiles. Hierarchical clustering analysis of correlation coefficients revealed metabolite clusters based on nitrogen and carbon sources. These clusters suggested a negative correlation between nitrogen and carbon. Pathway analysis revealed that black rice was rich in carbon-based secondary metabolites, with relatively low levels of primary metabolites compared with other rice cultivars. These data highlight the complex interactions between nitrogen and carbon metabolism of primary and secondary metabolites in rice. For the first time, the relationships and metabolic differences in terpenoid content (monoterpenes, triterpenes, and tetraterpenes) of non-pigmented and pigmented rice cultivars were analyzed. These findings should greatly contribute to the understanding of pigmented rice metabolome and inform breeding programs for new rice cultivars.

Effects of Soil Fertilization on Terpenoids and Other Carbon-Based Secondary Metabolites in Rosmarinus officinalis Plants: A Comparative Study.

Rosmarinus officinalis is an evergreen aromatic plant with important commercial interest as it contains numerous essential oils (composed of terpenoid compounds) and phenolic constituents (natural antioxidant compounds). This work aims at evaluating the concomitant effects of different inorganic and organic fertilization treatments and the subsequent increases in soil nutrient availability on terpenoids and other carbon-based secondary metabolites, e.g., flavonoids and phenolic compounds, in Rosmarinus officinalis leaves. The results showed that, as expected, the structural carbohydrate content (lignocellulosic compounds) in stems was higher in fertilized plants than in controls. Additionally, positive correlations were observed of the absolute amounts of total terpenoids and some single terpenoid compounds with N or P contents in leaves. On the contrary, the phenolic and flavonoid concentrations in all the rosemary plant parts were lower with the fertilization treatments. Indeed, negative correlations between the phenolic compounds (and flavonoids) and N in rosemary leaves were also found. Overall, the results suggest that the terpenoid production’s response to fertilization was due to N, which is essential for protein synthesis and terpene synthase activity, and to P, which is necessary for the synthesis of both terpenoid precursors and ATP and NADPH, also needed for terpenoid synthesis. On the other hand, the basis for the fertilization’s effects on the production of phenolic compounds is the direct nitrogen trade-off between growth and the shikimic acid pathway by which phenolics compounds are synthesized.

Whole-Plant Metabolic Allocation Under Water Stress.

Trade-offs between plant growth and defense depend on environmental resource availability. Plants are predicted to prioritize growth when environmental resources are abundant and defense when environmental resources are scarce. Nevertheless, such predictions lack a whole-plant perspective—they do not account for potential differences in plant allocation above- and belowground. Such accounting is important because leaves and roots, though both critical to plant survival and fitness, differ in their resource-uptake roles and, often, in their vulnerability to herbivores. Here we aimed to determine how water availability affects plant allocation to multiple metabolic components of growth and defense in both leaves and roots. To do this, we conducted a meta-analysis of data from experimental studies in the literature. We assessed plant metabolic responses to experimentally reduced water availability, including changes in growth, nutrients, physical defenses, primary metabolites, hormones, and other secondary metabolites. Both above- and belowground, reduced water availability reduced plant biomass but increased the concentrations of primary metabolites and hormones. Importantly, however, reduced water had opposite effects in different organs on the concentrations of other secondary metabolites: reduced water increased carbon-based secondary metabolites in leaves but reduced them in roots. In addition, plants suffering from co-occurring drought and herbivory stresses exhibited dampened metabolic responses, suggesting a metabolic cost of multiple stresses. Our study highlights the needs for additional empirical studies of whole-plant metabolic responses under multiple stresses and for refinement of existing plant growth-defense theory in the context of whole plants.

Tree growth indicates resource quality for foliage-feeding insects: Pattern and structure of herbivore diversity in response to productivity

In forests, local site conditions can affect both trees and herbivores and hence site-related factors act indirectly on herbivores mediated by tree growth rates. Here, tree foliage represents a fundamental prerequisite for insect herbivore development providing energy in the form of plant tissue quality. Resource-based theories, on the other hand, assume that the synthesis of defensive compounds is a trade-off with growth and peaks at low resource availability. However, the extent to which plant tissue quality in response to site productivity is relevant in the species-energy relationship is unknown. Therefore, we aimed at a better understanding of the form and structure of the species-energy relationship in forest insects. We used census data of foliage-feeding insects along a productivity gradient of Scots pine forests defined by relative growth rates of trees (RGR). As a result, diversity monotonically increases with decreasing RGR (as a proxy for energy) during almost two decades of sampling. Herbivore assemblages become more similar with available energy as species turnover linearly decreases and proportions of sites occupied by individual species rise. The results suggest that tree growth rate influences herbivore dynamics in this system by altering the chemical composition of needles, without necessarily affecting the form in the relationship. The site-specific resource availability requires trees to adjust their allocation to synthesis of carbon-based secondary metabolites or growth, which then results in fundamental differences in herbivore dynamics at low vs. lowest RGR (regular cycles (dominance) vs. dampened cycles (evenness)). However, these differences inevitably demonstrate that species richness is not necessarily a result of more individuals and implicate that different mechanisms are involved (facilitation vs. competition/temporal heterogeneity). The resulting pattern and structure of foliage-feeding insects advance our understanding of herbivore dynamics in response to site quality and tree growth, which may ultimately improve our knowledge of plant-insect interactions in the face of environmental change.

Seasonal variations in nutrients and secondary metabolites in semi-arid savannas depend on year and species

Semi-arid savannas are characterised by alternating wet and dry seasons and large inter-annual rainfall fluctuations that affect plant growth. Carbon-based secondary metabolites (CBSMs) vary inversely with growth and nutrients because of the physiological trade-off between cellular growth and differentiation. We predicted that (1) nutrient concentrations decrease during the wet season, (2) CBSM concentrations increase during the wet season and (3) nutrient concentrations are lowest and CBSM concentrations are highest in the dry season. We measured nitrogen, phosphorus, total polyphenols and condensed tannins in six woody species (including one evergreen) seasonally at the Nkuhlu exclosure, Kruger National Park, South Africa, for three consecutive years, including one ‘wet’ year (above-average rainfall) and two ‘dry’ years (below-average rainfall). Neither N nor P consistently decreased during wet seasons, while CBSMs did not consistently increase. Neither N nor P in the evergreen species was consistently lowest in dry seasons, while CBSMs were not consistently highest in dry seasons. We discuss the inconsistent responses in the context of species-specificity and high inter-annual rainfall variation. We conclude that seasonal variations in N, P and CBSMs in semi-arid savannas cannot be easily generalised because they depend on species and year.

A potential trade-off with stictic acid improves ascospore viability inXanthoparmelia cumberlandia

Abstract The goal of this paper was to explore trade-offs between fecundity and carbon based secondary metabolites (CBSM) production within the thallus of three species of Rock-shield and Ring lichens, Arctoparmelia centrifuga, Xanthoparmelia viriduloumbrina and X. cumberlandia. Fecundity was measured by numbers of apothecia per thallus, number of spores per apothecium and percent spore germination, and it was compared with the quantity of the major CBSMs produced by the thallus. Results showed negative correlations between stictic acid and all three measures of fecundity for X. cumberlandia, which are consistent with trade-offs. When thallus and apothecium quantities of CBSMs were compared, stictic acid was the only CBSM with levels in the apothecium that were significantly higher than those in the thallus supporting a function that may protect ascospores against herbivory. Other CBSMs are hypothesized to have functions associated with the thallus and UV protection of green algae.

Nutrient and secondary metabolite concentrations in a savanna are independently affected by large herbivores and shoot growth rate

Carbon-based secondary metabolites (CBSMs) such as tannins are assumed to function as plant defences against herbivores. CBSMs are thought to be inversely related to growth rate and nutrient concentrations because a physiological trade-off exists between cellular growth and differentiation, but CBSM concentrations can be altered by herbivory-induced changes in the trade-off. We predicted that a significant interaction exists between herbivory and growth phase, such that the effects of large herbivores (or their exclusion) on nutrient or CBSM concentrations are greatest during phases of rapid shoot or leaf growth. Leaf samples were collected during phases of different growth rate from six woody species 4 years after establishment of a large-scale long-term herbivore exclusion experiment in Kruger National Park, South Africa. Samples were analysed for N, P, condensed tannins and total phenolics. Interactions between growth phase and herbivores were rare. However, the assumption that elevated nutrients and reduced CBSMs occurs during fast phases of growth was supported by four species (consistent with the growth-differentiation balance hypothesis), but not the other two. Large herbivores generally did not affect nutrients, but CBSMs in four species were reduced by large herbivores other than elephants, while CBSMs in two species were reduced by elephants. Carbon limitation ultimately prevailed among woody plants taller than 2 m under long-term browsing. Large herbivores and plant growth phase are independent and important determinants of nutrients or CBSMs in African savannas, but the effects depend on the interacting assemblages of species, which poses challenges to the application of current general hypotheses of plant defence.

Does large herbivore removal affect secondary metabolites, nutrients and shoot length in woody species in semi-arid savannas?

We hypothesised that exclusion of long-term browsing leads to decreased nutrient concentrations, increased carbon-based secondary metabolites (CBSMs) and decreased shoot length in woody plants in semi-arid savannas. To test this, we sampled four deciduous woody species in a large-scale, long-term exclusion experiment. We found a tendency for large herbivore exclusion to either decrease nitrogen or increase tannin in two tree species. Phosphorus and total polyphenol responses indicated complex interactions with herbivore assemblage. Shoot length decreased under exclusion in two species, while a similar tendency existed in the remaining species. Therefore, we found limited support for our predictions that exclusion of browsing results in reduced nitrogen or increased CBSMs, but moderate support for decreased shoot length in semi-arid savannas. Dichrostachys cinerea, a fast-growing, aggressive invader in savannas, displayed C-limitation with removal of large herbivores because both shoot length and CBSMs decreased. In conclusion, effects of long-term browsing on deciduous trees in semi-arid savannas depend on the plant species, specific nutrients or CBSMs measured and the composition of the herbivore assemblages. Because elephant densities in the study area were high and most responses were associated with their exclusion, we conclude that elephants are important drivers of browse quality in the study area.

Sex influences the taxanes content in Taxus baccata

Like other species of the genus Taxus, European yew trees contain taxanes, including paclitaxel (T) and its precursor 10-deacetylbaccatin III (10-DAB). Taxanes are one of the most effective anticancer drugs. This study was undertaken to describe the levels and patterns of taxane variation in the Sudetian region (SW Poland). Paclitaxel (T) and 10-deacetylbaccatin III (10-DAB) concentrations were analysed in five populations. Needles and twigs were analysed from 60 individuals (30 males and 30 females) in each population. In addition, morphometric measurements were taken in the populations to obtain light intensity coefficients (specific leaf area, SLA). High variability in the taxane contents at both intra and interpopulational levels was found. Nevertheless, females had a significantly higher taxane content compared to males. Because taxanes are carbon-based secondary metabolites, females have higher rate of gas exchange of females compared to males. This was probably an adaptation to greater reproductive effort incurred by females. In this regard, female individuals seem to be better for selecting elite cultivars with a higher taxane production. The relationship between light intensity and taxane content was not significant. Shading, important for optimizing crop production, should not reduce the concentration of taxanes.

Pomegranate (Punica granatum L.) fruit response to different deficit irrigation conditions

No information exits on the effects of water stress on pomegranate (Punica granatum L.) fruits. In this paper, the influence of three irrigation treatments on the physical and chemical characteristics of the fruits of adult pomegranate trees was studied. Control (T0) plants were drip irrigated (60% ETo from the beginning of the season to the end of the first half of linear fruit growth phase, 117% ETo during the second half of linear fruit growth phase and 99% ETo during the end of fruit growth and ripening phase) in order to guarantee non-limiting soil water conditions, T1 plants were drip irrigated according to the criteria frequently used by the growers in the area (32% ETo from the beginning of the season to the end of the first half of linear fruit growth phase, 74% ETo during the second half of linear fruit growth phase and 36% ETo during the end of fruit growth and ripening phase) and T2 plants were irrigated as T1, except during the second half of linear fruit growth phase, in which irrigation was withheld. To ensure the recovery of T2 plants re-irrigation was performed at the levels used in T0. T1 plants achieved a moderate water stress level, whereas T2 plants achieved a more pronounced water stress level, from which they completely recovered when plants were rewatered. During ripening the peel of pomegranate fruits changes to show higher luminosity and greater red saturation. Also, the colour of the arils changes to a more perceptible red colour as a consequence of the increasing total anthocyanin content. However, neither the intense red colour of the arils nor their total phenolic compounds content was correlated with the juice antioxidant capacity. Considerable differences were observed in the response of pomegranate fruits to both deficit irrigation treatments. In this sense, fruits from T1 plants showed a decrease in fruit growth, leading to a lower final fruit size and lower total yield, and some changes in colour and chemical characteristics, which reflected earlier ripening. In contrast, a more pronounced water stress during the second half of the fruit growth phase (T2) was more critical for fruit size than for the chemical characteristics of the fruit, probably because under this situation carbon assimilation should be allocated to the synthesis of primary metabolites, which did not exceeded the amount used for fruit growth to the detriment of the synthesis of carbon-based secondary metabolites.

Reduced photoinhibition under low irradiance enhanced Kacip Fatimah (Labisia pumila Benth) secondary metabolites, phenyl alanine lyase and antioxidant activity.

A randomized complete block design experiment was designed to characterize the relationship between production of total flavonoids and phenolics, anthocyanin, photosynthesis, maximum efficiency of photosystem II (Fv/Fm), electron transfer rate (Fm/Fo), phenyl alanine lyase activity (PAL) and antioxidant (DPPH) in Labisia pumila var. alata, under four levels of irradiance (225, 500, 625 and 900 μmol/m2/s) for 16 weeks. As irradiance levels increased from 225 to 900 μmol/m2/s, the production of plant secondary metabolites (total flavonoids, phenolics and antocyanin) was found to decrease steadily. Production of total flavonoids and phenolics reached their peaks under 225 followed by 500, 625 and 900 μmol/m2/s irradiances. Significant positive correlation of production of total phenolics, flavonoids and antocyanin content with Fv/Fm, Fm/Fo and photosynthesis indicated up-regulation of carbon-based secondary metabolites (CBSM) under reduced photoinhibition on the under low light levels condition. At the lowest irradiance levels, Labisia pumila extracts also exhibited a significantly higher antioxidant activity (DPPH) than under high irradiance. The improved antioxidative activity under low light levels might be due to high availability of total flavonoids, phenolics and anthocyanin content in the plant extract. It was also found that an increase in the production of CBSM was due to high PAL activity under low light, probably signifying more availability of phenylalanine (Phe) under this condition.

Effects of Nitrogen Fertilization on Synthesis of Primary and Secondary Metabolites in Three Varieties of Kacip Fatimah (Labisia Pumila Blume)

A split plot 3 by 4 experiment was designed to examine the impact of 15-week variable levels of nitrogen fertilization (0, 90, 180 and 270 kg N/ha) on the characteristics of total flavonoids (TF), total phenolics (TP), total non structurable carbohydrate (TNC), net assimilation rate, leaf chlorophyll content, carbon to nitrogen ratio (C/N), phenyl alanine lyase activity (PAL) and protein content, and their relationships, in three varieties of Labisia pumila Blume (alata, pumila and lanceolata). The treatment effects were solely contributed by nitrogen application; there was neither varietal nor interaction effect observed. As nitrogen levels increased from 0 to 270 kg N/ha, the production of TNC was found to decrease steadily. Production of TF and TP reached their peaks under 0 followed by 90, 180 and 270 kg N/ha treatment. However, net assimilation rate was enhanced as nitrogen fertilization increased from 0 to 270 kg N/ha. The increase in production of TP and TF under low nitrogen levels (0 and 90 kg N/ha) was found to be correlated with enhanced PAL activity. The enhancement in PAL activity was followed by reduction in production of soluble protein under low nitrogen fertilization indicating more availability of amino acid phenyl alanine (phe) under low nitrogen content that stimulate the production of carbon based secondary metabolites (CBSM). The latter was manifested by high C/N ratio in L. pumila plants.

Secondary metabolites and nutrients of woody plants in relation to browsing intensity in African savannas

Carbon-based secondary metabolites (CBSMs) are assumed to function as defences that contribute to herbivore-avoidance strategies of woody plants. Severe browsing has been reported to reduce concentrations of CBSMs and increase N concentrations in individual plants, causing heavily browsed plants to be characterised by N-rich/C-poor tissues. We hypothesised that concentrations of condensed tannins (CT) and total polyphenols (TP) should decrease, or N increase, in relation to increasing intensity of browsing, rendering severely browsed plants potentially more palatable (increased N:CT) and less N-limited (increased N:P) than lightly browsed ones. We sampled naturally browsed trees (taller than 2m) of four abundant species in southern Kruger National Park, South Africa. Species-specific relationships between N:CT, CT, TP and P concentrations and increasing browsing intensity were detected, but N and N:P were consistently invariable. We developed a conceptual post-hoc model to explain diverse species-specific CBSM responses on the basis of relative allocation of C to total C-based defence traits (e.g. spines/thorns, tough/evergreen leaves, phenolic compounds). The model suggests that species with low allocation of C to C-based defence traits become C-limited (potentially more palatable) at higher browsing intensity than species with high allocation of C to C-based defences. The model also suggests that when N availability is high, plants become C-limited at higher browsing intensity than when N availability is low.

INFLUENCE OF IRRIGATION AND NITROGEN FERTILIZATION ON BROCCOLI POLYPHENOLICS CONCENTRATION

Polyphenolics are phytochemical compounds that may be beneficial or toxic to humans. They are found in fruits and vegetables known for their health promoting characteristics. These carbon-based secondary metabolites have roles such as protection against ultraviolet radiation, free radicals, oxidative stress, herbivores and pathogens. Their synthesis in plant material is therefore linked to both biotic and abiotic stress sources. Under nitrogen stress, for instance, an increase in polyphenolics concentration has been reported. This behaviour has been exploited in the design and marketing of the Dualex, a new diagnostic instrument aimed at establishing the in-season requirements of crops for nitrogen (N) fertilizer. The Dualex can estimate leaf polyphenolics concentrations from the measurement of ultraviolet (UV, 375 nm) absorption of the leaf epidermis by the double excitation of chlorophyll fluorescence. The value of the UV absorbance is directly connected to the amount of leaf polyphenolics. The objective of this study was to determine if irrigation and nitrogen fertilization can influence broccoli polyphenolics concentrations. A broccoli experimental field was established in 2005 at the Agriculture and Agri-Food Canada experimental farm of L'Acadie (Quebec). The crop was fertilized with either 0, 105, 135, 165, 195 or 225 kg N/ha and provided or not with supplementary irrigation after crop establishment. Four Dualex measurement campaigns were made, from 26 to 47 days after planting, as well as one laboratory estimation of polyphenolics. The absence of irrigation increased significantly polyphenolics concentrations on the two last dates (by 36 and 35%, respectively). The increase in fertilizer N was related to a linear or curvilinear decrease in polyphenolics. At the last date, unfertilized plots had 86 and 16% higher polyphenolics than fully fertilized ones, in irrigated and non-irriguated plots, respectively. Yields of marketable heads increased in a curvilinear matter with N rate, with a tendency to plateau at high rates. The percent of hollow stem was increased at high N rates, only under the irrigated treatment. Plots placed under conditions to produce the highest possible polyphenolics content were however the least productive ones, from a fresh market standpoint. Since these stresses are impacting crop growth and development, there is a clear antagonism between the production of polyphenolics from broccoli, on one hand, and fresh market production, on the other hand. The Dualex has the potential to establish broccoli N requirements by its capacity of detecting polyphenolics concentration in leaves but irrigation may interfere in the diagnosis.

Carbon-based secondary metabolites and internal nitrogen pools in Populus nigra under Free Air CO2 Enrichment (FACE) and nitrogen fertilisation

The goal of this study was to investigate whether increased nitrogen use efficiency found in Populus nigra L. (Jean Pourtet) under elevated CO2 would correlate with changes in the production of carbon-based secondary compounds (CBSCs). Using Free-Air CO2 Enrichment (FACE) technology, a poplar plantation was exposed to either ambient (about 370 µmol mol−1 CO2) or elevated (about 550 µmol mol−1 CO2) [CO2] for 5 years. After three growing seasons, the plantation was coppiced and half of the experimental plots were fertilized with nitrogen. CBSCs, total nitrogen and lignin-bound nitrogen were quantified in secondary sprouts in seasons of active growth and dormancy during 2 years after coppicing. Neither elevated CO2 nor nitrogen fertilisation alone or in combination influenced lignin concentrations in wood. Soluble phenolics and soluble proteins in wood decreased slightly in response to elevated CO2. Higher nitrogen supply stimulated formation of CBSCs and increased protein concentrations. Lignin-bound nitrogen in wood ranged from 0.37–1.01 mg N g−1 dry mass accounting for 17–26% of total nitrogen in wood, thus, forming a sizeable nitrogen fraction resistant to chemical degradation. The concentration of this nitrogen fraction was significantly decreased by elevated CO2, increased in response to nitrogen fertilisation and showed a significant CO2 × fertilisation interaction. Seasonal changes markedly affected the internal nitrogen pools. Soluble proteins in wood were 52–143% higher in the dormant than in the growth season. Positive correlations existed between the biosynthesis of proteins and CBSCs. The limited responses to elevated CO2 and nitrogen fertilisation indicate that growth and defence are well orchestrated in P. nigra and that changes in the balance of both resources—nitrogen and C—have only marginal effects on wood chemistry.

Long- and short-term induction of defences in seedlings of Shorea leprosula (Dipterocarpaceae): support for the carbon:nutrient balance hypothesis

The induction of carbon-based secondary metabolites in leaves following damage has been proposed to be a result of a shift in the carbon:nutrient balance, when growth is limited by nutrients in relation to carbon. Here we test this hypothesis using seedlings of a tropical tree, Shorea leprosula (Dipterocarpaceae). In the short term, we analysed the phenolic content of leaves 7 d after damage on seedlings grown under differing light and nutrient treatments. In the long term, we examined the effect of nutrients, over 12 mo, on leaf phenolic concentration and seedling growth. In both the long and short term, levels of phenolics increased in damaged leaves under low nutrient treatments. No changes in leaf phenolics were detected under high nutrient regimes, or in the short term under low light. In addition, it was found that defoliation of seedlings in high-nutrient environments led to greater rates of leaf production than in undamaged seedlings, suggesting compensation.

Terpene deployment in Eucalyptus polybractea; relationships with leafstructure, environmental stresses, and growth.

Terpene deployment was examined in a population of Eucalyptus polybractea (R.Baker) trees. Eucalyptus polybractea is a terpene-accumulating species, which stores terpenes in oil glands beneath the leaf surface. Using regression analysis, we showed that leaf thickness, measured as leaf mass per area (LMA), influenced terpene content, apparently through regulation of gland dimensions, and thus, gland volume. We also examined how environmental factors affected terpene content through regulation of both LMA, and therefore, storage capacity, and the supply of resources for terpene synthesis. Neither water stress, measured using carbon isotope ratios as an indicator, nor nutrient stress, measured as foliar nitrogen and phosphorus content, accounted for observed variation in either terpene content or LMA. Phenolic content, measured as a possible competing carbon sink, did not account for variation in terpene content, and variation in environmental stresses could not account for differences in growth rate. However, both terpenes and total carbon-based secondary metabolites (terpenes and phenolics) showed positive correlations with growth, suggesting plants gain a growth advantage by deploying greater amounts of secondary metabolites.