Late-season biosynthesis of leaf fatty acids and n-alkanes of a mature beech (Fagus sylvatica) tree traced via 13CO2 pulse-chase labelling and compound-specific isotope analysis

Abstract

Leaf cuticular waxes play an important role in reducing evapotranspiration via diffusion. However, the ability of mature trees to regulate the biosynthesis of waxes to changing conditions (e.g., drought, light exposition) remain an open question, especially during the late growing season. This holds also true for one of the most widely distributed trees in Central Europe, the European beech tree (Fagus sylvatica L.). In order to investigate the ongoing formation of wax constituents like alkanes and fatty acids, we conducted a ¹³CO₂ pulse-chase labelling experiment on sun-exposed and shaded branches of a mature beech tree during the late summer 2018. The ¹³C-label was traced via compound-specific δ¹³C isotope analysis of n-alkanes and fatty acids to determine the de-novo biosynthesis within these compound classes. We did not observe a significant change in lipid concentrations during the late growing season, but we found higher n-alkane concentrations in sun-exposed compared to shaded leaves in August and September. The n-alkane and fatty acid composition showed ongoing modifications during the late growing season. Together with the uptake and following subsequent decrease of the ¹³C-label, this suggests ongoing de-novo biosynthesis, especially of fatty acids in European beech leaves. Moreover, there is a high variability in the ¹³C-label among individual branches and between sun-exposed and shaded leaves. At the same time, sun-exposed leaves invest more of the assimilated C into secondary metabolites such as lipids than shaded leaves. This indicates that the investigated mature beech tree could adjust its lipid production and composition in order to acclimate to changes in microclimates within the tree crown and during the investigated period.

Keywords: 13CO2 labelling; Fagus sylvatica; compound-specific isotope analysis (CSIA); cuticular leaf waxes; fatty acids; n-alkanes.

Figure 1

Schematic overview of the labelling chamber (modified after Kagawa et al., 2005).

Figure 2

Change of the carbon (A) and nitrogen (B) concentrations of sun-exposed and shaded leaves during the late growing season (field replicates n=8 with 2 analytical replicates, each, error bars indicate SE).

Figure 3

Change of intracuticular (A) and cuticular (B) lipid concentrations of sun-exposed and shaded leaves during late growing season (field replicates n = 4, error bars indicate SE).

Figure 4

Change of fatty acids (intracuticular waxes) (A) and n-alkane (cuticular waxes) (B) concentrations of sun-exposed and shaded leaves during the late growing season (field replicates n=4, error bar indicate SE).

Figure 5

Natural δ¹³C abundance (control spot) of bulk tissue (A), fatty acids (intracuticular waxes) (B), and n-alkanes (cuticular waxes) (C) of sun-exposed and shaded leaves (analytical replicates n=3, error bars indicate SE).

Figure 6

Weighted average of ¹³C-excess of bulk C (A), fatty acids (intracuticular waxes) (B), and n-alkanes (intracuticular waxes) (C) of sun-exposed and shaded leaves after ¹³CO₂ pulse.

Figure 7

Weighted average of ¹³C-excess of short-chain fatty acids C_16-18 (A), long-chain fatty acids C_20:0-32:0 (B) of intracuticular waxes as well as n-alkanes C₂₅ (C), C₂₇ (D), and C₂₉ (E) of the cuticular waxes of sun-exposed and shaded leaves (analytical replicates n=3, error bars indicate SE). The vertical line in the graphs indicates the application date of the isotope label.