Structural Characterization of 6-Halo-6-Deoxycelluloses by Direct-Dissolution Solution-State NMR Spectroscopy.

Regioselective modifications of cellulose using activated cellulose derivatives such as 6-halo-6-deoxycelluloses provide a convenient approach for developing sustainable products with properties tailored to specific applications. However, maintaining precise regiochemical control of substituent distribution in 6-halo-6-deoxycelluloses is challenging due to their insolubility in most common solvents and the resulting difficulties in precise structure elucidation by modern instrumental analytical techniques. Herein, an accessible NMR-based approach toward detailed characterization of 6-halo-6-deoxycelluloses, including the determination of the degrees of substitution at carbon 6 (DS6), is presented. It is shown that the direct-dissolution cellulose solvent, tetrabutylphosphonium acetate:DMSO-d6, converts 6-halo-6-deoxycelluloses to 6-monoacetylcellulose, enabling in situ solution-state NMR measurements. A range of 1D and 2D NMR experiments is used to demonstrate the quantitivity of the conversion and provide optimum dissolution conditions. In comparison with other NMR-based derivatization protocols for elucidating the structure of 6-halo-6-deoxycelluloses, the presented approach offers major advantages in terms of accuracy, speed, and simplicity of analysis, and minimal requirements for reagents or NMR instrumentation.

Estimating intraseasonal intrinsic water-use efficiency from high-resolution tree-ring δ13 C data in boreal Scots pine forests.

Intrinsic water-use efficiency (iWUE), a key index for carbon and water balance, has been widely estimated from tree-ring δ13 C at annual resolution, but rarely at high-resolution intraseasonal scale. We estimated high-resolution iWUE from laser-ablation δ13 C analysis of tree-rings (iWUEiso ) and compared it with iWUE derived from gas exchange (iWUEgas ) and eddy covariance (iWUEEC ) data for two Pinus sylvestris forests from 2002 to 2019. By carefully timing iWUEiso via modeled tree-ring growth, iWUEiso aligned well with iWUEgas and iWUEEC at intraseasonal scale. However, year-to-year patterns of iWUEgas , iWUEiso , and iWUEEC were different, possibly due to distinct environmental drivers on iWUE across leaf, tree, and ecosystem scales. We quantified the modification of iWUEiso by postphotosynthetic δ13 C enrichment from leaf sucrose to tree rings and by nonexplicit inclusion of mesophyll and photorespiration terms in photosynthetic discrimination model, which resulted in overestimation of iWUEiso by up to 11% and 14%, respectively. We thus extended the application of tree-ring δ13 C for iWUE estimates to high-resolution intraseasonal scale. The comparison of iWUEgas , iWUEiso , and iWUEEC provides important insights into physiological acclimation of trees across leaf, tree, and ecosystem scales under climate change and improves the upscaling of ecological models.

Drivers of intra-seasonal δ13 C signal in tree-rings of Pinus sylvestris as indicated by compound-specific and laser ablation isotope analysis.

Carbon isotope composition of tree-ring (δ13 CRing ) is a commonly used proxy for environmental change and ecophysiology. δ13 CRing reconstructions are based on a solid knowledge of isotope fractionations during formation of primary photosynthates (δ13 CP ), such as sucrose. However, δ13 CRing is not merely a record of δ13 CP . Isotope fractionation processes, which are not yet fully understood, modify δ13 CP during sucrose transport. We traced, how the environmental intra-seasonal δ13 CP signal changes from leaves to phloem, tree-ring and roots, for 7 year old Pinus sylvestris, using δ13 C analysis of individual carbohydrates, δ13 CRing laser ablation, leaf gas exchange and enzyme activity measurements. The intra-seasonal δ13 CP dynamics was clearly reflected by δ13 CRing , suggesting negligible impact of reserve use on δ13 CRing . However, δ13 CP became increasingly 13 C-enriched during down-stem transport, probably due to post-photosynthetic fractionations such as sink organ catabolism. In contrast, δ13 C of water-soluble carbohydrates, analysed for the same extracts, did not reflect the same isotope dynamics and fractionations as δ13 CP , but recorded intra-seasonal δ13 CP variability. The impact of environmental signals on δ13 CRing , and the 0.5 and 1.7‰ depletion in photosynthates compared ring organic matter and tree-ring cellulose, respectively, are useful pieces of information for studies exploiting δ13 CRing .

Estimating intra-seasonal photosynthetic discrimination and water use efficiency using δ13C of leaf sucrose in Scots pine.

Sucrose has a unique role in recording environmental and physiological signals during photosynthesis in its carbon isotope composition (δ13C) and transport of the signal to tree rings. Yet, instead of sucrose, total organic matter (TOM) or water-soluble carbohydrates (WSC) are typically analysed in studies that follow δ13C signals within trees. To study how the choice of organic material may bias the interpretation of δ13C records, we used mature field-grown Scots pine (Pinus sylvestris) to compare for the first time δ13C of different leaf carbon pools with δ13C of assimilates estimated by a chamber-Picarro system (δ13CA_Picarro), and a photosynthetic discrimination model (δ13CA_model). Compared with sucrose, the other tested carbon pools, such as TOM and WSC, poorly recorded the seasonal trends or absolute values of δ13CA_Picarro and δ13CA_model. Consequently, in comparison with the other carbon pools, sucrose δ13C was superior for reconstructing changes in intrinsic water use efficiency (iWUE), agreeing in both absolute values and intra-seasonal variations with iWUE estimated from gas exchange. Thus, deriving iWUE and environmental signals from δ13C of bulk organic matter can lead to misinterpretation. Our findings underscore the advantage of using sucrose δ13C to understand plant physiological responses in depth.

Explicitly accounting for needle sugar pool size crucial for predicting intra-seasonal dynamics of needle carbohydrates δ18 O and δ13 C.

We explore needle sugar isotopic compositions (δ18 O and δ13 C) in boreal Scots pine (Pinus sylvestris) over two growing seasons. A leaf-level dynamic model driven by environmental conditions and based on current understanding of isotope fractionation processes was built to predict δ18 O and δ13 C of two hierarchical needle carbohydrate pools, accounting for the needle sugar pool size and the presence of an invariant pinitol pool. Model results agreed well with observed needle water δ18 O, δ18 O and δ13 C of needle water-soluble carbohydrates (sugars + pinitol), and needle sugar δ13 C (R2 = 0.95, 0.84, 0.60, 0.73, respectively). Relative humidity (RH) and intercellular to ambient CO2 concentration ratio (Ci /Ca ) were the dominant drivers of δ18 O and δ13 C variability, respectively. However, the variability of needle sugar δ18 O and δ13 C was reduced on diel and intra-seasonal timescales, compared to predictions based on instantaneous RH and Ci /Ca , due to the large needle sugar pool, which caused the signal formation period to vary seasonally from 2 d to more than 5 d. Furthermore, accounting for a temperature-sensitive biochemical 18 O-fractionation factor and mesophyll resistance in 13 C-discrimination were critical. Interpreting leaf-level isotopic signals requires understanding on time integration caused by mixing in the needle sugar pool.

Carbon flux from decomposing wood and its dependency on temperature, wood N2 fixation rate, moisture and fungal composition in a Norway spruce forest.

Globally 40-70 Pg of carbon (C) are stored in coarse woody debris on the forest floor. Climate change may reduce the function of this stock as a C sink in the future due to increasing temperature. However, current knowledge on the drivers of wood decomposition is inadequate for detailed predictions. To define the factors that control wood respiration rate of Norway spruce and to produce a model that adequately describes the decomposition process of this species as a function of time, we used an unprecedentedly diverse analytical approach, which included measurements of respiration, fungal community sequencing, N2 fixation rate, nifH copy number, 14 C-dating as well as N%, δ13 C and C% values of wood. Our results suggest that climate change will accelerate C flux from deadwood in boreal conditions, due to the observed strong temperature dependency of deadwood respiration. At the research site, the annual C flux from deadwood would increase by 27% from the current 117 g C/kg wood with the projected climate warming (RCP4.5). The second most important control on respiration rate was the stage of wood decomposition; at early stages of decomposition low nitrogen content and low wood moisture limited fungal activity while reduced wood resource quality decreased the respiration rate at the final stages of decomposition. Wood decomposition process was best described by a Sigmoidal model, where after 116 years of wood decomposition mass loss of 95% was reached. Our results on deadwood decomposition are important for C budget calculations in ecosystem and climate change models. We observed for the first time that the temperature dependency of N2 fixation, which has a major role at providing N for wood-inhabiting fungi, was not constant but varied between wood density classes due to source supply and wood quality. This has significant consequences on projecting N2 fixation rates for deadwood in changing climate.

Modeling the response of Norway spruce tree-ring carbon and oxygen isotopes to selection harvest on a drained peatland forest.

Continuous cover forestry (CCF) has gained interest as an alternative to even-aged management particularly on drained peatland forests. However, relatively little is known about the physiological response of suppressed trees when larger trees are removed as a part of CCF practices. Consequently, studies concentrating on process-level modeling of the response of trees to selection harvesting are also rare. Here, we compared, modeled and measured harvest response of previously suppressed Norway spruce (Picea abies) trees to a selection harvest. We quantified the harvest response by collecting Norway spruce tree-ring samples in a drained peatland forest site and measuring the change in stable carbon and oxygen isotopic ratios of wood formed during 2010-20, including five post-harvest years. The measured isotopic ratios were compared with ecosystem-level process model predictions for ${\kern0em }^{13}$C discrimination and ${\kern0em }^{18}$O leaf water enrichment. We found that the model predicted similar but lower harvest response than the measurements. Furthermore, accounting for mesophyll conductance was important for capturing the variation in ${\kern0em }^{13}$C discrimination. In addition, we performed sensitivity analysis on the model, which suggests that the modeled ${\kern0em }^{13}$C discrimination is sensitive to parameters related to CO2 transport through stomata to the mesophyll.

Progress in high-resolution isotope-ratio analysis of tree rings using laser ablation.

Stable isotope ratio analysis of tree rings has been widely and successfully applied in recent decades for climatic and environmental reconstructions. These studies were mostly conducted at an annual resolution, considering one measurement per tree ring, often focusing on latewood. However, much more information could be retrieved with high-resolution intra-annual isotope studies, based on the fact that the wood cells and the corresponding organic matter are continuously laid down during the growing season. Such studies are still relatively rare, but have a unique potential for reconstructing seasonal climate variations or short-term changes in physiological plant properties, like water-use efficiency. The reason for this research gap is mostly technical, as on the one hand sub-annual, manual splitting of rings is very tedious, while on the other hand automated laser ablation for high-resolution analyses is not yet well established and available. Here, we give an update on the current status of laser ablation research for analysis of the carbon isotope ratio (δ13C) of wood, describe an easy-to-use laser ablation system, its operation and discuss practical issues related to tree core preparation, including cellulose extraction. The results show that routine analysis with up to 100 laser shot-derived δ13C-values daily and good precision and accuracy (ca. 0.1‰) comparable to conventional combustion in an elemental analyzer are possible. Measurements on resin-extracted wood is recommended as most efficient, but laser ablation is also possible on cellulose extracted wood pieces. Considering the straightforward sample preparation, the technique is therefore ripe for wide-spread application. With this work, we hope to stimulate future progress in the promising field of high-resolution environmental reconstruction using laser ablation.

Spatio-Temporal Variations in Carbon Isotope Discrimination Predicted by the JULES Land Surface Model.

Stable carbon isotopes in plants can help evaluate and improve the representation of carbon and water cycles in land-surface models, increasing confidence in projections of vegetation response to climate change. Here, we evaluated the predictive skills of the Joint UK Land Environmental Simulator (JULES) to capture spatio-temporal variations in carbon isotope discrimination (Δ13C) reconstructed by tree rings at 12 sites in the United Kingdom over the period 1979-2016. Modeled and measured Δ13C time series were compared at each site and their relationships with local climate investigated. Modeled Δ13C time series were significantly correlated (p < 0.05) with tree-ring Δ13C at eight sites, but JULES underestimated mean Δ13C values at all sites, by up to 2.6‰. Differences in mean Δ13C may result from post-photosynthetic isotopic fractionations that were not considered in JULES. Inter-annual variability in Δ13C was also underestimated by JULES at all sites. While modeled Δ13C typically increased over time across the UK, tree-ring Δ13C values increased only at five sites located in the northern regions but decreased at the southern-most sites. Considering all sites together, JULES captured the overall influence of environmental drivers on Δ13C but failed to capture the direction of change in Δ13C caused by air temperature, atmospheric CO2 and vapor pressure deficit at some sites. Results indicate that the representation of carbon-water coupling in JULES could be improved to reproduce both the trend and magnitude of interannual variability in isotopic records, the influence of local climate on Δ13C, and to reduce uncertainties in predicting vegetation-environment interactions.

Tree organ growth and carbon allocation dynamics impact the magnitude and δ13C signal of stem and soil CO2 fluxes.

Incomplete knowledge of carbon (C) allocation dynamics in trees hinders accurate modeling and future predictions of tree growth. We studied C allocation dynamics in a mature Pinus sylvestris L. dominated forest with a novel analytical approach, allowing the first comparison of: (i) magnitude and δ13C of shoot, stem and soil CO2 fluxes (Ashoot, Rstem and Rsoil), (ii) concentration and δ13C of compound-specific and/or bulk non-structural carbohydrates (NSCs) in phloem and roots and (iii) growth of stem and fine roots. Results showed a significant effect of phloem NSC concentrations on tracheid growth, and both variables significantly impacted Rstem. Also, concentrations of root NSCs, especially starch, had a significant effect on fine root growth, although no effect of root NSC concentrations or root growth was detected on Rsoil. Time series analysis between δ13C of Ashoot and δ13C of Rstem or δ13C of Rsoil revealed strengthened C allocation to stem or roots under high C demands. Furthermore, we detected a significant correlation between δ13C of Rstem and δ13C of phloem sucrose and glucose, but not for starch or water-soluble carbohydrates. Our results indicate the need to include C allocation dynamics into tree growth models. We recommend using compound-specific concentration and δ13C analysis to reveal C allocation processes that may not be detected by the conventional approach that utilizes bulk organic matter.

Compound-specific carbon isotope patterns in needles of conifer tree species from the Swiss National Park under recent climate change.

Elevated CO2 along with rising temperature and water deficits can lead to changes in tree physiology and leaf biochemistry. These changes can increase heat- and drought-induced tree mortality. We aim to reveal the impacts of climatic drivers on individual compounds at the leaf level among European larch (Larix decidua) and mountain pine (Pinus mugo) trees, which are widely distributed at high elevations. We investigated seasonal carbon isotope composition (δ13C) and concentration patterns of carbohydrates and organic acids in needles of these two different species from a case study in the Swiss National Park (SNP). We found that average and minimum air temperatures were the main climatic drivers of seasonal variation of δ13C in sucrose and glucose as well as in concentrations of carbohydrates and citric acid/citrate in needles of both tree species. The impact of seasonal climatic drivers on larch and mountain pine trees at the needle level is in line with our earlier study in this region for long-term changes at the tree-ring level. We conclude that the species-specific changes in δ13C and concentrations of carbohydrates and organic acids are sensitive indicators of changes in the metabolic pathways occurring as a result of climatic changes.