Periodic Reporting for period 4 - VOCO (Biochemical link between plant volatile organic compound (VOC) emissions and CO2 metabolism - from sub-molecular to ecosystem scales)
Reporting period: 2020-04-01 to 2021-09-30
The overall goal of VOCO is the development of a novel technological and theoretical basis to couple investigation of CO2 fluxes and VOC emissions, establishing a mechanistic linkage between primary and secondary carbon metabolism. VOCO will evaluate carbon investment into VOCs, respiratory CO2 emission and the associated isotope effects among species with different plant functional traits, bridging scales from sub-molecular to the whole-plant and ecosystem processes in an interdisciplinary approach.
This radically new approach uses stable isotope fractionation of central metabolites (glucose, pyruvate) to trace carbon partitioning at metabolic branching points. A unique combination of cutting-edge technology (δ13CO2 laser spectroscopy, high sensitivity PTR-TOF-MS and quantitative isotopic NMR spectroscopy) will allow a significant advancement through innovative leaf and whole-tree position-specific labelling experiments and an entire ecosystem assessment by merging complementary approaches.
Creation of a unique technological platform will allow an unprecedented assessment of carbon partitioning, bridging scales from sub-molecular to whole-plant and ecosystem processes in an interdisciplinary approach. Innovative positional 13C-labelling will break new ground quantifying real-time sub-molecular carbon investment into VOCs and CO2, enabling mechanistic descriptions of the underlying biochemical pathways coupling anabolic and catabolic processes, particularly the long overlooked link between secondary compound synthesis and CO2 emission in the light. This approach will permit the development of a novel mechanistic leaf model and its integration into a state-of-the-art ecosystem flux model.
VOCO will set a new dimension with a world-wide first ecosystem positional labelling experiment in the unique Biosphere 2 enclosure (Arizona, US). Jointly with the novel process-based ecosystem model, VOCO2 will open new frontiers for assessing biogenic emissions of greenhouse gases at the ecosystem scale. This will deliver important information for global change related aspects, as these greenhouse gases can impact atmospheric chemistry and enhance global warming.
Additionally, VOCs of selected samples are identified by thermal desorption-gas chromatograph-mass spectrometry (TD-GC-MS, Markes International, Ltd. and Agilent Technologies, Inc.) with detection limits of < 5 pptv for most VOCs in air, collected on thermal desorption sample tubes. The first experiments showed the vast potential of combining these methods. However, they also indicated that due to the rapid dynamics in VOC and CO2 fluxes (ranging from seconds to minutes) precise, coordinated, and synchronised measurements are required, which are not attainable within a conventional setup. Hard- and software of both instruments were coupled into one innovative device directly linked to an automated Teflon valve switching unit enabling synchronised sampling of 13CO2 and VOCs on several enclosures/atmospheric samples, calibration gases and background values. Moreover, a hydrocarbon-free zero-air-generator was developed, as commercial available instruments turned out to not meet requested pureness of air. This unique infrastructure (PTR-TOF-IRIS) for automated real-time VOC and 13CO2 analysis (Fig. 1) does now allow an unprecedented real-time detection of (13C)-carbon allocation into respiratory CO2 and VOC emission in intact plants and atmospheric trace gases and will be used throughout all subsequent experiments. Interest in this approach is also reflected by the invitation for key-note talk on world expert conferences on VOC and Isotopes (Gordon research conference, Spain 2016, Isotopes 2017, Switzerland). WP1 has now been successfully concluded and publication is in progress (Fasbender et al., under revision in PlosOne).
For work package 2, a series of experiments was started on different plant functional groups of high and low VOC-emitters of different biomes including tropical rainforest species of the dominant species of the Biosphere 2. Assessment of carbon fluxes into net carbon assimilation, respired δ13CO2 and VOC emissions are made throughout the day and after darkening at the leaf and branch level simultaneously in custom-built plant cuvettes (Fig. 1). Stress experiments testing the temperature and drought resistance of tropical and native species and regulation of carbon allocation in VOC for stress protection were conducted. Additionally, VOC samples from species in their natural environment as well as natural drought conditions were added to the data set (Manuscripts in preparation; Yanez-Serrano et al. 2018 Haberstroh et al. moderate revision in Frontiers).
Position-specific 13C labelled metabolites enable tracing the fate of every single carbon atom of the molecule through the metabolic pathways, but in spite of its enormous potential, the application is still rare. Yet a huge step forward can be attained when combining position-specific 13C labelling with the PTR-TOF-IRIS (WP1), which allows now real-time recording of carbon partitioning into different pathways such as respiratory CO2 and VOCs in intact plants. Moreover, using central metabolites like pyruvate which are linking anabolic and catabolic metabolism, real-time information on carbon partitioning between these pathways can be retrieved. A plethora of difference VOC can be emitted by plants, which are synthesized in different metabolic pathways and cell compartments. Experiments started to explore pathways with pyruvate (and acetyl-CoA) as precursor molecule. Many less-regarded volatile compounds have been detected the different set of experiments, as well as under natural conditions (Yanez-Serrano et al. 2018).
The radically new approach of VOCO2 is based on molecular isotope analysis and will deliver a mechanistic description and model of carbon partitioning into different biochemical pathways. At the metabolic scale, the innovative setup of position-specific labelling breaks new ground in tracing real-time carbon allocation into VOCs and CO2. First exciting results have been gain by detecting rare compounds such as volatile diterpenes emitted by plants (Yañez-Serrano et al., 2018).