Hardorp 2024 Bachelor-Thesis
| Hardorp RA (2024) Oxygen consumption and production measured by high-resolution respirometry in leaf preparations from Robinia pseudoacacia. Bachelor-Thesis 47 pp. |
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Hardorp Rebecka A (2024) Bachelor-Thesis
Abstract: This study presents a novel approach to measure oxygen consumption and production under controlled light conditions in preparations of leaf samples using the Oroboros O2k highresolution respirometer. Dark respiration, photosynthesis-irradiance (PI) curves, and photodegradation described by oxygen consumption-irradiance (CI) curves were obtained accurately and reproducibly on leaf preparations. The developed protocols for high-resolution respirometry and photobiology provide a novel approach for understanding plant respiratory and photosynthetic processes. Green and yellow senescent leaves were collected from a Robinia pseudoacacia tree in the fall. The sample preparation involved the cutting and homogenization of leaves in two different media. Fresh and frozen samples of leaf homogenate were compared. First experimental test series are reported using the O2k equipped with the PhotoBiology (PB) Module. The instrument was controlled by, and data were recorded with DatLab 8, and the obtained data were further analyzed with Excel and GraphPad.
Dark respiration and photosynthetic activity of cut leaf pieces were plotted as a function of lightintensity to obtain photosynthesis-irradiance (PI) curves in the range of light intensities of 10 to 2000 ΞΌmolΒ·s-1Β·m-2. Measurements of oxygen consumption in the dark and at different light intensities revealed in the leaf homogenates an unexpected increase of oxygen consumption with blue, red, and white light. Mirror images of PI curves were remarkably similar to the shape of oxygen consumption-irradiance (CI) curves.
Mitochondrial dark respiration and chloroplast photorespiration are superimposed by residual oxygen consumption (Rox), defined as the oxygen consumption due to oxidative side reactions remaining after inhibition of the mitochondrial and chloroplast electron transfer pathways to oxygen. Application of selective inhibitors of the mitochondrial electron transfer system provided evidence against the involvement of mitochondrial electron transfer in dark and lightinduced oxygen consumption of the leaf homogenates which, therefore, was characterized as photodegradation. The effect of inhibitors of the chloroplast electron transfer system on photodegradation remains to be investigated.
Further studies are required with leaves at different stages of their life cycle and with sample preparations which preserve the functional integrity of mitochondria and chloroplasts. The production of reactive oxygen species is probably involved in photodegradation, which is an ecologically relevant process in various stages of the lifecycle of leaves and becomes progressively or abruptly dominant towards the final phase of leaf decomposition. Integrating this perspective into studies of dark respiration and photosynthesis of different species is an entirely new application field of high-resolution respirometry. It has a great potential to advance our understanding of plant physiology and analysis of the carbon balance in forest ecosystems and agriculture.
β’ Bioblast editor: Gnaiger E
Labels: MiParea: Respiration
Pathology: Aging;senescence
Stress:Oxidative stress;RONS, Hypoxia
Organism: Plants
Preparation: Homogenate
Regulation: Oxygen kinetics
HRR: Oxygraph-2k, NextGen-O2k
PhotoBiology, Photodegradation
