Volatile apocarotenoid biosynthesis and carotenoid catabolism in Arabidopsis thaliana
Date
2017
Authors
Rivers, John Yates
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Abstract
Carotenoids are essential for plant photosynthesis. But, the
story doesn’t end there: volatile apocarotenoids, carotenoid
oxidative cleavage products, are important plant fragrances and
semiochemicals, and some are believed to be so-called
‘apocarotenoid signals’ (ACS), regulating plant processes.
From in vitro experiments, apocarotenoids are known to be formed
either by carotenoid cleavage dioxygenases (CCDs) or
non-enzymatic oxidation. But, in planta mechanisms are in many
cases unclear. Are there volatile apocarotenoids yet to be
discovered? Are volatile apocarotenoids formed via CCD or
non-enzymatic processes in planta? And are some volatile
apocarotenoids ACSs?
To study volatile apocarotenoid metabolism in planta, a solid
phase microextraction (SPME) gas chromatography mass spectrometry
(GC/MS) protocol for analysing volatile apocarotenoids was
developed. I also detoured to examine a new MS ionisation
technology, cold electron ionisation (cold-EI). This technology
improves molecular mass determination via GC/MS. The ability of
cold-EI to enhance GC/MS plant volatile identification (via
provision of molecular mass data) was evaluated.
SPME-GC/MS was then applied to analyse volatile apocarotenoid
profiles from the model plant Arabidopsis thaliana. Col-0
(wild-type) and ccd-loss-of-function single and double mutants
were analysed, as were the Arabidopsis ccr2 mutant and
corresponding ccr2-background ccd mutants. ccr2 accumulates
linear carotenoids not typically observed in Arabidopsis, and
I’ve found it also emits exotic volatile apocarotenoids. A
more-targeted study of volatile apocarotenoids and CCD1 and CCD4
activity during senescence was also performed.
This thesis discusses aspects of analytical method development,
specifically optimisation and applications of SPME sampling and
cold-EI technology. Using untargeted SPME-GC/MS volatile
metabolomics, a remarkable breadth of known and putative volatile
apocarotenoids was uncovered in Arabidopsis. Evidence suggests
CCDs barely contribute to volatile apocarotenoid production, most
likely producing non-volatile apocarotenoids instead. Volatile
apocarotenoids seem to be formed mainly via carotenoid
non-enzymatic oxidation. My results suggest carotenoid turnover
in photosynthetic tissues is important: volatile apocarotenoid
production often increases markedly when CCD function is lost.
Despite previous studies’ results, no new evidences were found
for CCD-derived ACS contributing to either the ccr2 phenotype or
senescence retardation.
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Keywords
Arabidopsis thaliana, apocarotenoids, carotenoids, apocarotenoid signals, ACS, volatiles, carotenoid cleavage dioxygenase, CCD, plant metabolism, solid-phase microextraction, SPME, gas-chromatography mass spectrometry, GC/MS, electron ionisation, EI
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Thesis (PhD)
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