2. ENVIRONMENTAL ChEMISTRy & TEChNOLOGy 2.1. Lectures

Chem. Listy, 102, s265–s1311 (2008) Environmental Chemistry & Technology
P54 ChANGES IN CAROTENOIDS PATTERN IN
MOugeOTiA sP. ALGAE INDuCED by hIGh
LIGhT STRESS
eDWARD MUnTEAn, VICTOR BERCEA, nICOLETA
MUnTEAn and nICOLAE DRAGOş
University of Agricultural Sciences and Veterinary Medicine
Cluj-Napoca, 3–5 Calea Mănăştur, 400372 Cluj-Napoca,
Romania,
edimuntean@yahoo.com
Introduction
When exposure to light exceeds a maximum that can be
used productively by the photosynthesys, a violaxanthin deepoxidation
leads to antheraxanthin and finally to zeaxanthin,
the excessive energy being then dissipated as heat 1 . At lower
irradiance, zeaxanthin is re-epoxidated back to violaxanthin
by zeaxanthin-epoxidase (Fig. 1.).
Fig. 1. The xanthophyll cycle
This reversible interconversion of zeaxanthin and violaxanthin
via antheraxanthin was called xanthophyll cycle or
violaxanthin cycle, being initially studied in higher plants 6,7 ;
further researches established that it has a photoprotective
role, removing the excess excitation energy from the photosynthetic
antennae 1–4 , protecting in this way photosynthetic
organisms from dammage by excessive light. The aim of this
research was to establish the way in which the carotenoid
biosynthesis is influenced by high light stress in the green
algae Mougeotia sp. Agardt.
Experimental
The carotenoid standards were kindly provided by F.
Hoffmann – La Roche, Basel, Switzerland. All solvents were
HPLC grade purity (ROMIL Chemicals). The green algae
Mougeotia sp. Agardt (AICB 560) originated from the collection
of the Institute of Biological Researches Cluj-napoca;
it was grown in a Bold nutritive solution mixed by introducing
air containing 5 % CO 2 , under continuous illumination
(300 µmol m –2 s –1 , measured with a Hansatech Quantum Sensor
QSPAR), at an average temperature of 20 °C for 15 days.
Extraction and high performance liquid chromatography analysis
(HPLC) were conducted according to a previous published
procedure 5 . Separations were performed on an Agilent
1100 system, using a nucleosil 120-5 C 18 column and the
s439
following mobile phases: A – acetonitrile : water (9 : 1) and
B – ethyl acetate. The flow rate was 1 ml min –1 . and the solvent
gradient was as follows: from 0 to 20 min. – 10 % to 70 % B,
then from 20 to 30 min. – 70 % to 10 % B.Carotenoids identification
was completed based on HPLC co-chromatography
with authentic carotenoid standards.
Results
The HPLC chromatogram from Fig. 2.a reveals the
carotenoid pattern for the saponified extract of Mougeotia sp.
control sample, dominated by two major carotenoids: lutein
and β-carotene. Besides, four xanthophylls (violaxanthin,
lutein, zeaxanthin and 5,6-epoxy-β-carotene) and four carotenes
(α-carotene, β-carotene, 9Z-β-carotene and 15Z-β-carotene)
were also identified.
When the Mougeotia culture was exposed to a high light
irradiation (4,500 µmol m –2 s –1 ), the content of antheraxanthin
increased strongly as a result of de-epoxidation (Fig. 2.b,
Table I), the carotenoid pattern being dominated by lutein and
antheraxanthin, while among minor carotenoids 5,6-epoxyβ-carotene
moved out and zeaxanthin appeared.
Table I
The carotenoid concentrations of target carotenoids [μg ml –1
algal suspension]
Control Irradiation Recovery after
Carotenoids sample with 4,500 irradiation
[μmol m –2 s –1 ]
Violaxanthin 0.02 0.01 0.10
Antheraxanthin 0.10 0.60 0.05
Lutein 1.00 0.56 0.37
Zeaxanthin 0.00 0.05 0.02
5,6-epoxy-β-carotene 0.04 0.00 0.03
α-carotene 0.04 0.01 0.02
β-carotene 0.19 0.03 0.10
9Z – β-carotene 0.04 0.01 0.02
15Z – β-carotene 0.01 traces 0.01
The whole carotenoid pattern was affected by the light
stress (Fig. 2.a and 2.b), not only the xanthophylls involved
in the xanthophyll cycle. Chromatograms emphasize another
important aspect in the studied matrix: the xanthophyll cycle
converts violaxanthin mainly in antheraxanthin, not in zeaxanthin;
this finding agrees with results reported for Mantoniella
squamata 3 , where they were attributed as consequences for
the mechanism of enhanced non-photochemical energy dissipation.
The recovery after the light stress leads to a reversible
epoxidation to violaxanthin, revealed by the chromatogram
from Fig. 2.c, the final higher violaxanthin level being correlated
with a strong decrease in antheraxanthin concentration
(Fig. 2.c, Table I), while the new chromatographic pattern is
dominated by four major carotenoids: lutein, β-carotene, violaxanthin
and 5,6-epoxy-β-carotene.