PESTICIDE
Biochemistry & Physiology
Pesticide Biochemistry and Physiology 78 (2004) 161–170
www.elsevier.com/locate/ypest
Effect of herbicide clomazone on photosynthetic processes
in primary barley (Hordeum vulgare L.) leaves
R. Ka
na,a,b,* M. Spundov
a,a P. Ilık,a D. Laz
ar,a K. Klem,c
P. Tomek,a J. Naus,a and O. Pr
asilb
b
a
Laboratory of Biophysics, Faculty of Science, Palacky University, tř. Svobody 26, Olomouc 771 46, Czech Republic
Photosynthesis Research Center at the Institute of Microbiology, Opatovicky Mly n, 379 81 Třeborň and Institute of Physical Biology,
University of South Bohemia, Nove Hrady 373 33, Czech Republic
c
Agricultural Research Institute Ltd., Havlıckova 2787, Kromerız 767 01, Czech Republic
Received 7 August 2003; accepted 17 December 2003
Abstract
The effect of pre-emergently applied herbicide clomazone on the photosynthetic apparatus of primary barley leaves
(Hordeum vulgare L.) was studied. Clomazone application caused a reduction in chlorophyll (a þ b) and carotenoid
levels that was accompanied by a decline in the content of light harvesting complexes as judged from the increasing
chlorophyll a/b ratio. The pigment reduction also resulted in changes in 77 K chlorophyll fluorescence emission spectra
indicating lower chlorophyll (Chl) fluorescence reabsorption and absence of the long-wavelength emission forms of
photosystem I. The maximal photochemical yield of photosystem II (PSII) and the reoxidation kinetics of the primary
quinone acceptor Q
A were not significantly influenced by clomazone. A higher initial slope of Chl fluorescence rise in
the Chl fluorescence induction kinetic indicated an increased delivery of excitations to PSII. Simultaneously, analysis of
the Chl fluorescence quenching revealed that clomazone reduced function of the electron transport chain behind PSII.
The decrease in the saturation rates of CO2 assimilation paralleled the decrease of the Chl content and has been
suggested to be caused by a suppressed number of the electron transport chains in the thylakoid membranes or by their
decreased functionality. The obtained results are discussed in view of physiological similarity of the clomazone effect
with changes of photosynthetic apparatus during photoadaptation.
Ó 2004 Elsevier Inc. All rights reserved.
Keywords: Clomazone; Herbicide; Photosynthesis; Pigments; Carotenoid; Chlorophyll; Photosynthetic rate; Chlorophyll fluorescence;
Fluorescence quenching; Photoadaptation
1. Introduction
Clomazone [2-(2-chlorobenzyl)-4,4-dimethyl1,2-oxazolidin-3-one] is a pre-emergence herbicide
*
Corresponding author. Fax: +420-58-522-57-37.
E-mail address: [email protected] (R. Ka
na).
used against broad-leafed and grassy weeds [1,2].
It is widely used for the weed control in canopies
of soybeans, cotton, sugar cane, corn, rice, tobacco, and various vegetable crops [3]. It is generally accepted that clomazone prevents the
accumulation of chloroplast pigments and plastidic isoprene evolution [4–6]. In the first papers
0048-3575/$ - see front matter Ó 2004 Elsevier Inc. All rights reserved.
doi:10.1016/j.pestbp.2003.12.002
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R. Kana et al. / Pesticide Biochemistry and Physiology 78 (2004) 161–170
dealing with clomazone action [4,5,7] all findings
were explained mostly as a consequence of clomazone interference with carotenoid biosynthesis
that leads to a bleaching (photooxidation) of
chlorophylls (Chls). Later paper indicated that the
‘‘bleaching effect’’ is not caused by the inhibition
of carotenoid biosynthesis and subsequent
bleaching of Chls [8]. In contrast, after clomazone
treatment an enzyme involved in the synthesis of
plastidic isopenthenyl diphosphate (IPP—the biological precursor of all isoprenoids) is inhibited [9]
and clomazone reduces synthesis and accumulation of both carotenoids and Chls and other
compounds containing isoprenoid parts. Recently
it has been shown that clomazone toxicity is not
induced by clomazone itself but rather by its
breakdown product, 5-ketoclomazone that blocks
1-deoxy-D -xylulose 5-phosphate (DOXP)1 synthase, the first enzyme of the plastidic isopentenyl
diphosphate (IPP) synthesis pathway [9–11].
Most of experiments dealing with the effect of
clomazone on photosynthesis were done with
greening plants. Clomazone slows down the chloroplast development during greening [4,7,12].
These experiments showed that clomazone treatment greatly decreased the chlorophyllide a to Chl
a transformation monitored by the Shibata shift
[4,7], the development of photosynthetic oxygen
evolution capacity [12] and electron transport in
thylakoid membranes of etio(chloro)plasts [4]. The
clomazone-affected greening leaves are very sensitive to relatively low intensities of light. The light
intensity of 150 lmol photons of PAR m2 s1 used
for the greening of etiolated seedlings of pitted
morning glory caused changes explained as
photobleaching as was judged from the ultrastructural changes in developing etio(chloro)plasts
[4]. Duke and Kenyon [4] further found that clomazone does not directly influence the electron
transport in thylakoid membranes isolated from
fully green tissues.
In this work we examined changes in the photosynthetic apparatus of barley seedling using Chl
fluorometric and gasometric methods with the aim
to characterize the adverse effects of clomazone on
the function of thylakoid membranes under assumption of minimal photobleaching effect.
2. Materials and methods
2.1. Plant material
Barley seedlings (Hordeum vulgare L. cv. Akcent) were cultivated in Petri dishes on filter paper
soaked with distilled water or with 0.25 and
0.5 mM clomazone solution for 12 days (1.1
growth phase according to [13]). The seedlings
were grown in continuous light (10 lmol photons
of PAR m2 s1 ) at 10 °C. These conditions were
chosen in order to minimize the pigment photobleaching. Primary leaves were used for all measurements.
2.2. Pigment content
1
Abbreviations used: PSI (II), photosystem I (II); qP ,
coefficient of photochemical chlorophyll fluorescence quenching; qN , coefficient of non-photochemical chlorophyll fluorescence quenching; QA , primary stable quinone acceptor of
photosystem II; Ag , gross photosynthetic rate; LHC I (II), light
harvesting complex of photosystem I (II); IPP, isopentenyl
diphosphate; Chl, chlorophyll; Chl a (b), chlorophyll a (b); Rfd ,
fluorescence decrease ratio (index of vitality); F0 , minimal
chlorophyll fluorescence; FM , maximal chlorophyll fluorescence; FV , variable chlorophyll fluorescence; FV /FM , maximal
quantum yield of photosystem II photochemistry; PAR, photosynthetic active radiation; DOXP, 1-deoxy-D -xylulose 5phosphate; MEP, 2-C-methyl-D -erythritol 4-phosphate; Pmax ,
maximal photosynthetic rate; TR0 /RC, parameter of initial Chl
fluorescence rise to the J step of the fast Chl fluorescence
induction; Chl a/b, chlorophyll a to chlorophyll b ratio.
The Chl and carotenoid content was determined
spectrophotometrically in 80% acetone with a
double beam spectrophotometer Unicam UV 550
(ThermoSpectronic, Cambridge, UK) according to
[14]. The pigment content was related to the leaf
area.
2.3. Low-temperature Chl fluorescence spectroscopy
Chl fluorescence emission spectra were measured
with leaf segments immersed in liquid nitrogen
(77 K) using the fluorescence spectrophotometer
F-4500 (Hitachi, Tokyo, Japan). Chl fluorescence
was excited at 436 nm (5 nm spectral slit-width) and
R. Kana et al. / Pesticide Biochemistry and Physiology 78 (2004) 161–170
detected in the spectral range of 665–795 nm (2.5 nm
spectral slit-width) from the adaxial side of leaf
segments.
2.4. Fast chlorophyll fluorescence rise and Q
A
reoxidation
The fast Chl fluorescence rise from the adaxial
side of barley leaves was measured using a shutterless portable Chl fluorometer PEA (Hansatech,
Norfolk, UK). The leaves were dark-adapted for
15 min. The maximal and minimal Chl fluorescence
(FM and F0 ) and the normalized slope at the initial
part of the Chl fluorescence rise TR0 /RC were calculated from the fast Chl fluorescence rise. TR0 =
RC ð¼ ðdV =dt0 Þ=VJ Þ is defined as ½4ðF300 ls F50 ls Þ=½ðFM F50 ls ÞVJ , and VJ ¼ ðF2 ms F50 ls Þ=
ðFM F50 ls Þ is the relative height of the step J in the
Chl fluorescence rise, where F300 ls (F2 ms , F50 ls ) is the
Chl fluorescence intensity at 300 ls (2 ms, 50 ls).
The relative height of this step reflects a transient
accumulation of reduced primary stable quinone
acceptor of photosystem II (Q
A ) (for details see
[15,16]). The reoxidation rate of Q
A , the reduced
primary quinone acceptor of photosystem II (PSII)
was measured with the dual-modulated fluorimeter
FL 100 (Photon Systems Instruments, Brno, Czech
Republic). Dark-adapted leaf segments were
irradiated with saturation red light flash (650 nm)
and the subsequent decline of Chl fluorescence
intensity, reflecting the Q
A reoxidation kinetics
was detected for 10 s by a weak measuring light
(see e.g. [16]).
2.5. Chlorophyll fluorescence parameters
quenching analysis
and
The whole barley leaves of each leaf type (1
control and 2 clomazone-treated types) were darkadapted for 15 min. Then the blade of each leaf was
cut off and about 2.5-cm long central segment was
used for the measurements. Two segments of each
leaf type were used for one measurement and five
repetitions were performed. Chl fluorescence inductions were measured by the kinetic imaging
fluorometer FluorCam 700 MF (Photon Systems
Instruments, Brno, Czech Republic) using continuous actinic red light (650 nm; 300 lmol m2 s1 ).
163
Chl fluorescence was detected during short red light
(10 ls; 650 nm) measuring flashes. Strong white
light pulses (1000 lmol photons of PAR m2 s1 ,
1 s duration) were used to saturate electron transport in thylakoid membranes. The minimal Chl
fluorescence of dark-adapted segments (F0 ) was
determined as an average value of the Chl fluorescence signal detected by 3 measuring flashes placed
1 s apart before the onset of actinic light. Then, the
maximal Chl fluorescence of the dark-adapted segments (FM ) was determined as the Chl fluorescence
signal detected by the measuring flashes in the
middle of the saturating pulse period. The maximal
Chl fluorescence during the slow Chl fluorescence
induction (i.e., for the light-adapted segments) (FM0 )
was determined every 20 s during the actinic light
exposure of the leaves in the same way as FM . The
Chl fluorescence signal detected by the measuring
flashes during the Chl fluorescence induction just
before application of the saturating pulse to determine FM0 was taken as F (see below). The value of F
detected by the measuring flashes at the sixth minute
of exposure of the segments to the actinic light was
considered as the steady-state Chl fluorescence level
for the light adapted segments (FT ). The photochemical (qP ) and the non-photochemical (qN ) Chl
fluorescence quenching coefficients were calculated
as ðFM0 F Þ=ðFM0 F0 Þ and ð1 ðFM0 F0 ÞÞ=ðFM F0 Þ, respectively [17]. The maximal quantum yield of
photosystem II photochemistry was calculated as
FV =FM ¼ ðFM F0 Þ=FM [18] and the vitality index as
Rfd ¼ ðFM =FT Þ 1 [19]. For further details on the
Chl fluorescence parameters see the latest review on
this topic by [20].
The Chl fluorescence parameters were evaluated
for each whole segment area independently (one
segment consisted from about 400 pixels). The
average value of the calculated Chl fluorescence
parameter for a given leaf type was used for the
presentation of the results.
2.6. Gas exchange
The leaf CO2 exchange was measured by an
open gasometric system (LCA—4, ADC, Hoddeson, UK). The attached leaves were equilibrated in
the leaf chamber under standard conditions
(CO2 concentration in the air 350 lmol mol1 ,
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R. Kana et al. / Pesticide Biochemistry and Physiology 78 (2004) 161–170
temperature 24 0.3 °C and water pressure deficit
1 0.2 kPa).
The steady-state net photosynthetic rate (An ) was
calculated using the equations of Von Caemmerer
and Farquhar [21] for leaves adapted for 25 min at
given light intensity (27, 65, 131, 318, 435, 870,
1600 lmol photons of PAR m2 s1 ). The gross
photosynthetic rate (Ag ) was evaluated as a sum of
dark respiration (measured for dark-adapted
leaves) and net photosynthetic rate. The measured
light response curves were fitted by non-linear
convexity equation [22] to obtain the photosynthetic capacity (Pmax ) and the parameter of convexity (h).
3. Results
3.1. Photosynthetic pigments and pigment–protein
complexes
The levels of Chls (Chl a, Chl b) and carotenoids
in the leaves markedly decreased with increasing
clomazone concentration (Fig. 1). In correspondence with the previous results [6,8,23] the decrease
of Chls was more pronounced than the decrease of
carotenoids (Fig. 1B, inset). The observed clomazone-evoked increase in the Chl a/b ratio in barley
leaves (Fig. 1A, inset) already shown in [6] reflects a
preferential reduction of the Chl b-containing
light-harvesting complexes (LHCs) in thylakoid
membranes [24,25]. The changes at the level of
pigment–protein complexes in clomazone-treated
leaves were also observed in the 77 K Chl fluorescence emission spectra (Fig. 2). The clomazone
addition resulted in: (1) the shift of the long-wavelength emission band (F735) which is attributed to
the light harvesting complexes of PSI (LHCI) from
739 (control leaves) to 733 nm (500 lM clomazone)
and (2) in the increase of the short-wavelength
region (F685 and F695 bands, attributed to PSII
emission) relatively to that of the F735 band (Fig. 2).
The relative increase of the F685 and F695 bands
can be ascribed to the reduction of the Chl fluorescence reabsorption corresponding to the Chl decline
[26–28] and the shift of the F735 band to lower
wavelengths could reflect the absence of highly
organized LHCI complexes [29].
Fig. 1. Pigment content per leaf area in control and clomazonetreated leaves. (A) Chl level; inset in (A) Chl a/b ratio; (B)
total carotenoid concentration; inset in (B) Chl/total carotenoid
ratio. All data represent means and SD for n ¼ 7.
Fig. 2. Chlorophyll fluorescence emission spectra of control
and clomazone-treated barley leaves measured at 77 K. The
position of long-wavelength fluorescence maximum (F735)
shifted to lower wavelengths (739 nm, control sample; 734 nm,
250 lM clomazone; and 732 nm, 500 lM clomazone) and its
intensity in maximum relative to that in the maximum of F685
band was lowered (F685/F735 ¼ 0.47 for control sample; 0.89
for 250 lM; 1.03 for 500 lM) with higher clomazone concentration. Spectra were normalized to the maximum of F735
band. Excitation wavelength: 436 nm; emission and excitation
spectral slit-widths: 2.5 and 5 nm, respectively.
R. Kana et al. / Pesticide Biochemistry and Physiology 78 (2004) 161–170
3.2. Primary photosynthetic processes
The above-described results revealed that clomazone treatment reduces the photon absorption
capability of the photosynthetic apparatus. In order to characterize how the reduction of pigment
amount can influence the electron transport in
thylakoid membranes in vivo we measured several
Chl a fluorescence parameters.
Only minor changes observed in the Chl fluorescence ratio FV /FM (Fig. 3) imply that the maximal quantum yield of PSII photochemistry [18]
was not significantly influenced by the clomazone
treatment. Further, no changes in the kinetics of
reoxidation of the primary stable electron acceptor
(Q
A ) on the acceptor side of PSII (data not shown)
and the independence of the VJ value on the clomazone concentration (see Table 1) indicate that
the function of PSII and electron transport close
behind PSII were not significantly affected by the
clomazone treatment. This is in accordance with
Fig. 3. Maximal quantum yield of photosystem II photochemistry (FV /FM ) and ‘‘vitality index’’ of photosynthetic apparatus—Rfd (Chl fluorescence decrease ratio) for control and
clomazone-treated barley leaves. Means and SD are shown,
n ¼ 10.
165
the results of Duke and Kenyon [4] who observed
no direct effect of clomazone on the PSII electron
transport measured by Hill reaction (H2 O !
FeCN) in isolated cowpea thylakoids. Our results
also indicate that the Chl and other pigment
molecules required for the proper photochemical
function of PSII reaction centers were probably
supplied to the assembled PSII complexes in
sufficient extent regardless to their lower content
induced by the clomazone treatment (Fig. 1).
Similarly, the short-wavelength part of the lowtemperature Chl a fluorescence emission spectrum
contains both the F685 and F695 bands in the
usual ratio (Fig. 2) and thus the inner antennae of
PSII complex seem not to be affected by the clomazone tretment.
Another PSII characteristic, the kinetics of the
QA reduction, was determined by measurements of
the slope of variable Chl a fluorescence rise within
the 50–300 ls interval after the onset of the intensive excitation light. When the Q
A reoxidation
proceeds normally, as was observed for all our
samples (see above), the slope of this Chl fluorescence rise, normalized to the Chl fluorescence intensity at the J step in the Chl fluorescence
transient (TR0 /RC parameter, see also Section 2),
is proportional to the flux of excitations trapped
per reaction center of PSII in the sample [16]. We
expected this parameter to be lower for the clomazone-treated leaves due to their reduced content
of LHCs that can supply excitations to the
photosystems (see the results above). However, we
observed quite opposite effect (Fig. 4). The TR0 /
RC parameter was by about 30% higher for the
plants grown in the presence of 500 lM clomazone
than was in the control barley plants. The supply
of excitations to PSII centers was accelerated
probably due to the effect of increased light field
Table 1
Effect of clomazone treatment on Chl fluorescence intensity (F0 , minimal fluorescence; FM , maximal fluorescence; and FV , variable
fluorescence) and on the relative Chl fluorescence intensity in the J step of fluorescence rise
Clomazone
concentration (lM)
F0 (r.u.)
FM (r.u.)
FV (r.u.)
VJ
0
250
500
553 16
638 148
1000 286
2964 201
2950 403
2843 212
2411 201
2312 429
1843 356
0.557 0.046
0.524 0.025
0.557 0.056
Measured with PEA fluorometer (n ¼ 10).
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R. Kana et al. / Pesticide Biochemistry and Physiology 78 (2004) 161–170
Fig. 4. The TR0 /RC parameter that characterizes a slope of the
initial Chl fluorescence rise to the J step in the fast Chl fluorescence induction for control and clomazone-treated barley
leaves. This parameter is proportional to the flux of excitations
trapped per reaction center of photosystem II (for details see
Section 2). Means and SD are shown, n ¼ 10.
inside the leaves. The effect of ‘‘optical trap’’
leading to a relative enhancement of light intensity
inside the leaf has been already demonstrated for
leaves with reduced pigment content during
greening [30] and it has been associated with high
scattering and reflections on many surfaces inside
the leaf [31].
More detailed information about changes in the
utilization of absorbed light provide measurements
of the Chl fluorescence quenching coefficients
during irradiation of the dark-adapted leaves [17].
These coefficients reflect the extent to which the
utilization of absorbed light proceeds by photochemical (qP ) or non-photochemical (qN ) ways (for
details see e.g. [20]). The (1 qP ) value is approximately related to the actual fraction of PSII
with reduced QA ([20] and references therein). The
(1 qP ) coefficient was higher for clomazonetreated leaves in comparison to the control leaves
(Fig. 5A). Such increase usually implies some reduction of the electron transport processes beyond
QA [32]. It means that although electron transport
through PSII centers seems to be unaffected by
clomazone, the rest of the photosynthetic electron
transport chain is not able to efficiently withdraw
electrons from PSIIs. The observed increase in
(1 qP ) can be also caused by the above-mentioned increase in the excitation flux trapped
by the individual PSII reaction centers due to the
Fig. 5. Courses of Chl fluorescence quenching parameters
1 qP (A) and qN (B) during the first 6 min of actinic irradiation (300 lmol photons of PAR m2 s1 ) for control and clomazone-treated barley leaves. Leaves were kept in the dark for
15 min before the measurement. Means and SD are shown,
n ¼ 10.
increase in the internal light field inside leaves
treated with clomazone.
The parameter (1 qP ) is also known as the
excitation pressure [33]. For the non-stressed mature leaves, the enhancement of the PSII excitation
pressure by the increasing intensity of actinic light
leads to the increase of the non-photochemical
quenching coefficient qN [34,35]. In our case, on
the contrary, the higher PSII excitation pressure
found for clomazone-treated leaves was accompanied by a decrease in qN (Fig. 5B). This might
indicate inability to maintain balance between excitations used by PSII for photochemical events
and excitations dissipated in non-photochemical
way (coefficient qN ) because for the non-stressed
leaves the ratio between (1 qP ) and non-photochemical quenching coefficient is roughly constant
[36]. Recently, it has been observed that reduction
in specific LHCs strongly inhibits the non-photochemical Chl fluorescence quenching [37]. Thus
the observed decrease in qN might be caused by
the reduction of LHCs (see Fig. 1A, inset). The
R. Kana et al. / Pesticide Biochemistry and Physiology 78 (2004) 161–170
clomazone treated leaves with suppressed content
of carotenoids (Fig. 1B) and LHCs probably also
have reduced content of protective pigments of the
xanthophyll cycle. Both the incomplete composition of the LHC systems and the shortage of
xanthophyll molecules might be responsible for the
effect of reduced capability of non-photochemical
quenching.
3.3. The rate of CO2 assimilation
The decrease of another Chl a fluorescence parameter—Rfd (Fig. 3) is regarded as a consequence
of the decrease in both qP and qN parameters and is
interpreted as decrease in the potential photosyn-
167
thetic activity [19]. Since changes in this parameter
usually reflect changes in the rate of CO2 fixation
in leaves [38] we have decided to measure the impact of clomazone on the rate of CO2 fixation
(gross photosynthetic rate Ag ) at eight different
irradiancies (the so-called light–response curve of
photosynthesis). As can be seen in the Fig. 6A, the
clomazone action decreased the photosynthetic
rate for all used irradiancies, the result similar to
previous results [39].
We have fitted the experimental results by the
non-linear convexity equation to obtain the parameters of the photosynthesis versus irradiance
curves. For higher clomazone concentrations we
observed a decrease in convexity (h) (see legend to
Fig. 6). This result can be explained as a consequence of the reduced Chl content because the h
parameter decreases with lower leaf absorptance
[40], and also due to the reduced supply of
NADPH and ATP that limit the regeneration of
ribulose 1,5-bisphosphate [22,41]. If we present the
photosynthetic rate on Chl basis (Fig. 6B) there
are no significant differences for the control and
clomazone treated leaves. This indicates that the
photosynthetic rate and especially Pmax were reduced as a consequence of the changes in pigment
content.
4. Discussion
Fig. 6. Representative light–response curves of gross photosynthetic rate (Ag ) for control and clomazone-treated barley
leaves that characterize photosynthetic rate calculated on leaf
area (A) and chlorophyll content (B). The curves plotted in (A)
have been obtained by fitting experimental data with the
equation of convexity (for details see Section 2). The maximal
photosynthetic rate (Pmax ) was 12.64 (control), 7.04 (250 lM
clomazone), and 3.77 (500 lM clomazone) lmol (CO2 ) m2 s1 .
Parameter of convexity decreased with higher clomazone concentration (0.78 for control; 0.67 for 250 lM clomazone; 0.57
for 500 lM clomazone). The intensity of irradiance is characterized in lmol of photons of photosynthetic active radiation
(400–700 nm).
Our work confirmed the well-known fact (see
Section 1) that the main effect of clomazone action
on photosynthesis is a reduction in the content of
photosynthetic pigments, Chls, and carotenoids
(Fig. 1). A similar decrease in the pigment level, in
the inverse sense, can be also measured during
greening of etiolated leaves on continuous light
[42–44]. The 77 K Chl fluorescence emission spectra (Fig. 2) obtained for clomazone-treated leaves
are also similar to those observed for the leaves at
the early stages of greening [43,45–47]. This means
that the deficiency of absorbed light in the case of
etiolated barley plants retards Chl synthesis similarly like clomazone treatment inhibits pigment
synthesis. Consequently, both phenomena lower
the formation of pigment–protein complexes in
thylakoid membranes of chloroplasts.
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R. Kana et al. / Pesticide Biochemistry and Physiology 78 (2004) 161–170
Such reduction in the photosynthetic pigment
content has influence on photosynthesis at different levels that can be measured by several methods
like changes in the photosynthetic assimilation rate
or by the Chl fluorescence (Chl fluorescence
emission spectra, parameters of variable Chl fluorescence, and quenching of Chl fluorescence). Interdependence between changes in the photon
absorption capability and photosynthetic parameters has been also observed for leaves grown at
different irradiancies (see e.g. [48–51]). If we compare changes in the photosynthetic parameters
measured with clomazone treated leaves with
changes observed during the photoadaptation (for
the detailed study see e.g. [52–54]) we have found
that there is no clear similarity with the ‘‘sun’’ or
with the ‘‘shade’’ photoadaptation types of the
photosynthetic apparatus. In case of clomazone
treatment, the parameters that characterize the
pigment composition of leaves (Chl content,
Fig. 1A; Chl a/b, Fig. 1A, inset; and Chls/carotenoids, Fig. 1B, inset) revealed a similarity to the
‘‘sun’’ type adaptation response but on the other
hand Chl fluorescence parameters (F0 and FV ,
Table 1; FV /FM , Fig. 3; and Rfd , Fig. 3) and photosynthetic rate (Fig. 6) rather indicate ‘‘shade’’
type adaptation. Similar results were also observed
for transgenic tobacco leaves with reduced pigment content (see [55]). It can be deduced that the
regulatory mechanism of photoadaptation that
maintains balance between the content of photosynthetic pigments and optimal utilization of the
absorbed light energy for photochemistry cannot
operate to full extent in leaves with pigment content reduced by clomazone.
Acknowledgments
This project was supported by the Grant No.
522/00/1274 from the Grant Agency of the Czech
Republic and by the Ministry of Education of the
Czech Republic (projects with numbers MSM
was supported by
153100010 and 12300001). M.S.
the Grant No. 522/01/P098 from the Grant
Agency of the Czech Republic. The research in the
laboratory of R.K. and O.P. was supported by the
Ministry of Education of the Czech Republic
(project LN00A141) and by the Institutional Research Concept No. AV0Z5020903.
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