close

Вход

Забыли?

вход по аккаунту

?

553

код для вставкиСкачать
J Sci Food Agric 1998, 78, 208È212
The Eþect of Light and Temperature on
Glucosinolate Concentration in the Leaves and
Roots of Cabbage Seedlings
Eduardo A S Rosa* and Paula M F Rodrigues
Section of Horticulture, Department of Field Crops, Universidade de Tras-os-Montes e Alto Douro,
Apartado 202, 5001 Vila Real Codex, Portugal
(Received 17 April 1997 ; revised version received 23 January 1998 ; accepted 8 February 1998)
Abstract : In previous studies it was shown that the concentration of total and
individual glucosinolates in brassicaceous plants can vary signiÐcantly over a
24-h period grown either in the Ðeld or under controlled conditions. The present
study shows total and individual glucosinolate variation during a single day.
Seedlings of cabbage grown under controlled conditions and at 14 and 15 days
after emergence were moved to 20¡C (Exp A) and 30¡C (Exp B), with a constant
photosynthetic photon Ñux density of 480 lmol m~2 s~1 and 75% relative
humidity, over a 2-day period, during which time aerial parts and roots were
sampled at regular intervals. Whilst the glucosinolate patterns of the aerial part
of the plant and of the roots remained the same, the levels of major glucosinolates in the aerial part, averaged over all sampling times and 2 days, were
233 ^ 60 lmol 100 g~1 DW for 3-methylsulphinylpropyl and 72 ^ 22 for 2propenyl ; in the roots, 2-phenylethyl and 1-methoxyindol-3-ylmethyl showed the
highest average concentrations, with 678 ^ 355 lmol 100 g~1 DW and
411 ^ 122, respectively. Total and individual glucosinolate levels showed very
high signiÐcant di†erences between the two plant parts. Despite the constant
temperature, light and relative humidity, glucosinolates varied within a 24-h
period, showing ultradian rhythms that are common to several metabolic processes in plants. The results conÐrm previous observations that at a temperature
of 20¡C, close to the optimum for growth and development, the diurnal variation
in glucosinolate concentration, was smaller than at 30¡C. ( 1998 Society of
Chemical Industry
J Sci Food Agric 78, 208È212 (1998)
Key words : glucosinolate ; temperature ; light ; relative humidity
several reviews (Rosa et al 1997 ; Fenwick et al 1983 ;
Duncan 1991), which also discuss the role of glucosinolates on insect behavior such as attraction, oviposition
and feeding deterrence, matters of major concern in an
environmental agriculture system. Glucosinolate concentrations have been shown to vary not only between
plant species but also between individual parts of the
same plant, throughout the growth period and between
growing seasons and years (Macfarlane Smith and GrifÐths 1988 ; McGregor 1988 ; Clossais-Besnard and
Larher 1991 ; Fieldsend and Milford 1994a,b ; Rosa et al
1996), suggesting that climate factors are involved in
such variation (Rosa et al 1994, 1996 ; Rosa 1997). Since
brassicaceous plants are grown all year around in many
parts of the world and under the most diverse climate
INTRODUCTION
Glucosinolates are naturally occurring compounds in
brassicaceous plants which, on hydrolysis by the
enzyme myrosinase (thioglucoside glucohydrolase ; EC
3.2.3.1), liberate, in addition to sulphate ion and glucose,
a range of compounds which may include isothiocyanates, nitriles and thiocyanates. These hydrolysis
products are associated with several biological e†ects
that may be either beneÐcial, such as the anticarcinogenic activity of isothiocyanates, or detrimental
(eg the goitrogenic properties of thiocyanates and
oxazolidine-thiones). Such e†ects have been described in
* To whom correspondence should be addressed.
208
( 1998 Society of Chemical Industry. J Sci Food Agric 0022È5142/98/$17.50.
Printed in Great Britain
E†ect of light and temperature on glucosinolate concentration
conditions, it is likely that plant products might have
di†erent glucosinolate concentrations according to the
climate in which they are produced and even the period
of the day in which they are harvested. SigniÐcant
changes in glucosinolate concentration between
growing seasons and during a 24 h period might be
reÑected in the biological properties of brassicaceous
plants.
In previous work it was shown that glucosinolates in
the leaves of young cabbage plants grown under Ðeld
conditions can vary within a single day, suggesting a
rapid metabolism of these compounds (Rosa et al 1994).
In a more recent study under a constant 14 h photoperiod (0700È2100) and two temperature regimes (20
and 30¡C), the authors demonstrated variation in total
and individual glucosinolate for both aerial parts and
roots of developing young cabbage plants (Rosa 1997).
The present work describes the e†ect of two constant
regimes (20 and 30¡C) on total and individual glucosinolate levels throughout a 24 h period under controlled
constant conditions of light at a photosynthetic photon
Ñux density of 480 lmol m~2 s~1 and relative humidity
at 75%.
MATERIALS AND METHODS
Plant material
The young plants of the pointed cabbage (Brassica
oleracea var capitata cv Duchy F1) were grown in polypropylene trays with alveoles of 65 cm3 Ðlled with a
mixture of 1 vol sand (0É02È0É2 mm) to 3 vol peat
compost (Humobact Terreau, Frans Baele, Bailleul,
France). Seeds were placed at a depth of 1É5 cm and
covered with vermiculite. Trays were placed in a growth
cabinet (Conviron E15) under climatic controlled conditions of a 14 h photoperiod (0700È2100), corresponding
to a photosynthetic photon Ñux density of
480 lmol m~2 s~1, 75% relative humidity and 25¡C
during the light cycle and 18¡C during the dark cycle.
At 12 days after emergence (DAE), light and temperature regimes were changed to a constant
480 lmol m~2 s~1 and 20¡C, respectively (Exp A).
After 48 h, sampling was started with samples collected
at 0200, 0600, 1000, 1400, 1800 and 2200 over a period
of two consecutive days (14 and 15 DAE). Another
study (Exp B) was conducted in the same way as Exp A,
except that temperature was kept at a constant 30¡C,
with light and relative humidity exactly the same as in
the previous experiment.
Chemical analysis
In both experiments, plants were divided into aerial
parts and roots and analysed separately. At harvest,
the plants which were at 3È4 true leaf stage, lifted from
209
the alveoles and the intact aerial part was cut at the soil
level and immediately frozen in liquid nitrogen and
freeze-dried. The roots were separated from the peat
compost by soaking in water and gently washed in a
continuous water Ñow to remove all peat residues with
minimal damage. After removing excess water with
absorbent paper, the roots were frozen in liquid nitrogen and freeze-dried. A total of six plants were used in
each of three replicates. Plants were randomly selected,
ensuring that at each sampling time each plant was surrounded by others to avoid border e†ects. The freezedried material was reduced to a Ðne powder and
samples (about 200 mg) were extracted by addition of
boiling 90% methanol (3 ml plus 0É4 lmol benzyl glucosinolate as an internal standard) and maintaining
boiling for 10 min, a procedure which simultaneously
inactivates myrosinase. After Ðltration, the residue was
re-extracted twice using boiling 70% methanol (3 ml).
The extracts were combined to give a Ðnal volume of
10 ml and an aliquot (3 ml) was evaporated to dryness
and taken up in water (3 ml), of which 2 ml was applied
to small columns of DEAE Sephadex A 25 and the
absorbed glucosinolate desulphated as described by
Heaney and Fenwick (1980). Desulphoglucosinolates
were eluted with water and analysed using the highperformance liquid chromatography (HPLC) procedure
described by Spinks et al (1984). Glucosinolate concentration was expressed in lmol 100 g~1 DW. Statistical
analysis was done using a SuperAnova 1.1 software.
RESULTS AND DISCUSSION
The glucosinolate patterns of the aerial part and the
roots of the cabbage plants were similar in both Exp A
and B, which is contrary to previous studies (Rosa
1997). The di†erence between the two studies may be
due to the use of di†erent cultivars of the same species,
which are likely to a†ect glucosinolate patterns (Rosa et
al 1997).
The major glucosinolates in the aerial part of the
cabbage seedlings (Figs 1 and 2), were 3-methylsulphinylpropyl, 2-propenyl and indol-3-ylmethyl, with
minor amounts of 2-hydroxybut-3-enyl, 1-methoxyindol-3-ylmethyl, 4-hydroxyindol-3-ylmethyl and 2phenylethyl glucosinolates. In the roots (Figs 1 and 2),
2-phenylethyl, 1-methoxyindol-3-ylmethyl, 2-propenyl
and indol-3-ylmethyl were the main glucosinolates, with
minor amounts of 3-methylsulphinylpropyl, 2hydroxybut-3-enyl and 4-hydroxyindol-3-ylmethyl glucosinolates. The major glucosinolates in both parts of
the plant were the same as in a previous study (Rosa
1997).
As previously reported (Rosa 1997), the mean standard errors for total and individual glucosinolates in the
roots were larger than for the aerial part of the plant,
probably due to inevitable damage during the cleaning
210
Fig 1. Individual and total glucosinolate variation in the
aerial part (K) and roots (L) throughout a 24 h period at a
constant temperature of 20¡C, 480 lmol m~2 s~1 and 75%
relative humidity (Exp A). When SE bars are not shown, the
SE is smaller than the symbols.
process of the roots leading to some glucosinolate degradation. Negligible mean standard errors were determined for the aerial part which required only minimal
cleaning prior to extraction.
Total glucosinolates
In both experiments (Figs 1 and 2), the levels of total
glucosinolates in the roots, averaged over all sampling
times, were signiÐcantly higher (P \ 0É001) than in the
leaves. Higher levels for glucosinolates in roots were
E A S Rosa, P M F Rodrigues
Fig 2. Individual and total glucosinolate variation in the
aerial part (K) and roots (L) throughout a 24 h period at a
constant temperature of 30¡C, 480 lmol m~2 s~1 and 75%
relative humidity (Exp B). When SE bars are not shown, the
SE is smaller than the symbols.
also reported in previous studies (Josefsson 1967 ; Rosa
1997). When comparing the same plant part for total
glucosinolate levels at 20 and 30¡C, no signiÐcant di†erences were noted in leaves (386 ^ 71 lmol 100 g~1 DW
in Exp A vs 409 ^ 104 in Exp B) whilst, in roots, total
glucosinolate levels at 30¡C (1635 ^ 684 lmol 100 g~1
DW) were signiÐcantly higher (P \ 0É01) than at 20¡C
(1342 ^ 269 lmol 100 g~1 DW). Although these results
are in agreement with previous studies conducted in the
Ðeld (Rosa et al 1996) in which summer conditions tend
to increase the glucosinolate levels (between 4 and 35%
E†ect of light and temperature on glucosinolate concentration
in the leaves of Ðve Brassica species) when compared to
winter seasons, they showed less magnitude of variation
probably because the di†erences in the temperature
regimes were lower than in the Ðeld experiment and
because all the other climatic factors were kept constant. In the present study, glucosinolate levels tend to
be higher at higher temperature, suggesting that temperature (the only variable) could be responsible for
such di†erences.
Consideration of total glucosinolate levels between
sampling times at 20¡C (Exp A) showed no signiÐcant
di†erences in the leaves and signiÐcant variation
(P \ 0É05) in the roots ; at 30¡C (Exp B) the variation of
total glucosinolate levels between sampling times in
both parts of the plant was larger (P \ 0É001), probably
as a result of temperature stress. These Ðndings are in
agreement with previous results (Rosa 1997), further
demonstrating that photoperiod is likely to have little
interference in glucosinolate variation, since such variation is independent of light/dark conditions. Figure 1
shows that, in the present study, total glucosinolate
concentration in the leaves in Exp A are fairly stable ; in
the roots, there was a sinusoidal-type trend with peaks
at 0600 and 2200 following the major glucosinolates 2phenylethyl and 1-methoxyindol-3-ylmethyl. In Exp B,
at a constant 30¡C (Fig 2), total glucosinolate concentration in the leaves showed an upward trend also with
a sinusoidal-type curve ; for concentration, in the roots,
this curve type was much clearer, with peaks at 0200
and 1400. This also reÑected the trend in the concentration of the major glucosinolate. This type of evolution
curve corresponds to that described in the ultradian
rhythms which have been reported to occur in plants
for several metabolic processes such as glycolysis, sap
Ñow, enzyme activity and protoplasmic streaming in
less than 20 h (Salisbury and Ross 1992). This occurs
even under constant illumination and steady conditions,
after plants have been tuned to natural conditions.
Thus, it is likely that glucosinolates and their precursors
will follow a similar trend, since many photoresponses
in plants become so enmeshed with the daily change in
environmental factors that they become self-sustaining.
Di†erences between the evolution curves at both temperature regimes might reÑect the sensitivity to temperature of most of the ultradian rhythms.
Individual glucosinolates
The concentration of the major individual glucosinolate, 3-methylsulphinylpropyl, was signiÐcantly higher
(P \ 0É001) in the aerial part (228 ^ 55 lmol 100 g~1
DW in Exp A and 238 ^ 67 lmol 100 g~1 DW in Exp
B) than in the roots (94 ^ 39 lmol 100 g~1 DW in Exp
A and 103 ^ 53 lmol 100 g~1 DW in Exp B) when
averaged over all sampling times. This tendency had
been noted in previous studies (Rosa 1997). When comparing 3-methylsulphinylpropyl glucosinolate levels
211
within the same plant part at the two temperature
regimes, no signiÐcant di†erences were noted. At a constant 20¡C and constant light and relative humidity
throughout the day (Fig 1), an inverse relationship was
noted between the aerial part and roots, with no signiÐcant variations between sampling times in both parts of
the plant. This Ðnding suggests, as indicated for total
glucosinolates, an ultradian rhythm of the 3methylsulphinylpropyl glucosinolate associated with a
turnover between roots and aerial part. When seedlings
were submitted to 30¡C (Exp B, constant light and relative humidity) (Fig 2), signiÐcant di†erences (P \ 0É001)
were noted between sampling times in both parts of the
plant, with levels in the leaves being characterised by an
upward trend. Thus, it is likely that, at a temperatures
close to stress levels, plants respond with an increase in
3-methylsulphinylpropyl glucosinolate, which is in
agreement with previous Ðndings (Rosa 1997). On the
other hand, when all the three climatic parameters
(temperature, permanent light and relative humidity)
were kept constant and under the optimum growing
temperatures, 3-methylsulphinylpropyl glucosinolate
levels were relatively steady. This is contrary to the
results observed when plants were submitted to the
same conditions except photoperiod (light cycle 0700È
2100) (Rosa 1997). Thus, light may interfere in the variation of this glucosinolate.
Despite its lower concentration, 2-propenyl glucosinolate showed a similar trend to 3-methylsulphinylpropyl
in both experiments. However, in this case, the levels in
the roots (113 ^ 43 lmol 100 g~1 DW in Exp A and
151 ^ 83 lmol 100 g~1 DW in Exp B) were signiÐcantly higher (P \ 0É001) than in the aerial part of the
plant (67 ^ 18 lmol 100 g~1 DW in Exp A and
78 ^ 25 lmol 100 g~1 DW in Exp B). The levels of 2propenyl glucosinolate in both aerial parts and roots
varied signiÐcantly (P \ 0É001) when temperature was
changed from 20 to 30¡C. At a constant 20¡C and constant light and relative humidity, no signiÐcant variations were noted throughout the day in either part of
the plant, but, when temperatures was changed to 30¡C,
the variations were signiÐcant (P \ 0É001), as observed
in total and 3-methylsulphinylpropyl glucosinolates.
These observations and previous results (Rosa 1997)
indicate that, under normal growing temperatures,
photoperiod has an inÑuence on glucosinolate variation
throughout the day ; at stress temperatures, variations
are larger and are almost completely dependent on temperature. This masks the e†ect of photoperiod. The
inverse relationship between the aerial part and roots
suggested in the previous study (Rosa 1997) was only
partly supported at a constant 20¡C and constant light
and relative humidity (Fig 1), probably as an e†ect of
permanent light.
Indol-3-ylmethyl glucosinolate generally follows the
same trend as the other two major glucosinolates in the
aerial part of the plant and in the roots. When averaged
E A S Rosa, P M F Rodrigues
212
over all sampling times and in both experiments, levels
in the roots (127 ^ 50 lmol 100 g~1 DW) were signiÐcantly higher (P \ 0É001) than in the aerial part
(71 ^ 17 lmol 100 g~1 DW). SigniÐcant di†erences
(P \ 0É001) were noted between the two temperature
regimes in the aerial parts (67 ^ 13 lmol 100 g~1 DW
in Exp A and 75 ^ 21 lmol 100 g~1 DW in Exp B) and
roots (89 ^ 22 lmol 100 g~1 DW in Exp A and
165 ^ 39 lmol 100 g~1 DW in Exp B). However,
throughout the day, signiÐcant di†erences (P \ 0É05)
were noted only for the aerial part of the plant at 20¡C ;
at 30¡C and despite the large variation in the roots, no
signiÐcant di†erences were noted due to the large SE
errors.
The concentration of the two major glucosinolates in
the roots, 2-phenylethyl and 1-methoxyindol-3-ylmethyl
were signiÐcantly higher (P \ 0É001) than in the aerial
parts where levels were less than 7 lmol 100 g~1 DW.
Levels of both glucosinolates at a constant 20¡C were
signiÐcantly lower than at 30¡C (629 ^ 181 lmol
100 g~1 DW vs 727 ^ 475 lmol 100 g~1 DW for 2phenylethyl and 391 ^ 83 lmol 100 g~1 DW vs
431 ^ 152 lmol 100 g~1 DW for 1-methoxyindol-3ylmethyl). When plants were submitted to 30¡C,
changes in levels of 2-phenylethyl glucosinolate
throughout the day were larger (P \ 0É001) than at
20¡C. Thus, it seems that stress temperatures induced
larger 2-phenylethyl variations between sampling times,
which is in agreement with previous results (Rosa 1997).
For 1-methoxyindol-3-ylmethyl glucosinolate, signiÐcant di†erences (P \ 0É01) were noted throughout the
day only at 20¡C, the variation at 30¡C probably being
covered by the large SE. In common with other individual glucosinolates in this study, 2-phenylethyl and 1methoxyindol-3-ylmethyl also showed the ultradian
rhythms in a sinusoidal curve type with more or less
variation according to temperature but with the inÑuence due to light. Similar evolution trends were noted
under photoperiodic conditions (0700È2100) (Rosa
1997).
In comparison with the variations in the daily glucosinolate level observed either under Ðeld conditions
(Rosa et al 1994) or controlled climatic conditions (Rosa
1997), the present study reveals that no large changes
throughout the day could be observed when temperature, light and relative humidity are kept constant
at an optimum growth and development temperature ;
at temperatures close to stress, variations are larger.
CONCLUSIONS
There were large di†erences in total glucosinolate concentrations between the aerial part of the plant and the
roots when growing conditions, except temperature,
were kept constant. Even under constant conditions,
seedlings exhibit the ultradian rhythms for glucosinolates which are more pronounced when temperatures are
above the optimum for growth and development. At
optimum temperatures, photoperiod inÑuences the synthesis of major glucosinolates throughout the day ;
however, when the temperature is closer to that inducing stress, the e†ect of light is almost negligible, variations being mainly due to temperature. Thus, it is likely
that glucosinolates accompany the evolution of other
plant constituents throughout the day as a response to
external factors of environment.
REFERENCES
Clossais-Besnard N, Larher F 1991 Physiological role of glucosinolates in Brassica napus : concentration and distribution pattern of glucosinolates among plant organs during a
complete life cycle. J Sci Food Agric 56 25È38.
Duncan A J 1991 Glucosinolates. In : T oxic Substances in
Crop Plants, ed Felix DÏMello J P, Du†us C M & Du†us J
H. Royal Society of Chemistry, Cambridge, UK, pp 126È
147.
Fenwick G R, Heaney R K, Mullin W J 1983 Glucosinolates
and their breakdown products in food and food plants.
CRC Crit Rev Food Sci Nutr 18 123È201.
Fieldsend J, Milford G F J 1994a Changes in glucosinolates
during crop development in single- and double-low genotypes of winter oilseed rape (Brassica napus). I : Production
and distribution in vegetative tissues and developing pods
during development and potential role in the recycling of
sulphur within the crop. Ann Appl Biol 124 531È542.
Fieldsend J, Milford G F J 1994b Changes in glucosinolates
during crop development in single- and double-low genotypes of winter oilseed rape (Brassica napus). II : ProÐles and
tissue-water concentrations in vegetative tissues and
developing pods. Ann Appl Biol 124 543È555.
Heaney R K, Fenwick G R 1980 Glucosinolates in Brassica
vegetables : analysis of 22 varieties of Brussels sprouts
(Brassica oleracea var gemmifera). J Sci Food Agric 31 785È
793.
Josefsson E 1967 Distribution of thioglucosides in di†erent
parts of Brassica plants. Phytochemistry 6 1617È1627.
Macfarlane Smith W, Griffiths D W 1988 A time-course study
of glucosinolates in the ontogeny of forage rape (Brassica
napus L). J Sci Food Agric 43 121È134.
McGregor D I 1988 Glucosinolate content of developing
rapeseed (Brassica napus L “MidasÏ) seedlings. Can J Plant
Sci 68 367È380.
Rosa E 1997 Daily variation of glucosinolate concentration in
the leaves and roots of cabbage seedlings in two constant
temperature regimes. J Sci Food Agric 73 364È368.
Rosa E, Heaney R K, Rego F C, Fenwick G R 1994 The
variation of glucosinolate concentration during a single day
in young plants of Brassica oleracea var acephala and capitata. J Sci Food Agric 66 457È463.
Rosa E, Heaney, R K, Fenwick G R, Portas C A M 1996
Changes in glucosinolates concentrations in Brassica crops
(B oleracea and B napus) throughout growing seasons. J Sci
Food Agric 71 237È244.
Rosa E, Heaney, R K, Fenwick G R, Portas C A M 1997
Glucosinolates in crop plants. Hort Rev 19 99È215.
Salisbury F B, Ross C W 1992 Plant Physiology (4th edn).
Wadsworth, Belmont, CA, USA.
Spinks A, Sones K, Fenwick G R 1984 The quantitative
analysis of glucosinolates in cruciferous vegetables, oilseeds
and forage crops using high-performance liquid chromatography. Fette Seif Anstrichmitte 86 228È231.
Документ
Категория
Без категории
Просмотров
2
Размер файла
204 Кб
Теги
553
1/--страниц
Пожаловаться на содержимое документа