全 文 ：西北植物学报 2003, 23( 6): 954— 957
Acta Bot . Boreal.-Occident. Sin .
文章编号: 1000-4025( 2003) 06-0954-04
渗透胁迫对绿豆叶圆片叶绿素荧光的影响
杨甲定 ,赵哈林 ,张铜会
(中国科学院寒区旱区环境与工程研究所 生态与农业研究室 ,兰州 730000)
摘　要: 绿豆叶圆片用 0、 - 0. 3、 - 0. 6、 - 1. 2、 - 1. 8、 - 2. 4 M Pa等 6个渗透梯度处理 24 h后 ,其叶绿素荧光的反
应表明:光系统Ⅱ的潜在量子产量 (Fv /Fm )和电子传递受体 (QA )的库容 ( poo l size )均随胁迫强度的增大而减小 ,
但 QA库容的下降明显比 Fv /Fm的下降缓和 . 结合原初荧光 Fo和最大荧光 Fm的变化 ,分析认为本实验中渗透胁
迫并未导致明显的光系统Ⅱ反应中心的失活或破坏 . 造成光系统Ⅱ潜在量子效率降低的原因主要是通过光系统Ⅱ
反应中心到 QA的电子传递受到阻抑 .
关键词: 叶圆片 ;绿豆 ;渗透胁迫 ;叶绿素荧光
中图分类号: Q945. 11　　　文献标识码: A
Effects of osmotic stress on chlorophyll f luorescence in
leaf discs of mungbean (Phaseolus aureus )
YANG Jia-ding , ZHAO Ha-lin, ZHANG Tong-hui
( Departmen t of Ecolog y and Ag ricul tu re Research , Cold and Arid Region s Environm ental and Engineering Research Ins ti tute,
Chinese Academy of Sciences , Lan zh ou 730000, China)
Abstract: Leaf discs o f mungbean ( Phaseolus aureus Roxb. ) w ere submitted to six o smo tic t reatments, i.
e. , 0, - 0. 3, - 0. 6, - 1. 2, - 1. 8, - 2. 4 M Pa , for 24 hours. Af ter dark adaptation, randomly-selected 10
leaf discs w ere measured using a por table f luo rometer ( Handy PEA, Hansatech Inst ruments Ltd. , U K) in
the da rk room. Results show ed that the potential quantum yield of PSⅡ and the poo l size of PSⅡ elect ron
t ranspor t accepters (QA ) w ere bo th reduced by treatments of o smo tic st ress. The mild o smo tic treatments
o f - 0. 3 and - 0. 6 M Pa had smaller, w hi le modera te and severe t reatments had la rg er influence. Associat-
ed wi th the Fo and Fm values in all t reatments, i t w as suggested that o smo tic st ress had sligh t ef fects on
the poo l size of QA and did no t induce the apparent inactiva tion or damage to PSⅡ reaction centers. The
main reason fo r the reduced po tential quantum yield of PSⅡ ( Fv /Fm ) wa s the blocking o f elect ron t rans-
port f rom PSⅡ reaction center to QA .
Key words: leaf discs; mungbean; osmotic st ress; chlo rophyll f luo rescence
　　 It has long been repo rted tha t inhibi tion of leaf
photosynthesis is one of the common metabo lic
va ria tions of plants during period of wa ter
st ress[1～ 3 ] , as found w ith low and high tempera-
ture[ 4, 5] , salini ty st ress[6 ] etc. This inhibition can be
resulted f rom leaf stomatal closure, inhibi tion of
chlo roplast membrane reactions ( elect ron t ranspo rt
and photopho spho rylation) , and inf luenced pho to-
synthetic enzymes, etc
[ 2]
. Besides gas exchange
measurement of pho to synthesis ra te and in vit ro
收稿日期: 2003-02-24;修改稿收到日期: 2003-04-30
基金项目:国家重点基础研究规划发展项目 ( G2000048704) ;中国科学院创新项目资助 ( CHX210048)
作者简介:杨甲定 ( 1971- ) ,男 (汉族 ) ,在读博士 ,从事植物逆境生理与植物基因工程研究 .
assays of elect ron t ranspo rt rate and enzyme activ i-
ties, chlorophyll fluo rescence emit ted f rom PSⅡ
ref lects the state of pho to synthesis in vivo. This
method has found w idespread use in pho to synthe-
sis resea rch because i t is quickly measured and not
dest ructiv e to leaf ma terials
[7～ 9 ]
.
The objectiv e of this study w as to examine
changes in pho tochemical ef ficiency of PSⅡ in leaf
discs during a set of o smotic st resses, thus to in-
vestiga te the possible acting si tes of osmo tic st ress
on the light reaction process o f pho tosynthesis.
1　Materials and methods
1. 1 Plant material
　　 The experimental field is located in the south-
w est bo rder o f Ho rqin sandy land, belong ing to the
eastern pa rt o f Inner M ongolia, China ( 42°55′N,
120°42′E, 345 m above mean sea lev el) . M ungbean
( Phaseolus aureus Roxb. ) seeds w ere g row n and
seedling s w ere managed exact ly according to local
cultiva tion modes. The experiments w ere carried
out in the fi rst ten days of August w hen mungbean
plants w ere at anthesis stag e. Fo r a ll experiments,
fully-expanded mature leaf lets ( on dif ferent com-
pound leaves) w ere used.
1. 2　Osmotic stress treatment
Around 7: 00 A. M. ( Beijing time) about three
hours af ter sunrise, an appropria te number of com-
pound leaves was removed from plants and brought
to the labo rato ry wi th the petioles soaked in dis-
ti lled w ater. Leaf discs ( diameter= 1. 5 cm ) were
removed from the lamina betw een the major veins
and immersed in PEG8000 solutions wi th w ater po-
tential of - 0. 3, - 0. 6, - 1. 2, - 1. 8, - 2. 4 M Pa
respectiv ely , acco rding to Ha rdeg ree and Em-
merich [10 ] . Simul taneously, a set o f leaf discs wa s
put into distilled w ater as the contro l o f 0 M Pa.
These six sets o f t rea ted lea f discs w ere put a t the
room temperature ( 25～ 27℃ ) under natural light /
dark ( 16 h /8 h) condition for 24 hours.
1. 3　 Chlorophyll fluorescence
After dark adapta tion fo r at least 30 min, the
potential quantum yield o f PSⅡ ( Fv /Fm ) and pool
size of primary elect ron acceptor (QA ) ( Area ) of
random ly-selected 10 leaf discs were determined
using a po rtable f luo rometer ( Handy PEA, Hansat-
ech Inst ruments Ltd. , U K) in the dark room. The
sa turating PPFD of the actinic light w as 3000μmo l
· m- 2· s- 1 and the time of ex ci tation w as 0. 6 s. In
order to compare the pool size of PSⅡ elect ron
t ranspo rt accepto r (QA ) in dif ferent samples, the
Area ha s to be normalized by Fv . The normalized
expression is defined as: Sm≡ Area /(Fm- Fo ) [ 11 ] .
2　 Results
2. 1　 Chl f luorescence parameterFv /Fm
　　 As shown in Fig. 1, th e values o f Fv /Fm de-
clined slight ly f rom 0 to - 0. 6 M Pa. Treatment of
- 1. 2, - 1. 8 and - 2. 4 M Pa resulted in the de-
crease of Fv /Fm as 62% , 36% and 28% o f the con-
trol respectiv ely. Obv iously it w as indicated that
the potentia l quantum yield of PSⅡ ( Fv /Fm )
w ould be reduced by wa ter stress. The mild w ater
stress ( - 0. 3 and - 0. 6 M Pa) had smal ler ef fects
than moderate wa ter stress ( - 1. 2 and - 1. 8
M Pa) and severe st ress ( - 2. 4 M Pa ) on the de-
cline of Fo /Fm in leaf discs of mungbean.
Fig. 1　 Values o f Fv /Fm in leaf discs with six
trea tments o f osmo tic str ess fo r 24 h
( Ver tical ba rs indica te the standa rd err or s,n= 10)
2. 2　 Initial (Fo) and maximal (Fm ) f luorescence
Measurements of Fo and Fm in leaf discs in
six osmo tic t reatments ( Fig. 2) show ed that os-
mo tic stress had very sligh t ef fects on Fo but
caused appa rent reduction in Fm w ith the intensi-
fying of o smo tic stress. So the decrease of Fv /Fm
( Fig. 1) i s mainly due to the declined Fm rather
955
6期 杨甲定 ,等:渗透胁迫对绿豆叶圆片叶绿素荧光的影响 (英 )
than the Fo lev el.
Fig. 2　 Values o f the initial (Fo) a nd maximal (Fm )
fluo r escence in leaf discs with six trea tments
of o smotic stress for 24 h
( V ertical bar s indicate the standard er ro rs, n= 10)
2. 3　 The pool size of PSⅡ electron transport ac-
ceptorQA (Sm )
Compared w ith the 0 M Pa control, Sm in five
t reatments of osmotic st ress decreased ( Fig. 3) .
But the t rends w ere much gentler than tho se of
Fv /Fm ( Fig. 1) . Sm in t reatments o f - 0. 3 and -
0. 6 M Pa w as clo se, w hile in - 1. 2 and - 1. 8 M Pa
treatments wa s simi lar. In - 3. 0 M Pa trea tment ,
Sm was about 80% of that in 0 M Pa contro l.
Fig. 3　Values of QA pool size of ( Sm )
in the six trea tments o f osmo tic str ess fo r 24 h
( V ertical bar s indicate the standard er ro rs, n= 10)
3 Discussion
The po tential quantum yield of PSⅡ as indi-
cated by Fv /Fm has been shown to be propo rtional
to the quantum yield of pho tochemist ry and to be
related clo sely to the quantum yield of net pho to-
synthesis[12 ] . So the decreases o f the Fv /Fm values
in osmotic t reatments f rom - 0. 3 to - 2. 4 M Pa
( Fig. 1) indicated that osmotic st ress inf luenced
the int rinsic status o f leaf discs and led to reduction
in pho tochemical yield of PSⅡ .
In the present experiment , the no rmalized pa-
rameter Area ( the area above the fluo rescence
curve between Fo and Fm , i. e. , Sm ) i s proportion-
al to the pool size of the electron accepto rs QA on
the reducing side o f PSⅡ ( Users manual, Handy
PEA, Hansa tech Instruments Ltd. , U K) . In other
w ords, Sm is a measure o f the energy needed to
clo se all reaction centers
[11 ]
. Judged from Fig. 3, i t
w as show n that the pool size o f QA in t reatments of
- 0. 3 to - 2. 4 M Pa declined mo re slight ly com-
pared wi th the values of Fv /Fm. For example,Fv /
Fm in - 2. 4 M Pa treatment was 31% , whi le Sm
was 80% of that in 0 M Pa trea tment respectiv ely.
These resul ts meant that there should be some oth-
er facto rs responsible for the larg e decrease o f the
po tential quantum yield o f PSⅡ besides the slight-
ly reduced pool size o f QA , although the slight ly de-
creased Sm may decrease the Fv /Fm , as indicated
by Woodrow& Berry
[13 ]
and Xu et al
[ 8] . It wa s sug-
gested that damage or inactiv ation of PSⅡ reaction
centers wil l result in increasing of the ini tial f luo-
rescence (Fo )
[ 7, 14] . As show n in Fig. 2, Fo were ap-
proxima tely similar, i. e. , no net increase of Fo in
t rea tments o f o smo tic stress. This meant that there
w as no reversible inactiva tion or ir rev ersible dam-
age in PSⅡ reaction centers. On the other side, the
maximal f luorescence ( Fm ) ref lects the elect ron
t ranspo rt th rough PSⅡ [15 ] . Wi th the intensifying of
osmo tic stress, Fm was apparently reduced ( Fig.
2) . This st rongly indicated that the elect ron t rans-
po rt f rom the PSⅡ reaction centers to the quinone
pool was partially blocked o r the quanti ty of t rans-
ferred elect ron decreased. These resul ts w ere com-
patible w ith the fo rmer repo rts o f st ress-induced
956
西　北　植　物　学　报 23卷
inhibi tion of the ligh t reactions of pho to synthe-
sis
[1 ]
.
Briefly , in the present w o rk, o smo tic st ress
had sligh t ef fects on the pool size of QA and did not
induce the apparent inactiva tion or damage to PSⅡ
reaction centers in leaf discs of mungbean. The
main reason fo r the reduced potential quantum
yield o f PSⅡ ( Fv /Fm ) w as the pa rtially blocking
of elect ron transport f rom PSⅡ reaction center to
QA .
参考文献:
[1 ]　 HANSON A D, HIT Z W D. Metabolic responses of mesophytes to plant w ater deficit s [ J ]. Ann . Rev. Plant Physiol . , 1982, 33: 163- 203.
[2 ]　 BERRY J A, DOWN TON W J S. Envi ronmen tal regulation of photosyn th esis [ A ] . Govind jee, ed s. Ph otos ynth esi s (V ol. Ⅱ ) [ C ] . New
Yo rk: Academic Press , 1982: 263- 343.
[3 ]　 BOW LER C M, V AN M ONT AGU M, INZE D. Superoxid e dismutase and s tress tolerance[ J ]. Ann . Rev. P lant Physiol . Plant Mol .B iol. ,
1992, 43: 83- 116.
[4 ]　 DU FF T D, BEARD J B. Sup raoptimal temp erature effect s upon A grost is p alustr is. Part Ⅱ . Inf luence on carbohyd rate level s, ph otos yn-
th etic rate, and respi rat ion rate [ J] . Physiol . Plant , 1974, 32: 18- 22.
[5 ]　 HODGSON R A J, RAISON J K. Superoxide production b y thylakoids du ring chi lling and i ts implication in th e sus ceptibi lit y of plan ts to
chil ling-ind uced ph otoinhibiti on [ J] . Plan ta , 1991, 183: 222- 228.
[6 ]　 HEU ER B, PL AUT Z. Ph otos ynthesis and osmot ic adju stment of tw o sugarbeet cul tivars grow n und er s aline condit ions [ J ]. J . Exp . Bot. ,
1989, 40: 437- 440.
[7 ]　 KRAUSE G H, WEIS E. Ch lorophyl l fluores cence and ph otos ynthesis: the basics [ J ]. Ann. Rev . Plant Physiol Plant Mol . Biol . , 1991, 42:
313- 349.
[8 ]　X U D Q(许大全 ) , ZHANG Y ZH(张玉忠 ) , ZHANG R X(张荣铣 ) . Ph otoinhibi ti on of ph otosynthesis in plants [ J ]. Plant Physiology Com -
municat ion (植物生理学通讯 ) , 1992, 28( 4): 237- 242( in Chines e) .
[9 ]　 GOV IND JEE. Six ty-th ree years since Kau tsk y: chloroph yll a f luorescence [J ]. Aust. J . Plant Physiol . , 1995, 22: 131- 160.
[10 ]　 HARDEGREE S P, EMM ERICHW E. Ef fect of polyethylene glycol exclusion on the water potential of solu tion-satu rated fi l ter paper
[ J] . Plant Physiol. , 1990, 92: 462- 466.
[11 ]　 STRASSER R J, SRIV ASTAV A A, TSIM ILLI-M ICHAEL M. The f luo rescence t ransien t as a tool to ch aracteri ze and screen ph otos yn-
th etic s am ples [ A]. Yunus M , Path re U, M oh an ty P, eds. Probing photosynthesis: mech anism, regulation& adaptation [ C] . Lond on: Tay-
lor & Francis , 2000: 445- 483.
[ 12 ]　 GEN TY B, BRIAN TAIS J M , BAKER N R. Th e relationsh ip betw een th e quan tum yield of ph otos ynthetic elect ron t ransport and
quenching of chloroph yll f luorescence[ J] . Biochim .B iophys. Acta. , 1989, 990: 87- 92.
[13 ]　 WOODROW I E, BERRY J A. Enzymatic regulation of ph otos ynthetic CO 2 fixat ion in C3 plants [ J] . Ann . Rev. Plant Physiol. Plant Mol.
B iol. , 1988, 39: 533- 594.
[14 ]　 DEM MIG B, WIN TER K, KRUGER A, CZYGAN F C. Photoinhibi tion and zeaxanthin fo rmation in intact leav es. a possible role of the
xanthophyl ls cycle in the dis sipation of excess light energy [ J] . Plant Physiol . , 1987, 84: 218- 224.
[15 ]　 ZHAN G S R(张守仁 ) . A dis cus sion on chloroph yll fluores cence kin etics parameters and thei r sig ni ficance [J ]. Chinese Bul let in of Botany
(植物学通报 ) , 1999, 16( 4): 444- 448( in Chines e) .
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6期 杨甲定 ,等:渗透胁迫对绿豆叶圆片叶绿素荧光的影响 (英 )