全 文 ：植　　　物　　　研　　　究
BULLETIN OF BOTANICAL RESEARCH
第 19 卷　第 3 期 1999 年　7 月
Vol.19　No.3 July , 　1999
碱性盐胁迫下星星草幼苗中几种渗透调节物质的变化
阎秀峰1　孙国荣2　李　晶3　肖　玮2
(1.东北林业大学森林植物生态学开放研究实验室 , 哈尔滨 150040)
(2.哈尔滨师范大学生物系 , 哈尔滨 150080)　(3.东北农业大学基础部 ,哈尔滨　150030)
摘　要　模拟松嫩盐碱草地碱化土壤的离子组成配制碱性盐溶液处理星星草
(Puccinellia tenui f lora)幼苗 ,测定了碱性盐胁迫下星星草幼苗地上部分几种渗透
调节物质含量的变化 。随着碱性盐浓度及胁迫天数的增加 ,Na+、可溶性盐 、游离脯
氨酸和可溶性糖的含量逐渐增加 ,其中游离脯氨酸的变化倍数最大。K+含量在浓
度小于 20g L-1碱性盐胁迫下随胁迫天数增加而略为减少 ,在 40 g L-1和 60 g L-1
碱性盐胁迫下先是降低 ,而后迅速增加 。
关键词　星星草;碱性盐胁迫;渗透调节物质
CHANGES OF SEVERAL OSMOTICA IN PUCCINELLIA
TENUIFLORA SEEDLING UNDER ALKALI SALT STRESS
Yan Xiu-feng 1　Sun Guo-rong2　Li Jing1　Xiao Wei2
(1.Open Research Labo rato ry of Forest Plant Ecology , Northeast Forestry University , Harbin 150040)
(2.Department of Biology , Harbin Normal University , Harbin 150080)
(3.Northeast Ag riculture University , Herbin 150030)
Abstract　Puccinellia tenuif lora seedlings w ere t reated by alkaline salt solution that
was made up according to ions composition of the alkaline soil of Songnen alkaline
grassland.Contents of several osmotica in above -ground part of P.tenui f lora
seedling under alkaline salt stress w ere determined.With the increment of alkaline salt
concentrat ion and st ress day s , content of Na+ , soluble salt , f ree proline and soluble
carbohydrate increased g radually , and the change of f ree proline is the g reatest.Under
the stress of alkaline salt w hich concentration is smaller than 20 g L-1 , K+content de-
creased a lit tle w ith the stress days , and under the concentration of 40 g L -1 and 60 g
L
-1 , K+ content decreased fi rst , and then increased rapidly.
收稿日期:1997-3-10
Key words　Puccinellia tenui f lora;Alkaline salt st ress;Osmotica
Osmoregulation means the course that make the balance between osmotic po tential in cell
and the potent ial energy of external envi ronment through removing or adding solute to the cell.
It includes the catatonosis that is decreasing cell osmot ic potential by adding solute , and also in-
cludes anatonosis that is increasing cell osmotic po tential by removing solute.Osmo tic adjust-
ment means that solute concentration of cell increases and cause the decreasing of osmotic po-
tential w hen advanced plant are under stress such as drought and salinity〔1〕.It indicates a kind
of ability that plant can continue to abso rb w ater f rom the ex ternal medium w ith a low w ater
potential through the increasing of cell solute concentration , and the decreasing of cell osmotic
potential(as a result of the decreasing of water potential).This can make plant keep a certain
cell turg or pressure and maintain no rmal metabolism under st ress〔2〕.There are tw o kinds of so-
lutes that can adjust the osmotic potential.One is ino rganic ions that get into the cell f rom the
ex ternal environment , the other is organic solute that is synthesized inside the cell , including
amino acid , betaine , soluble carbohydrate , polyhydric alcohol and so on , of w hich proline and
betaine have generally been researched.Among all the amino acids proline is the most ef fective
to osmotic adjustment.Whether the plant is st ressed by drought , high temperature , low tem-
perature , f reezing , salt , low pH , malnutrition or disease , proline will be accumulated.Plant
w ill have the most accumulation of proline especially under drought st ress , i t can be scores o r
even hundreds times higher than original content〔3〕.Under salt stress , Whether lower plant o r
higher plant , halophy te or g lycophy te , proline will be accumulated〔4〕.
Puccinellia tenui f lora is a perennial graminaceous plant that has a st rong tolerance to salt
and alkali.It dist ributes w idely in the Songnen alkaline g rassland.It is used in improving alka-
line g rassland〔5〕.Osmotic adjustment of P.tenuif lora seedling under the stress of neutral salt
NaCl has been researched
〔6 , 7〕.In this paper , we made up the alkaline salt solution according to
the ion ing redients of the soil in Songnen alkaline g rassland , and treated P.tenui f lora
seedlings with this solution in order to research the changes of several osmotica in above -
g round part of P.tenui f lora seedling.
1　Materials and methods
1.1　Culture and treatment of P.tenui f lora seedlings
The plastic pots w hich diameter is 25cm were full filled w ith pearlite(graininess , w ithout
pollution), fill Hoagland solution into the pots until the appearance of pearlite was moist.
Sowed the seeds of P .tenui f lora on the pearlite , and cultured in the controlled environment
chamber.Day/night temperature is 25℃/20℃, light application is 65 klx , and relative hu-
midity is 75%.After the seedlings grew two leaves they were t reated by salt solution.Accord-
ing to the analysis results of the alkaline soil sample of 0 ～ 20 cm layer in Songnen grassland ,
we made up the alkaline salt stock solution wi th Na2CO3 , NaHCO3 , K2CO3 , Na2SO4 and
M gCl2.The Seedlings w ere divided into 6 groups.According to the quantity of solution in each
348 植　　物　　研　　究　　　　　　　　　　　　　　　19 卷
plastic pot calculated the quantity of stock solution that each concentration need.Fill the stock
solut ion to the bo ttom of pearlite by a funnel , refill proper quantity of Hoagland solution to
make 6 kinds of concentration in the 6 g roups of plastic pots to 0 , 5 , 10 , 20 , 40 , 60 g L-1.
1.2　Determination of osmotica content
According to the method of He Zhili〔8〕 , the fresh sample w as homogenated by redistilled
w ater then filter it , and Na+ , K+ content of colature w ere determined w ith 6400-f lame pho-
tometer.Soluble salt content w as determined w ith DDS-11A conductivity meter according to
conductometry.Free proline content w as determined according to the method of Zhang Zhil-
iang〔9〕.Soluble carbohydrate content w as determined according to Zhang Zhenqing〔10〕.
Osmotica content is calculated by unit f resh w eight of the above -g round part of P.
tenui f lora seedling .Because the w ater condition of the plant will change under salt st ress(es-
pecially high concentration salt stress), the osmotica content based on unit fresh weight w ill
bring some false appearances because of the change of w ater higher than that of dry w eight.In
this experiment , seedlings of P .tenui f lora were t reated in a short time , the water content
changed lit tle(only w hen the salt concentration is higher than 40g·L-1 water content changed
obviously , Table1).
Table 1　Relative water content in the above -ground part of Puccinellia tenuif lora
seedling after 6 days alkaline salt stress
S alt concent rat ion(g·L -1) 0 5 10 20 40 60
Relat ive w ater content(%) 86.04 86.24 86.02 85.44 83.75 80.75
2　Results and Discussion
2.1　Changes of K+ , Na+ and soluble salt content in above-ground part of P.tenui f lora
seedlings under alkali salt stress
Af ter 1 day salt stress , Na+ content in the above-g round part of P .tenui f lora seedling
increased obviously , and then continue to accumulate.3 days later the speed of accumulation
became smaller , af ter 5 day s Na+ content w as almost stable (Fig.1).
Na
+
is a kind of basic ion wi thout act ivation in plant.Too much Na+ will do harm to
plant by dest roying the enzyme to a protein conformation w ithout activi ty.In fact , the me-
chanics of plant tolerance to salt st ress is the problem of the metabolism of Na+ and other
ion〔11〕.Fig 2 show ed the changes of Na+ content under different alkaline salt concentration.
Within the concentration of 10 g·L-1 , Na+ content in the above-ground part of P .tenuif lo-
ra seedling had a linear increment w ith the stress.From 10 g·L-1 to 40g·L-1 , the accumula-
tion speed of Na+ became slow.When the concentrat ion w as higher than 40g·L-1 , Na+ con-
tent increased rapidly again.The inhibited grow th condit ion of P.tenui f lora seedling indicat-
ed , salt stress of 5 g·L-1 almost had no effect and inhibition of 10 g·L-1 was also not very
strong
〔12〕.So w e can see P .tenui f lora seedling can bear the accumulation of certain quantity
3493 期　　　　　　　阎秀峰等:碱性盐胁迫下星星草幼苗中几种渗透调节物质的变化
Fig.1　Changes of K +, Na+and soluble salt content in the above-g round part of Puccinellia tenui flora
seedling under alkaline salt stress
Na
+
w ithout affect ing its normal grow ing.This probably because the bioplasm of P.tenuif lo-
ra seedling is adaptive to Na+ , small quantity of Na+ is no t enough to hurt the plant.Another
possibility is that Na+ is isolated in the sap vesicle and does no harm to bioplasm.From 10 g·
L-1 to 40 g·L-1 salt stress , the g row th of above-g round part of P.tenui f lora seedling began
to be inhibited.At the same time the accumulation speed of Na+ content slowed down , i t indi-
cates that P .tenui f lora seedling had active reaction to the ex ternal environment w ith high
concentration of Na+.
Fig.2　The contents of Na+ , K + and
soluble salt of the Puccinellia
tenuif lora seedling above -
g round part after 6 day s alka-
line salt stress
Many haloduric plants have selective absorption to in-
organic ions f rom ex ternal envi ronment.So they can ab-
sorb more favorable ions and less harmful ions〔13〕.The ex-
periment results indicated that P .tenuif lora also has this
ability .But the ability of this selective absorption is limi ted
in some ex tent.When the salt stress is too strong (higher
than 40 g·L-1)to the P .tenui f lora seedling , the mem-
brane w ill be destroyed completely , and the selective ab-
sorption will be lost
〔14〕.As a result all kinds of ions en-
tered the cell(Fig.1 and Fig.2), and the P.tenuif lora
seedling will he severely destroyed〔12〕.
Most plants have antagonistic action to the absorbing
of K+ and Na+〔15〕 , that is to say K+ content will decrease
w ith the increasing of Na+ content.Fig.1 and Table 2
showed the same results.The difference is that K+content
350 植　　物　　研　　究　　　　　　　　　　　　　　　19 卷
didn t decrease af ter 4 day s salt st ress of 40g·L -1and 3 days salt st ress of 60 g·L-1 , on the
o ther hand it increased rapidly.This possibly because the membrane sy stem of P .tenui f lora
seedling had been dest royed and a lot of K+ entered the cell.
Soluble salt content of above-ground part of P .tenuif lora seedling is almost equivalent
to the sum of all kinds of inorg anic ions.The change of soluble salt content is similar to that of
Na+ under alkaline salt stress(Fig.1 and Fig.2).
Table 2　The value of K+/Na+ in above-ground part of Puccinell ia tenuiflora seedling
under alkaline salt stress
St ress days
S alt concen trat ion(g·L -1)
0 5 10 20 40 60
1 8.40 5.72 4.82 4.20 3.24 2.24
2 8.86 4.84 4.04 3.10 2.10 1.00
3 7.86 4.81 3.39 2.62 1.80 1.14
4 7.81 4.61 3.33 2.30 1.95 1.45
5 7.73 4.46 3.18 2.14 2.03 1.65
6 7.80 4.35 3.07 2.08 2.05 1.66
2.2　Changes of f ree proline and soluble carbohydrate content in above -g round part of P.
tenui f lora seedling under alkaline salt st ress.
The accumulation quanti ty of free proline under 5 g·L-1 salt st ress w as very small.But
speed of accumulation increased rapidly under the salt stress , which concentration is 5 ～ 20g·
L-1 then it slow ed down when the salt concentration is over 20 g·L-1.The accumulation
speed of soluble carbohydrate was almost stable.The carbohydrate content had a linear incre-
ment w ith salt stress(Fig.3 and Fig .4).In addition , the increment times of f ree proline and
soluble carbohydrate w ere far more different.The content of free proline after 6 days salt st ress
of 60 g·L-1 was 15.47 times of the original content , and soluble carbohydrate was only 2.31
times of the o riginal content.Though free proline content is small , i ts accumulation speed
largely exceeds that of soluble carbohydrate.There w as a good correspondence between the salt
stress concentration at w hich free proline began to accumulate rapidly and the salt st ress concen-
tration at which the above-ground part of P.tenui f lora seedling began to be inhibited〔12〕.
This show ed the close relation betw een free proline and osmotic adjustment.
2.3　Comparison of several measured osmotica
Though some of the inorganic ions(such as Na+)are harmful to the action of plant , they
have osmotic adjustment function w hile accumulated under low concentration salt st ress as long
as the quantity is not enough for doing any harm.The total amount of these osmotica that been
had determined is the sum of inorg anic ions(that is soluble salt including Na+ and K+), f ree
proline and soluble carbohydrate(Table 3).Whether the increment of salt stress concentration
o r the increment of st ress time all make the total amount of osmotica increasing.
3513 期　　　　　　　阎秀峰等:碱性盐胁迫下星星草幼苗中几种渗透调节物质的变化
Table 3 The total amount of several measured osmotica(mg g-1FW)
St ress days
S alt concen trat ion(g·L -1)
0 5 10 20 40 60
1 25.775 25.859 27.000 27.899 28.849 30.304
2 25.947 27.361 28.276 29.644 31.525 37.227
3 25.959 27.827 29.669 31.966 36.310 48.792
4 26.339 28.236 30.107 33.794 40.341 53.524
5 26.098 28.386 30.822 34.776 42.433 55.328
6 26.510 28.468 31.493 35.578 43.267 57.174
Fig.3　The changes of free pro line and soluble carbohydrate content in above-g round par t of Puccinellia
tenuif lora seedling under alkaline salt stress
Fig.4　The content of free proline and
soluble carbohydrate in above-
ground part of Puccinellia
tenuif lora seedling under 6 day s
alkaline salt stress
According to the experiment results the proportion of
each osmoticum in the to tal amount of measured osmo tica
can be calculated(Table 4 to 8).The proportion of soluble
carbohydrate w as relatively stable with bo th the changes of
salt st ress concentration and st ress time(Table 8).But the
proportion of inorg anic ions (soluble salt including Na+
and K+)and free proline changed a lot (Table 4 to 7).
That is to say w ith the changes of st ress intensity and
stress time , the function of each osmoticum changed too.
Generally , when the stress intensity o r the st ress t ime of
certain concentrat ion increases , the proportion of free pro-
line increases (Table 7)and the proportion of inorg anic
ions decrease(Table 4).Conclusion is that , in the course
of osmotic adjustment under alkaline salt st ress , the func-
tion of inorganic ions maybe mo re important at low concentration salt st ress or the prophase of
stress , and org anic osmotica especially f ree proline maybe more important at high concentration
salt st ress or the anaphase of st ress.
352 植　　物　　研　　究　　　　　　　　　　　　　　　19 卷
Table 4 The proportion of soluble salt in the measured osmotica(%)
St ress days
S alt concen trat ion(g·L -1)
0 5 10 20 40 60
1 77.60 77.56 77.96 77.88 77.59 77.43
2 77.63 78.66 77.89 76.39 74.63 75.42
3 77.53 79.01 76.80 75.20 72.73 72.24
4 77.94 78.35 76.41 73.69 70.81 70.37
5 77.62 77.93 75.31 71.96 68.63 69.44
6 78.02 77.78 75.17 71.19 68.35 69.33
Table 5 The proportion of Na
+
in the measured osmotica(%)
St ress days
S alt concen trat ion(g·L -1)
0 5 10 20 40 60
1 3.28 4.74 5.39 5.89 7.15 9.71
2 3.17 5.28 6.04 7.42 9.86 16.71
3 3.45 5.25 6.78 7.82 9.90 13.94
4 3.42 5.30 6.72 8.05 9.43 12.82
5 3.49 5.39 6.82 8.30 9.31 12.52
6 3.42 5.42 6.77 8.17 9.24 12.25
Table 6 The proportion of K+ in the measured osmotica(%)
St ress days
S alt concen trat ion(g·L -1)
0 5 10 20 40 60
1 27.53 27.11 26.00 24.74 23.20 21.79
2 27.37 25.57 24.41 23.02 20.71 16.72
3 27.13 25.24 22.99 20.48 17.85 15.86
4 26.72 24.44 22.40 18.49 18.41 18.62
5 27.00 24.03 21.70 17.78 18.95 20.71
6 26.69 23.56 20.82 16.97 18.92 20.29
3533 期　　　　　　　阎秀峰等:碱性盐胁迫下星星草幼苗中几种渗透调节物质的变化
　　Table 7 The proportion of free prol ine in the measured osmotica(%)
St ress days
S alt concen trat ion(g·L -1)
0 5 10 20 40 60
1 1.25 1.26 1.23 1.64 2.13 2.60
2 1.32 1.22 1.78 3.33 5.08 5.77
3 1.28 1.23 3.44 5.35 7.44 8.30
4 1.24 1.42 3.98 6.68 7.84 8.64
5 1.26 1.63 4.40 7.18 8.45 8.67
6 1.25 1.79 4.47 7.30 8.52 8.69
Table 8　　The proportion of soluble carbohydrate in the measured osmotica(%)
St ress days
S alt concen trat ion(g·L -1)
0 5 10 20 40 60
1 21.15 21.18 20.81 20.48 20.28 19.97
2 21.05 20.11 20.33 20.28 20.30 18.81
3 21.19 19.76 19.76 19.46 19.83 19.46
4 20.82 20.23 19.60 19.63 21.34 20.99
5 21.12 20.44 21.22 20.86 22.92 21.88
6 20.73 20.43 20.35 21.52 23.13 21.98
Generally , it will take less energy using inorganic ions as osmo tica than small molecule o r-
ganic substance , because the synthesis of org anic substance consumes energy and it seizes sub-
st rate w ith respirato ry metabolism〔16〕.Though P .tenuif lora seedling didn t show severe af-
fected symptom , the g row th had been severely inhibited〔12〕.Acco rding to the results of this
experiment , on one hand the inhibition due to the toxic action of salt st ress , on the other hand
i t may have some relation w ith the energy consumption of osmotica accumulation.At the pre-
sent time , high content of free proline in plant is considered favorable to st rong tolerance , but
at the same time it is one reason of i ts low production〔7〕.
Otherw ise , though the proportion of inorganic ions decreased gradually (Table 4), the
composi tion probably changed.The results of the experiment showed that af ter 6 day s st ress
the propo rtion of Na+ increased w ith the increment of salt solution concentration(Table 5)and
the propo rt ion of K+ decreased with the increment of salt concentration (Table 6).Content
changes of Na+ and K+ are mainly af fected by the function of membrane.Under low concen-
tration salt stress , membrane has not been dest royed.It s selective absorption keeps a large
quantity Na+ out of the cell.At the same time , Na+ entered the cell are isolated in the sap
vesicle , this will keep the Na+ from doing harm to phy siology metabolism action , and these
Na+also have the funct ion of osmotic adjustment.Under high concentration salt stress , the
membrane w ill be severely dest royed.It s selective absorption w as lost〔14〕.So a large quantity
354 植　　物　　研　　究　　　　　　　　　　　　　　　19 卷
of Na+ can enter into the cell.The toxic action can lead a series of disorders of metabolism ac-
tion , it inhibi ts the grow th and even hurt the plant to death.Under salt st ress the ratio of K+/
Na
+
in plant is related to the tolerance closely.The bigger the ratio is the st ronger tolerance the
plant has.So a lo t of people considered the ratio K+/Na+ as an index to estimate the vitality
and the ability of cell maintaining its ion balance〔7〕.
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3553 期　　　　　　　阎秀峰等:碱性盐胁迫下星星草幼苗中几种渗透调节物质的变化