Docsity
Docsity

Prepare for your exams
Prepare for your exams

Study with the several resources on Docsity


Earn points to download
Earn points to download

Earn points by helping other students or get them with a premium plan


Guidelines and tips
Guidelines and tips

Inorganic Salts' Effects on Plant Membrane: Betacyanin Leakage & Leaf Senescence Study, Slides of Inorganic Chemistry

This document from a 1976 journal explores the effects of inorganic salts on plant membrane permeability using beet root slices and rumex obtusifolius l. Leaf discs. The study reveals that cacl2 decreases betacyanin leakage and delays senescence, while (nh4)2so4 increases leakage and hastens senescence. The authors suggest that these effects may be due to alterations of interactions between water and macromolecules in the tissues, and discuss the possible relationship between these effects and the macromolecular stabilization or destabilization properties of inorganic salts.

Typology: Slides

2021/2022

Uploaded on 09/12/2022

merielynd
merielynd 🇬🇧

4.7

(9)

218 documents

1 / 4

Toggle sidebar

This page cannot be seen from the preview

Don't miss anything!

bg1
Plant
Physiol.
(1976)
58,
182-185
Effects
of
Inorganic
Salts
on
Tissue
Permeability1
Received
for
publication
February
19,
1975
and
in
revised
form
February
24,
1976
B.
W.
POOVAIAH2
AND
A.
CARL
LEOPOLD3
Department
of
Horticulture,
Purdue
University,
ABSTRACT
Inorganic
solutes
are
shown
to
alter
the
permeability
of
root
and
leaf
tissues.
Experiments
with
beet
root
tissues
reveal
that
CaCI2
decreases
leakage
of
betacyanin
from
the
tissue,
that
(NH4)2SO4
increases
leakage,
and
that
each
salt
can
relieve
the
effects
of
the
other.
A
comparison
of
cations
and
anions
shows
a
range
of
effects
with
the
various
solutes.
Experiments
with
Rumex
obtusifolius
L.
leaf
discs
reveal
that
whereas
CaCI2
defers
the
development
of
senescence,
(NH4)2SO4
hastens
senes-
cence
and
increases
the
leakage
of
materials
out
of
the
leaf
discs.
The
solute
effect
on
Rumex
obtusifolius
L.
is
prevented
by
gibberellin.
CaC12
can
relieve
the
(NH4)2SO4
effect.
The
results
are
interpreted
as
indicat-
ing
that
the
inorganic
solutes
may
serve
to
alter
the
permeability
of
membranes
through
alterations
of
interactions
between
water
and
mac-
romolecules
in
the
tissues;
the
interpretation
is
consistent
with
the
evidence
for
opposite
effects
of
Ca
and
NH4,
the
effective
concentrations
being
about
1O-3
M,
and
the
reversibility
of
the
effects
of
one
solute
by
another
of
opposite
stabilization-destabilization
effect.
It
is
well
known
that
inorganic
salts
can
have
large
effects
on
the
configuration
of
macromolecules;
Hofmeister
(2)
described
the
range
of
effects
of
various
salts
on
protein
solubilities,
and
in
a
more
current
context,
von
Hippel
and
Wong
(16)
have
de-
scribed
the
effects
of
inorganic
salts
as
an
alteration
of
macromo-
lecular
stabilization
or
destabilization.
We
have
reported
that
inorganic
salts
can
alter
the
actions
of
each
of
the
known
plant
hormones
(3),
and
have
raised
the
question
of
whether
the
alterations
of
hormonal
actions
might
be
related
to
effects
of
the
solutes
on
plant
membranes.
It
is
the
intent
of
this
paper
to
explore
the
effects
of
inorganic
salts
on
some
plant
membranes,
and
to
consider
the
possibility
that
the
observed
effects
might
relate
to
their
effects
as
stabilization
or
destabilization
agents.
MATERIALS
AND
METHODS
Beet
roots
(Beta
vulgaris
L.)
were
purchased
locally
and
cut
into
sections
1
cm
in
diameter
and
1
to
2
mm
thick.
The
sections
were
washed
for
about
2
hr
in
several
changes
of
aerated
distilled
H20.
Five
sections
were
transferred
to
10
ml
of
test
solution.
Membrane
damage
was
assessed
by
spectrophotometric
deter-
mination
of
the
amount
of
betacyanin
pigment
that
leaked
out
into
the
ambient
solution
over
12-
to
18-hr
period.
For
the
Rumex
assay
(Rumex
obtusifolius
L.),
following
the
procedure
of
Whyte
and
Luckwill
(17),
old
leaves
which
were
uniformly
green
were
selected
from
matured
plants
in
the
green-
1
Journal
Paper
No.
5813,
Purdue
University
Agricultural
Experi-
ment
Station,
Lafayette,
Ind.
47907.
2
Present
address:
Department
of
Horticulture,
Washington
State
University,
Pullman,
Wash.
99163.
3
Present
address:
Graduate
College,
University
of
Nebraska,
Lin-
coln,
Neb.
68583.
Lafayette,
Indiana
47907
house.
Leaf
discs
(1
cm)
were
cut
with
a
cork
borer
and
random-
ized,
and
10
discs
were
floated
in
each
Petri
dish
containing
10
ml
of
solution
to
be
tested
in
the
darkroom
at
25
C
for
about
4
or
5
days.
Chl
was
extracted
in
ethanol.
Ten
leaf
discs
were
trans-
ferred
to
10
ml
of
ethanol
in
a
closed
vial
and
allowed
to
stand
overnight,
and
absorbance
was
read
at
665
nm.
The
absorbance
of
ambient
solution
was
measured
at
280
nm
to
assess
the
leakage
from
leaf
discs.
All
experiments
were
repeated
on
at
least five
different
occasions.
RESULTS
As
a
simple
test
of
membrane
permeability,
we
used
slices
of
beet
root
suspended
in
water,
and
measured
the
extent
of
beta-
cyanin
leaking
out
into
the
ambient
solution
as
an
index
of
membrane
leakiness.
As
a
comparison
of
an
extreme
macromo-
lecular
destabilizer,
CaCl2,
and
an
extreme
stabilizer,
(NH4)2SO4,
serial
concentrations
of
these
two
salts
were
applied
separately
to
beet
root
slices
(Fig.
1);
it
can
be
seen
that
CaC12
produced
a
decrease
in
the
amount
of
pigment
leaking
out,
whereas
(NH4)2SO4
caused
an
increase
in
the
amount.
If
these
opposite
effects
are,
in
fact,
related
to
destabilization
and
stabilization
effects,
then
it
should
be
possible
to
relieve
the
effects
of
each
salt
with
increasing
amounts
of
the
other.
In
order
to
test
that
possibility,
aseries
of
beet
root
sections
was
treated
with
3
x
10-3
M
(NH4)2SO4,
and
the
effects
of
increasing
amounts
of
CaCI2
were
examined
(Fig.
2).
It
can
be
seen
that
whereas
the
(NH4)2SO4
produced
a
marked
increase
in
the
amount
of
pigment
leaking
out
of
the
root
sections,
the
further
addition
of
CaCl2
at
slightly
above
10-3
M
CaCl2
brought
the
leakage
back
down
to
the
level
of
the
sections
in
water
alone.
The
converse
experiment
was
carried
out
as
shown
in
Figure
3;
here,
a
group
of
sections
was
treated
with
CaCI2
at
10-3
M
concentration,
and
the
effects
of
further
additions
of
(NH4)2SO4
were
examined.
It
can
be
seen
that
the
CaCI2
alone
reduced
the
leakage
of
pigment
by
about
half;
the
further
addition
of
(NH4)2SO4
brought
the
amount
of
leakage
back
to
the
value
of
water
controls.
These
two
experiments
indicate
that
the
effects
of
each
of
these
two
salts
may
be
erased
by
the
simultaneous
application
of
the
other
salt.
As
further
evidence
about
the
possibility
that
destabilization-
stabilization
effects
were
causing
the
leakage
responses
ob-
served,
we
made
a
comparison
of
various
cations
and
various
anions.
An
array
of
10
cations
was
added
as
their
chlorides,
each
at
3
x
10-3
M
concentration,
and
the
amount
of
pigment
leaking
out
of
the
beet
sections
is
shown
in
Table
I;
here
again,
ammo-
nium
caused
the
greatest
leakage,
and
Ca
depressed
leakage
by
more
than
half.
The
monovalent
ions
K,
Li,
and
Na
gave
a
graded
series
of
effects
toward
a
lessening
of
leakage,
and
the
divalent
cations
Mg,
Ca,
Ba,
Sr,
and
Mn
produced
greater
depression
of
leakage
in
that
order;
the
trivalent
lanthanum
was
markedly
more
effective
in
depressing
leakage,
as
expected
for
a
"supercalcium"
(7).
Various
anions
were
tested
as
the
K
salts
at
a
concentration
of
3
x
10-3
M,
and
the
comparative
effectiveness
of
the
five
anions
is
shown
in
Table
II.
Relatively
little
effects
were
obtained
for
phosphate,
chloride,
and
sulfate,
and
a
slight
182
pf3
pf4

Partial preview of the text

Download Inorganic Salts' Effects on Plant Membrane: Betacyanin Leakage & Leaf Senescence Study and more Slides Inorganic Chemistry in PDF only on Docsity!

Plant Physiol. (1976) 58, 182-

Effects of Inorganic Salts on Tissue Permeability

Received for publication February 19, 1975 and in revised form February 24, 1976

B. W. POOVAIAH2 AND A. CARL LEOPOLD

Department of Horticulture, Purdue University,

ABSTRACT

Inorganic solutes are shown to alter the permeability of root and leaf tissues. Experiments with beet root tissues reveal that CaCI2 decreases leakage of betacyanin from the tissue, that (NH4)2SO4 increases leakage, and that each salt can relieve the effects of the other. A comparison of cations and anions shows a range of effects with the various solutes. Experiments with Rumex obtusifolius L. leaf discs reveal that whereas CaCI2 defers the development of senescence, (^) (NH4)2SO4 hastens senes- cence and increases the leakage of materials out of the leaf discs. The solute effect on Rumex obtusifolius L.^ is prevented^ by^ gibberellin.^ CaC can (^) relieve the (^) (NH4)2SO4 effect. The results are interpreted as indicat- ing that the inorganic solutes may serve to alter the^ permeability^ of membranes through^ alterations^ of^ interactions^ between water and mac- romolecules in^ the tissues; the^ interpretation^ is^ consistent^ with^ the evidence for opposite effects of Ca and NH4, the^ effective^ concentrations being about 1O-3^ M, and the reversibility of^ the^ effects^ of^ one^ solute by another of opposite stabilization-destabilization effect.

It is well known that inorganic salts can have large effects on the configuration of macromolecules; Hofmeister (2) described the range of effects of various salts on protein solubilities, and in

a more current context, von Hippel and Wong (16) have de-

scribed the^ effects^ of^ inorganic salts^ as^ an^ alteration^ of^ macromo-

lecular stabilization or destabilization. We^ have^ reported that

inorganic salts can alter the actions of each of^ the known^ plant

hormones (3), and have raised the question of whether^ the

alterations of hormonal actions might be related to^ effects^ of the

solutes on plant membranes. It is the intent of this paper to

explore the^ effects^ of^ inorganic^ salts^ on some^ plant^ membranes,

and to consider the possibility that the observed effects might

relate to their effects as stabilization or destabilization agents.

MATERIALS AND METHODS

Beet roots^ (Beta vulgaris L.) were^ purchased locally and^ cut

into sections 1 cm^ in^ diameter and^1 to^2 mm^ thick. The^ sections

were washed for about 2 hr in several changes of aerated distilled

H20. Five sections were transferred to^10 ml of^ test^ solution.

Membrane damage was assessed by spectrophotometric deter-

mination of the amount of betacyanin pigment that leaked^ out

into the ambient solution over^ 12-^ to^ 18-hr^ period.

For the Rumex assay (Rumex obtusifolius L.), following the

procedure of Whyte and^ Luckwill^ (17), old leaves which^ were

uniformly green were^ selected^ from^ matured^ plants in the green-

1 Journal Paper No. 5813, Purdue University Agricultural Experi-

ment (^) Station, Lafayette, Ind. 47907. (^2) Present address: Department of Horticulture, Washington State University, Pullman, Wash. 99163. (^3) Present address: Graduate College, University of Nebraska, Lin- coln, Neb. 68583.

Lafayette, Indiana 47907

house. Leaf discs (1 cm) were cut with a cork borer and^ random- ized, and 10 discs were floated in each Petri dish^ containing^10 ml of solution to be tested in the darkroom at 25 C for about 4 or 5 days. Chl was extracted in ethanol. Ten leaf discs were trans- ferred to 10 ml of ethanol in a closed vial and allowed to stand overnight, and absorbance was read at 665 nm. The absorbance of ambient solution was measured at 280 nm to assess the leakage from leaf discs. All experiments were repeated on at least five different occasions.

RESULTS

As a simple test of^ membrane permeability,^ we^ used^ slices of beet root suspended in water, and measured the^ extent^ of beta- cyanin leaking out into the ambient solution as an index of membrane leakiness. As a comparison of an extreme macromo- lecular destabilizer, CaCl2,^ and^ an^ extreme^ stabilizer, (NH4)2SO4, serial concentrations of^ these^ two salts^ were^ applied separately to beet root slices (Fig. 1); it can be seen that CaC produced a decrease in the amount of pigment leaking out, whereas (NH4)2SO4 caused an increase in the amount. If these opposite effects are, in fact, related to destabilization and stabilization effects, then it should be possible to relieve the effects of each salt with increasing amounts of the other. In order to test that possibility, a series of beet root sections was treated

with 3 x 10-3 M (NH4)2SO4, and the effects of increasing

amounts of CaCI2 were examined (Fig. 2). It can be seen that

whereas the (^) (NH4)2SO4 produced a marked increase in the amount of pigment leaking out of the root sections, the further addition of (^) CaCl2 at slightly above 10-3 M CaCl2 brought the leakage back down to the level of the sections in water alone. The converse experiment was^ carried out^ as shown^ in^ Figure^ 3; here, a group of sections was^ treated^ with^ CaCI2^ at^ 10-3^ M concentration, and the effects of further^ additions^ of^ (NH4)2SO

were examined. It can be seen that the^ CaCI2^ alone^ reduced^ the

leakage of pigment by about half; the^ further addition^ of (NH4)2SO4 brought the amount of leakage back to^ the value of water controls. These two^ experiments^ indicate^ that^ the^ effects of each of these two^ salts^ may be^ erased^ by the simultaneous application of^ the other salt. As further evidence about the possibility that destabilization- stabilization effects were causing the leakage responses ob- served, we made a comparison of various cations and^ various

anions. An array of 10 cations^ was^ added^ as^ their^ chlorides, each

at 3 x^ 10-3^ M^ concentration, and the^ amount^ of^ pigment leaking

out of the beet sections is shown^ in^ Table^ I; here^ again, ammo-

nium caused the greatest leakage, and Ca depressed leakage by

more than half. The monovalent ions K, Li, and^ Na^ gave a

graded series of effects toward^ a^ lessening of^ leakage, and the

divalent cations^ Mg, Ca, Ba, Sr, and^ Mn^ produced greater

depression of^ leakage in that^ order; the^ trivalent^ lanthanum^ was

markedly more effective in depressing leakage, as^ expected for^ a

"supercalcium" (7). Various anions^ were^ tested^ as^ the K salts^ at

a concentration of^3 x^ 10-3^ M, and the^ comparative effectiveness

of the five anions is shown in Table II. Relatively little effects

were obtained for phosphate, chloride, and^ sulfate, and^ a^ slight

182

Plant Physiol. Vol. 58, 1976

0

0

CD .4 w

SALTS AND PERMEABILITY

2x1I0 2 2x162 m CONC. OF^ SA LT FIG. 1.^ Effects of^ (NH4)2S04 and^ CaC12^ on^ leakage^ of^ betacyanin from beet slices.

0 In a 0 w CD 4 4 -i

also caused an increase in leakage out of the leaf sections. Even in the presence of GA, an enhancement of leakage by (NH4)2SO4 was evident at the higher concentrations. If the enhancement of the rate of leaf senescence and of leakage by (NH4)2SO4 is related^ to^ the solute^ effecting the destabilization-stabilization state of^ the^ membranes, then^ one would expect that^ CaCl2^ additions^ might^ relieve^ the^ effects^ of^ the (NH4)2SO4. Experiments^ in^ which various^ concentrations^ of CaCl2 were added in the incubation medium are presented in Figure 5, from which it can be seen that even in the presence of the (NH4)2SO4, CaCI2 was able to defer the rate of senescence, as indicated by the maintenance^ of^ higher Chl^ contents^ in^ the leaf discs. The lower^ figure indicates^ that^ the^ leakage of^ materi- als absorbing at 280 nm^ was^ stimulated^ by (NH4)2SO0 as^ was seen previously in Figure 4, but the^ CaCI2 additions relieved^ the effects of the 3 x^ 10-3 M (NH4)2SO4 both^ with respect to^ Chl retention (Figure 5, above) and limiting the^ development of leakiness (Fig. 5, below).

0

0 0

(^04) w 4 w -i

1.2-

0.6-

0.2v

J II CONC. CO C FIG. 2. Promotion of betacyanin leakage from beet slices^ with 3^ mM (NH4)2S04 and the reversal of its effect with increasing concentrations of CaC12-

reduction of leakage was obtained with^ nitrate^ and^ carbonate. Collectively, these cation and anion^ experiments indicate^ that the leakiness effects are not^ specific to^ special cations^ or^ anions, but that (^) graded series of effects can be obtained with a wide array of anions and cations. Some further experiments were done with another tissue: the 1-cm discs of Rumex leaves. We had previously reported that the senescence of corn leaf discs was associated with a marked increase in leakiness, as measured by apparent free space and by permeability to tritiated water (8). It is known that the senes- cence of Rumex leaves is deferred by gibberellin (17), and that CaCI2 also markedly slowed the development of senescence (8) and abscission (9). In the present experiments, Rumex leaf discs were treated with various concentrations of (NH4)2SO4 to deter- mine whether this stabilizing salt would have effects opposite to those of CaCl2. Effects were measured in the presence and in the absence of GA. Chi content was used as a measure of senes- cence, and the^ relative^ absorbancy^ of the ambient^ solution^ at 280 nm was taken as an indication of the amount of leakage of organic materials from the leaf discs. The data in Figure 4 indicate that (NH4)2SO4 at concentrations of 10-3 M and above enhanced the rate of leaf senescence, as indicated by the greater loss of Chl, than in control pieces. While GA deferred senescence effectively, the addition of (NH4)2SO in the presence of the GA did not alter the rate of leaf senes- cence. The measurements of leakage out of the leaf discs indi- cate that the (NH4)2SO4 concentrations of 3 x 10-3 M^ or above

1O-4 - CONC. (NH4)2 S FIG. 3.^ Inhibition^ of^ betacyanin leakage from beet^ slices^ with^1 mm CaC12 and the reversal of its^ effect^ with^ increasing concentrations^ of (NH4)2S04.

Table I. Comparative Effects of Cations on Leakage of Betacyanin from Beet Slices Each cation supplied as the chloride at 3 x^ 10-3 M concentration.

Solute Betacyanin leakage

supplied (OD540)

H20 0.266 + 0.

Ammonium 1.300 +^ 0. Potassium 0.325 -^ 0. Lithium 0.200 - 0. Sodium 0.180 -^ 0. Magnesium 0.160 -^ 0. Calcium 0.115 -^ 0. Barium 0.111 + 0. Strontium 0.098 - 0. Manganese 0.030 - 0. Lanthanum 0.017 ±^ 0.

(NH4)2SO

coci

I

0.so

SALTS AND PERMEABILITY

that ammonium salts could increase the leakage of photosyn- thates out of cotton leaf cells, and Ca having the opposite effect. In some experiments with^ dynamic pumping^ systems,^ Satter^ et al. (13) have shown that the rhythmic movements^ of^ Albizzia pulvini are caused to exercise an opening action in the presence of calcium salts, a closing action in the presence of ammonium salts, and lesser or intermediate effects of other cations in^ the Hofmeister series. These experiments can be interpreted^ as indicating that inorganic solutes of the Hofmeister^ series^ can increase permeability of membranes or^ decrease the^ permeabil- ity, depending upon the stabilization or^ destabilization^ effects of the solutes. Through the experiments on^ interactions between solutes, our experiments have added strength to^ the^ concept^ of solute effects being through the^ destabilization-stabilization^ ef- fects. Our special interest in the solute alterations of membrane functions lies in the area of hormonal action; if hormones act through an attachment to a site of action on a membrane, then solute alterations of membrane characteristics might be expected to alter hormonal effectiveness.^ We^ have shown that^ each^ of the plant hormones is altered^ in^ effectiveness^ by^ calcium^ and^ ammo- nium salts (3, 11), and that^ the^ attachment^ of^ auxin^ to^ mem- brane pieces from^ corn^ coleoptiles can^ also be^ altered^ by^ these salts (10).

LITERATURE CITED

  1. GARY-BoBo, C. M. 1970. Effect of Ca-+ on the water and non-electrolyte permeability of phospholipid membranes.^ Nature^ 228: 1101-1102.
  2. HOFMEISTER, F. 1888. Zur Lehre von der Wirkung der Salze. Zweite Mitteilung. Arch.

Exp. Pathol. Pharmakol. 24: 247-260.

  1. LEOPOLD,^ A. C.,^ B.^ W.^ POOVAIAH,^ R. K.^ DELA^ FUENTE,^ AND^ R.^ J. WILLIAMS.^ 1974. Regulation of growth with inorganic^ solutes.^ In: Plant^ Growth^ Substances, 1973.^ Hiro- kawa Publishing Co., Tokyo. pp. 780-788.
  2. LEVIN, Y. K., A. G. LEE, N. J. M. BIRDSALL, J. C. METCALFE, AND J. D. ROBINSON. 1973. The interaction of paramagnetic ions and spin labels with lecithin bilayers. Biochim. Biophys. Acta^ 291:^ 592-607.
  3. (^) MARINOS, N. (^) G. 1962. Studies on submicroscopic aspects of mineral deficiencies. I. Calcium deficiency in the shoot apex of barley. Am. J. Bot. 49: 834-841.
  4. MURAKAMI, S. AND L. PACKER. 1971. The role of cations in the organization of chloroplast membranes. Arch. Biochem. Biophys. 146: 337-347.
  5. PICKARD, B. G. 1970. Comparison of calcium and lanthanum ions in the Avena coleoptile growth test. Planta 91: 314-320.
  6. POOVAIAH, B. W. AND A. C. LEOPOLD. 1973. Deferral of leaf senescence with calcium. Plant Physiol. 52: 236-239.
  7. POOVAIAH, B. W. and A. C. LEOPOLD. 1973. Inhibition of abscission by calcium. Plant Physiol. 51: 848-851.
  8. POOVAIAH, B. W. AND A. C. LEOPOLD. 1976. Effects of inorganic solutes on the binding of auxin. Plant Physiol. In press.
  9. (^) POOVAIAH, B. W. AND A. C. LEOPOLD. 1976. Modification of hormonal responses with ammonium sulfate. Plant Physiol. In press.
  10. REHFELD, D. W. AND R. G. JENSEN. 1973. Metabolism of separated^ leaf cells.^ Ill. Plant Physiol. 52: 17-22.
  11. SATTER, R. L., P. B. APPLEWHITE, D. J. KREBS, AND A. W. GALSTON. 1973. Rhythmic leaflet movement in Albizzia julibrissin. Plant Physiol. 52: 202-207.
  12. VAN STEVENINCK, R. F. M. 1965. The significance of calcium on the apparent permeability of cell membranes and the effects of substitution with other divalent ions. Physiol. Plant 18: 54-69.
  13. VON HIPPEL, P. H. AND T. SCHLEICH. 1969. The effects of neutral salts on the structure and conformational stability of macromolecules in solution. In: S. N. Timashef and G. D. Fasman, eds., Structure and Stability of Biological Macromolecules. Marcel Decker Inc., New York. pp. 417-573.
  14. VON HIPPEL, P. H. AND K. Y. WONG. 1964. Neutral salts: the generality of their effects on the stability of macromolecular conformations. Science 145: 577-580.
  15. WHYTE, P. AND L. C. LUCKWILL. 1966. A sensitive bioassay for gibberellins based on retardation of leaf senescence. Nature 210: 1360-1361.

Plant Physiol. Vol. 58, 1976