Anda di halaman 1dari 6

Plant Cell Physiol.

27(2): 187-192 (1986)


JSPP © 1986

Effect of Sodium Application on Growth of


Amaranthus tricolor L.

Toru Matoh, Daisaku Ohta and Eiichi Takahashi

Plant Nutrition Laboratory, Department of Agricultural Chemistry,


Faculty of Agriculture, Kyoto University, Kyoto 60S, Japan

Sodium application of 0.5 meq./liter to water-cultured Amaranthus tricolor L,. cv.


Tricolor plants brought about a three-fold increase in dry matter production compared

Downloaded from pcp.oxfordjournals.org by guest on January 12, 2011


with those of plants deprived of sodium. This increase was due to sodium itself and
not to a supplementary effect of sodium on potassium shortage nor to the accompanying
anion. In sodium-sufficient plants, the total nitrogen and potassium contents were
lower and calcium, chlorophyll and betacyanin contents were higher than in the deficient
plants.

Key words: Amaranthus tricolor — Cj-plants — Sodium requirement.

Many plant species show positive growth response to sodium. The effect of sodium,
however, has been ascribed to a poor supply of potassium, that is, sodium can in part compensate
for the shortage of potassium. (Flowers and Lauchli 1983). Brownell and Wood (1957) first
demonstrated the essentiality of sodium for the growth of Atriplex vesicaria, and their group has
done extensive studies on the function of this element. Brownell and Crossland (1972, 1974)
demonstrated that the sodium requirement is remarkable only in C4 and CAM plants, and
not in C3 plants. In two genera of Chenopodiaceae, Atriplex and Kochia, which contain both
C3 and C4 species, sodium was shown to be essential only for the C4 species (Brownell 1979).
In this paper, we report the sodium requirement of the NAD-malic enzyme type C4 plant
Amarantus tricolor L. cv. Tricolor, and discuss the possible functions of this element.

Materials and Methods


Plant materials and growth conditions—Seeds ofAmaranthus tricolor L. cv. Tricolor were purchased
from Takii Seed Co., Ltd., Kyoto, Japan, in May 1983. The seeds used in the growing seasons
of 1984 were obtained from plants grown under sodium-deficient conditions in 1983 in our
greenhouse. The seeds (1 g) were washed with distilled water five times each for 5 min. The
seeds were sown on a sheet of cheesecloth covering acid-washed polyethylene beads (diameter
5 mm, packed in a 20 X 25 X 3 cm polyethylene container) wetted with distilled water, and were
kept at 30°C under continuous illumination (8,000 lux). After germination, a half-strength
culture solution (see below) was given and just before the second leaf pair appeared, the seedlings
were transplanted to water culture, using 3-liter plastics pots. The solution was aerated
for 30 min every 2 hours without air purification. The pots were kept in a greenhouse,
where they were dipped into a water bath with circulating water of 20°C. The standard culture
solution had the following salts; 1 mti KC1, 0.5 nw MgSO 4 -7H 2 O, 0.25 mM (NH 4 )2HPO 4 and
1 mM Ca(NO3)2-4H 2 O. The salts were purified by recrystallization from ethanol-water, except

187
188 T. Matoh, D. Ohta and E. Takahashi

for calcium nitrate. The micronutrient composition was that of Arnon's solution cited in
Hewitt (1966) except that all the iron was supplied as ferrous-citrate. The culture solutions
were prepared using distilled and deionized water. Sodium was supplied as NaCl or Na2SO4
and in the reference treatment, KC1 or K2SO4 was supplemented to give the same anion con-
centration. Every pot had four or two seedlings and the solution was changed every four or
two days, as the plant growth proceeded.
The growth experiments were carried out for four times in the summer of 1983 and 1984.
In the 1983 experiments, the plants were allowed to grow until their seed formation stage.
The sodium-deprived plants had only about 20% seeds compared with the sodium-supplied
plants, but the germination efficiency was not different.
Analyses—The plants were dissected into leaves, stems and roots, and the parts were washed
with distilled water, blotted dry, weighed and dried in an oven at 70°C. The materials were
finely ground using a ball mill and appropriate amounts were digested with a nitric acid-sulfulic

Downloaded from pcp.oxfordjournals.org by guest on January 12, 2011


acid mixture (10 : 1) at 150°C. Sodium and potassium contents were determined by emission
spectrophotometry and calcium, magnesium and iron by atomic absorption spectrophotometry.
Total nitrogen and phosphorus contents were determined colorimetrically using the indophenol
blue (Weatherburn 1967) and the molybdenum blue method (Murphy and Riley 1962), respec-
tively, after Kjeldahl digestion. Chlorine was extracted with boiling water and measured by
a colorimetric method (Iwasaki et al. 1952). Chlorophyll content was estimated in 80% (v/v)
acetone-water extracts (Arnon 1949) and betacyanin content in 67% (v/v) methanol-water
extracts (Elliott 1979) prepared from the fresh leaves.

Results
Growth—Dry matter production by A. tricolor under various growth conditions is shown in
Fig. 1. Growth of the plants receiving sodium salts was about 300% of those grown without
sodium salts. As the reference plants received potassium salts instead of sodium salts to give
the same anion concentrations, the growth stimulation induced by the application of sodium
salts is a sodium specific effect and is due neither to compensation of potassium shortage nor to
the accompanying anion.
Mineral contents—Mineral contents of A. triclor plants grown under various conditions are
shown in Tables 1 and 2. For elements other than Na and K, only the contents in the leaves
are presented. The plants supplied with K2SO4 at 0.5 mM showed poorer growth (Fig. 1) and
lower chlorine and potassium contents (Table 1) than the sodium-deprived plants. Sodium
contents in the leaves increased 40- to 50-fold with addition of sodium salts at 0.5 or 1 mM, but
the distribution of this element among the organs was almost the same as in the sodium-deprived

Table 1 Contents of sodium, potassium and chlorine in A. tricolor plants under various conditions
Na(ppm) K (%) Cl (ppm)
Leaves Stems Roots Leaves Stems Roots Leaves
Standard 32.6 48.9 110 6.69 12.7 9.03 5,940
+0.5 mM KC1 41.8 54.3 93.9 7.30 13.9 11.3 6,740
+1 mM KC1 32.5 38.9 68.8 6.53 12.9 10.8 7,070
+0.5 mM K2SO4 59.6 56.2 54.3 4.38 7.46 5.58 3,350
+0.5 mM NaCl 1,700 6,500 6,930 4.80 9.26 5.19 9,150
+ 1 mM NaCl 2,430 10,500 6,300 4.08 7.25 5.29 8,570
+0.5 mM Na 2 SO 4 2,650 12,400 7,520 4.30 9.16 5.86 8,220
The values are the means of three replicates.
Sodium requirement of Amaranthus tricolor 189

Standard
+ 0.5 mM KCI
+ I mM KCI

+ 0.5 mM K2SO«

+ 0.5 mM NaCI

+ I mM NaCI

+ 0.5 mM No2SO4

Downloaded from pcp.oxfordjournals.org by guest on January 12, 2011


Fig. 1

Fig. 1 Dry matter production of Amaranthus tricolor plants grown with or without sodium salts. The seeds were
sown on May 4, 1984. Sodium salts were given for 27 days and the plants were harvested on July 9, 1984. The
results are shown as an average of three plants and the bars show standard deviations. The growth experiment was
repeated three times with similar results.
Fig. 2 Acidification of the Amaranthus tricolor rooting medium by sodium salt application. The medium pH
changes were traced in the 1984 growing period using the same plants as in Fig. 1. Determinations were done from
the 17th day after sodium salt application. Standard culture solution (-O-); standard culture solution containing
0.5 meq./liter chloride salts ( ) of sodium (•) and potassium (O), the same at 1 meq./liter cation concentration
( ); the same at 1 meq./liter cation concentration, but given as sulfate salts ( ).

plants. A. tricolor plants responded to sodium but the sodium content and distribution in the
sodium-supplied plants are not different from those of other crop plants grown under ordinary
conditions, for which the sodium requirement has not been reported (Flowers and Lauchli 1983).
The chlorine content (Table 1) was somewhat higher in the sodium-recieving plants, but even
that of the reference plants far exceeded the critical content for chlorine deficiency (Broyer et al.
1954).
Comparing the mineral contents between + NaCI and +KC1 plants, significant differences
were found also for N, K and Ca contents. Nitrogen and potassium contents were higher in
the sodium-deprived plants. These results suggest that sodium improves the efficiency of

Table 2 Contents of nitrogen, phosphorus, calcium, magnesium and iron in A. tricolor plant leaves under various
conditions
Culture conditions N (%) P (%) Ca (%) Mg (%) Fe (ppm)
Standard 5.91 ±0.249 0.743 ±0.010 2.84±0.100 1.12 ±0.058 128±5.50
+0.5 mM KCI 6.20±0.370 0.855 ±0.058 2.85±0.064 1.21 ±0.067 N.D.
+1 mM KCI 5.99±0.050 0.807 ±0.071 2.54±0.50 1.08 ±0.130 249±12.4
+0.5 mM K2SO4 5.50±0.189 0.509±0.044 3.33±0.162 1.01 ±0.025 213±10.0
+0.5 mM NaCI 5.20±0.161 0.722 ±0.058 3.01 ±0.020 1.08 ±0.052 N.D.
+1 mM NaCI 5.28±0.I17 0.628 ±0.023 3.20±0.016 1.12 ±0.027 331 ±12.5
+0.5 mM Na2SO4 5.I0±0.U6 0.641 ±0.006 3.17 ±0.067 1.16±0.063 205 ±4.49
The values are the means of three replicates±standard errors. N.D., Not determined.
190 T. Matoh, D. Ohta and E. Takahashi

Table 3 Effect of sodium application on the chlorophyll and betacyanin contents


in the leaves of A. tricolor plants

Treatment (mg/gfr P wf») , B f ^cyanin


(Chlorophyll a/b ratio) ^ m ° l e s / g fr- w t - ' )

Without sodium (+0.5 mi* KC1) J 1.49 0.305


(3.41)

With sodium (+0.5 mu NaCl) ° 2.30 1.23


(3.72)

" 0.5 DIM KC1 or 0.5 HIM NaCl was supplemented to the standard culture solution.
* Leaves just reaching their maximum sizes were used.
Determinations were carried out six times using different plants.

Downloaded from pcp.oxfordjournals.org by guest on January 12, 2011


nitrogen and potassium utilization; sodium enhances dry matter production markedly with
relatively little increase in N and K uptake. Phosphorus contents did not significantly differ
between the sodium-supplied and sodium-derpived plants. Calcium contents are generally
higher in the sodium-receiving plant leaves but no differences were found in the stems and roots.
Magnesium and iron contents in the leaves were also determined (Table 2), since the sodium-
deprived plants easily suffered from chlorosis. Although significant fluctuations were observed
in the iron contents, magnesium contents were not different.
Medium pH changes—Fig. 2 shows the medium pH changes. The pH decrease was faster
in the Na-containing medium. Although this difference was reproducible, it is not clear at
present whether sodium has a direct effect on this pH decrease. Since the sodium-receiving
plants grow vigorously, the faster pH drop in the sodium-containing medium may be due to a
greater unbalance of uptake between cations and anions, that is, the secondary effect of sodium.
Betacyanin—Brownell and Crossland (1972) demonstrated that the sodium-deprived A. tricolor
plants showed the deficiency symptoms of chlorosis and necrosis. We also noted symptoms
of sodium deficiency such as pale leaf color and low betacyanin contents (Table 3), but usually
did not find necrosis.

Discussion
Under our growth conditions, the contaminating sodium in the culture solutions was less
than 20 ppb, and the sodium content in the leaves of plants grown without added sodium were
30 to 60 ppm (Table 1) although in the steins and roots they were generally higher. According
to Brownell (1979), his group reduced the concentration of contaminating sodium in the culture
solutions to less than 2 ppb, but the sodium content in their Atriplex vesicaria leaves, showing
sodium-deficiency symptom, was 10 mmol/kg dry matter, equaling 230 ppm. Kushizaki and
Yasuda (1964) reported that when sugar beet plants showed sodium-deficiency symptoms, the
internal sodium content was 6 mmol/kg dry leaves. Comparing these findings indicates that
the critical concentration of sodium at which plants suffer sodium deficiency may differ from
species to species.
Brownell (1979) also pointed out that the sodium requirement of the Ci-Atriplex plants was
met by application of about 0.1 meq./liter sodium. Our results (Fig. 1) indicate that application
of 0.5 meq./liter of sodium satisfies the sodium requirement of A. tricolor but the precise threshold
concentration of sodium for the plants was not determined. Assuming that all the sodium in
the leaves was free, the sodium concentrations were calculated to be about 0.2 nut in the sodium-
deprived leaves and about 10 DIM in the sodium-supplied leaves on the tissue water basis.
Sodium requirement of Amaranthus tricolor 191

Accordingly, if the system(s) which requires sodium occurs in the leaves, the internal threshold
concentration of sodium is in a mM range.
The physiological ground for the sodium requirement has never been made clear. Nable
and Brownell (1984) reported that in the sodium-deficient Amaranthus leaves, alanine accumula-
tion is significant compared with the contents in the sodium-sufficient plant leaves. They suggest
that sodium is required for in vivo activation of pyruvate orthophosphate dikinase. However,
we are not aware of any report on the sodium requirement of maize and sugar cane, representative
C4-plants having photosynthesis of the NADP-malic enzyme type. Hewitt (1983) summarized
the sodium effect on the C4 plant species and suggested that aspartate-forming C4-plants respond
to sodium. Comparison of the photosynthetic capacities between sodium-supplied and -deprived
plants is now in progress in our laboratory.
Nunes et al. (1983) demonstrated a NaCl-stimulated proton efflux from sugar beet leaf discs.
When the leaf discs absorbed sodium, stoichiometric release of potassium and proton occurred

Downloaded from pcp.oxfordjournals.org by guest on January 12, 2011


under light. They concluded that sugar beet has a sodium-stimulated proton pump and
discussed the possibility of the pump being a sodium-activated ATPase. As A. tricolor belongs
to the same family, Chenopodiaceae, as the sugar beet, there might be some similar response to
sodium as to proton efflux in the A. tricolor plants.
Elliott (1979) investigated cytokinin effects on betacyanin biosynthesis in dark-grown A. tricolor
seedlings and his intensive studies revealed that there is a Na-K synergism for the cytokinin-
dependent betacyanin synthesis. Elliott (1979) suggested involvement of a Na-K ATPase
activity, but its occurrence in higher plants has not yet been demonstrated. Our results (Table 3)
indicate a four-fold increase in the betacyanin contents in mature leaves. As cytokinin is known
to participate in many diverse aspects of plant cell metabolism, sodium may operate through
the cytokinin functions.

References

Arnon, D. I. (1949) Copper enzymes in isolated chloroplasts. Polyphenoloxidase in Beta vutgaris. Plant Physiol.
24: 1-5.
Brownell, P. F. (1979) Sodium as an essential micronutrient element for plants and its possible role in metabolism.
Advances Bot. Res. 7: 117-224.
Brownell, P. F. and C. J. Crossland (1972) The requirement for sodium as a micronutrient by species having the
C4 dicarboxylic photosynthetic pathway. Plant Physiol. 49: 794-797.
Brownell, P. F. and C.J. Crossland (1974) Growth responses to sodium by Bryophyllum tubiflorum under conditions
inducing crassulacean acid metabolism. Plant Physiol. 54: 416—417.
Brownell, P. F. and J. G. Wood (1957) Sodium as an essential micronutrient element for Atriplex vesicaria, Heward.
Nature 179: 635-636.
Broyer, T. C , A. B. Carlton, C. M. Johnson and P. R. Stout (1954) Chlorine—A micronutrient element for
higher plants. Plant Physiol. 29: 526-532.
Elliott, D. C. (1979) Ionic regulation for cytokinin-dependent betacyanin synthesis in Amaranthus seedlings. Plant
Physiol. 63: 264-268.
Flowers, T.J. and A. Lauchli (1983) Sodium versus potassium: Substitution and compartmentation. In Ency-
clopedia of Plant Physiology, N.S., vol. 15B, Inorganic plant nutrition. Edited by A. Lauchli and R. L. Bieleski.
pp. 651-681 Springer-Verlag.
Hewitt, E. J. (1966) The composition of the nutrient solution. In Sand and Water Culture Methods, pp. 190.
Commonwealth Agricultural Bureaux, Farnham Royal Bucks, England.
Hewitt, E.J. (1983) Essential and functional metals in plants. In Metals and Micronutrients: Uptake and
Utilization by Plants. Edited by D. A. Robb and W. S. Pierpoint. pp. 313-315. Academic Press.
Iwasaki, I., S. Utsumi and T. Ozawa (1952) New colorimetric determination of chloride using mercuric thiocyanate
and ferric ion. Bull. Chem. Soc. Japan 25: 226.
Kushizaki, M. and T. Yasuda (1964) Seasonal variations of inorganic and organic constituents of sugar beets
during their growth II. Sodium deficiency symptoms. Rts. Bull. Hokkaido Nat. Agr. Exp. Station. 84: 46-51.
192 T. Matoh, D. Ohta and E. Takahashi

Murphy, J . and J . P. Riley (1962) A modified single solution method for the determination of phosphate in natural
waters. Anal. Chimica Acta. 27: 31-36.
Nable, R. O. and P. F. Brownell (1984) Effect of sodium nutrition and light upon the concentrations of alanine in
leaves of C4 plants. Aust. J. Plant Physiol. 11: 319-324.
Nunes, M. A., M. M. Correia and M. D. Lucas (1983) NaCl-stimulated proton efflux and cell expansion in sugar-
beet leaf discs. Planta 158: 103-107.
Weatherburn, M. W. (1967) Phenol-hypochlorite reaction for determination of ammonia. Anal. Chem. 39: 971—
974.

(Received June 18, 1985; Accepted November 18, 1985)

Downloaded from pcp.oxfordjournals.org by guest on January 12, 2011