Environmental Pollution 133 (2005) 265–274
www.elsevier.com/locate/envpol
Influence of temperature and salinity on heavy metal
uptake by submersed plants
Å. Fritioff*, L. Kautsky, M. Greger
Department of Botany, Stockholm University, S-106 91 Stockholm, Sweden
Received 21 November 2003; accepted 31 May 2004
Metal concentrations increase with increasing temperature and decreasing
salinity in two aquatic plants.
Abstract
Submersed plants can be useful in reducing heavy metal concentrations in stormwater, since they can accumulate large amounts
of heavy metals in their shoots. To investigate the effects of water temperature and salinity on the metal uptake of two submersed
plant species, Elodea canadensis (Michx.) and Potamogeton natans (L.), these plants were grown in the presence of Cu, Zn, Cd, and
Pb at 5, 11, and 20 C in combination with salinities of 0, 0.5, and 5&. The metal concentrations in the plant tissue increased with
increasing temperature in both species; the exception was the concentration of Pb in Elodea, which increased with decreasing
salinity. Metal concentrations at high temperature or low salinity were up to twice those found at low temperature or high salinity.
Plant biomass affected the metal uptake, with low biomass plants having higher metal concentrations than did high biomass plants.
Ó 2004 Elsevier Ltd. All rights reserved.
Keywords: Elodea canadensis; Potamogeton natans; Phytoremediation; Stormwater; Cd; Cu; Zn; Pb
1. Introduction
Submersed plants may be useful in reducing heavy
metal concentrations in stormwater, since they have the
ability to take up heavy metals directly from water and
accumulate them in their shoots (St-Cyr et al., 1994;
Coquery and Welbourn, 1995; Greger et al., 1995; Rai
et al., 1995; Jackson, 1998). Stormwater comprises
rainwater and meltwater draining from hard surfaces,
such as roads and roofs in urban areas, and often
contains heavy metals, in particular Cu, Zn, Cd, and Pb
(Morrison, 1989; Sansalone et al., 1996; Boller, 1997;
Pettersson et al., 1999). The metal uptake of submersed
plants may be influenced by several factors, including
* Corresponding author. Tel.: C46 8 161348; fax: C46 8 165525.
E-mail address: fritioff@botan.su.se (Å. Fritioff).
0269-7491/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.envpol.2004.05.036
pH, redox potential, and surrounding metal concentrations as well as the temperature and salinity of the
surrounding stormwater. The chemistry of stormwater is
highly variable depending on its origin, on rain intensity,
and on the season; its pH, however, is often around 7
(Pettersson, 1999). In a temperate climate, like that of
Sweden, the temperature and salinity of stormwater
vary with the season: the water temperature may exceed
20 C in the summer and can decrease to below 0 C in
the winter. Salinity is very low most of the year, but
salinities up to 5& have been measured (Wittman, 1979;
Sandersson, 1997; Christer Lännergren, pers. comm.)
during winter when NaCl is used for deicing roads.
During summer, slightly elevated salinities up to 0.5&
can be measured (Wittman, 1979; Sandersson, 1997;
Christer Lännergren, pers. comm.), probably due to the
resuspension of sediment (Odum, 2000).
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Å. Fritioff et al. / Environmental Pollution 133 (2005) 265–274
Water temperature may influence water chemistry,
metal solubility, metal uptake by plants, and plant
growth. According to Zumdahl (1992), seasonal variation in water temperature has no direct effect on the
solubility of metal in water. However, cool water
contains more dissolved oxygen than does warm water.
Thus, metal concentration in the interstitial water of the
sediment may decrease with decreasing temperature, as
more metals are bound to sediment colloids at high
rather than low redox potentials (Förstner, 1979). In
addition, temperature has a profound effect on plant
growth rates and higher temperatures will thus result in
greater biomass production and distribution of submersed macrophyte communities (Marschner, 1995;
Rooney and Kalff, 2000). A plant of relatively high
biomass may have a greater metal uptake capacity; this
results from lower metal concentration in its tissue
because of a growth rate that exceeds its uptake rate
(Ekvall and Greger, 2003). Changes in temperature
further change the composition of the plasma membrane
lipids (Lynch and Steponkus, 1987). This alters the plant
membrane fluidity, resulting in lower membrane permeability at low temperatures and lower metal uptake
(Marschner, 1995). In the aquatic environment, the Cu
adsorption to the alga Dunaliella tertilecta increases with
increasing temperature (Gonzalez-Davila et al., 1995).
Further, in the lichen Peltigera horizontalis (Huds.)
(Beckett and Brown, 1984) and the liverwort Dumortiera
hirsuta (Sw.) (Mautsoe and Beckett, 1996), both intracellular and extracellular uptake of Cd was stimulated
by an increase in temperature. Similarly, several studies
of terrestrial plants grown at high root temperatures
found higher uptakes of Zn, Pb, Ag, Cr, Sb, and Cd
than was the case with plants grown at low root
temperatures (Hooda and Alloway, 1993; Macek et al.,
1994; Baghour et al., 2001; Albrecht et al., 2002).
Therefore, a general increase of metal uptake with
increasing temperature seems likely.
The salinity of stormwater may affect the plant
growth rate and plant metal uptake through the toxic
effects of both the NaC and Clÿ ions. NaC ions may
release Cd from the sediment to the water, thereby
increasing the Cd concentration in the water (Greger
et al., 1995). In seawater, chloride commonly forms
complexes with Zn, Cd, and, to a lesser extent, Cu, but
not with Pb, and thus the free ion concentration of the
former metals will be reduced (Förstner, 1979; Williams
et al., 1994). Fewer free Cd ions in water of higher
salinity correlates with a lower uptake of Cd by
Potamogeton pectinatus (L.) growing in higher salinity
water (Greger et al., 1995). At a higher salinity, the
increased NaC concentration reduced both intracellular
and extracellular uptake of Cd in the free-floating plant,
Lemna polyrhiza (L.) (Noraho and Gaur, 1995). Not
only in submersed (Greger et al., 1995) and free-floating
(Noraho and Gaur, 1995) plants, but also in macro-
algae, such as the seaweed Fucus vesiculosus (L.), did the
uptake of metal (Zn) decrease with increasing salinity
(Munda and Hudnik, 1988). Thus, a general decrease in
metal uptake with increasing salinity could be expected.
Variations in the temperature and salinity of stormwater result in different combinations of these two
factors over the course of a year (Wittman, 1979;
Sandersson, 1997). In winter/early spring, low temperature is combined with low to high (5&) salinity, while
in summer, high or medium temperatures are combined
with low to medium (0.5&) salinity. Furthermore, in
late autumn/early winter both temperature and salinity
may be low. To the best of our knowledge, no
investigation has as yet determined the effects of
temperature in combination with salinity on the heavy
metal uptake of submersed plants. This study evaluates
the combined effects of temperature and salinity on the
uptake of a set of heavy metalsdCu, Zn, Cd, and Pbd
in two morphologically different submersed aquatic
plant species, Elodea canadensis and Potamogeton
natans. In a previous study (Fritioff and Greger, 2003),
these two species were shown to accumulate higher
concentrations of heavy metals when grown in stormwater ponds than were emergent species sampled.
Although these are known as freshwater species, they
also grow in somewhat brackish water (Luther, 1951).
Furthermore, both species grow in ponds and wetlands
until an ice layer covers the water surface. Our
hypotheses were that metal concentrations and accumulations in submersed plant species would: (1) independent of salinity, increase with increasing water
temperature and (2) independent of temperature, decrease with increasing salinity.
2. Materials and methods
2.1. Plant material
The two studied species commonly grow in stormwater treatment ponds and wetlands. Although both are
submersed, E. canadensis (L.) and P. natans (Michx.)
have quite different morphologies. Elodea has very thin
submersed leaves, almost lacking a cuticle, arranged
around the stem, giving a high surface-to-volume ratio.
In contrast, Potamogeton has long petioles and leaves
that are several cell layers thick floating on the water
surface. These leaves are covered with a comparatively
thick cuticle, resulting in a much lower surface-tovolume ratio. Both studied species grow and remain
green until an ice layer covers the wetland (during cold
winters), causing a die back. Below the ice, Potamogeton
over-winters by means of its hardy rootstocks, while
Elodea over-winters by means of short vegetative shoots.
Whole plants of Potamogeton were collected
in August 2001 from an unpolluted pond called
Å. Fritioff et al. / Environmental Pollution 133 (2005) 265–274
Skåbydammen, located south of Stockholm, Sweden.
Plants were gently cleaned with tap water to remove the
sediment, planted in a greenhouse pond containing tap
water and minimally contaminated sediment, and left
growing for 4 weeks until the start of the experiment.
For Elodea, 10-cm-long vegetative shoots were collected
from a greenhouse pond. The shoots were planted in
a new pond containing the same types of sediment and
water as did the pond containing the Potamogeton
plants, and grown for about 5 weeks until new
vegetative shoots had developed.
2.2. Growth conditions
The two species were grown in a greenhouse equipped
with supplementary lamps. During the pre-experimental
phase, Osram daylight lamps (HQI-BT 400W) were used
for a light period of 12 h. Temperature in the pond
water was 18 G 1 C. The acclimatization and experimental phases were carried out in a climate-controlled
chamber with an air temperature of 10 G 1 C. Halogen
lamps (Osram Poverstar HQI-E) were used to create
light intensity of 50 G 5 mmol photons mÿ2 sÿ1 for a light
period of 18 h.
2.3. Experiment
To study the effects of salinity and water temperature
on the heavy metal uptake of Potamogeton and Elodea,
a two-factorial design, 32 (Sokal and Rohlf, 1995), was
set up with temperature and salinity as the two factors.
Each Potamogeton plant (with 1–5 leaves) was thoroughly cleaned under running tap water to remove
sediment and other particles and then rinsed in
redistilled water before being put in a container
containing 2 L of 2% Hoagland medium (Eliasson,
1978) with a pH of 5.0 G 0.2; the medium was not
aerated. In the experiment with Elodea, four vegetative
shoots (one each of 10, 15, 20, and 25 cm in length) were
cut and placed together in each container, a single
container comprising one replicate. Seven replicates
were used for each treatment and each species. Containers with plants were randomly placed in large, waterfilled tubs. Water in one-third of the tubs was chilled to
5 C using a compressor (TECO RA 680, Italy),
thermostats (Jäger, Type LZRH, Wüstenrot) to keep it
at 20 C, heated the water of another third of the tubs
and the water of the remaining third of the tubs was
kept at 11 C. To reduce evaporation, all containers
were covered with transparent plastic film. After 10 days
of acclimatization, the fresh weights (FW) of the
individuals of both species were measured after first
gently shaking the plants for 10 s to remove water.
Additionally, the lengths of the Elodea plants were
measured. Plants were then again set in containers of
fresh medium kept at the same temperatures as before.
267
After another day of acclimatization, plants from three
containers at each temperature were used to determine
the baseline heavy metal content in the plants. Thereafter, heavy metals and NaCl were added to all
containers. To simulate a storm event, when a flush of
highly polluted water originating from rainfall enters the
pond, the plants were treated for 48 h with metal levels
of 1.5, 20, 1, and 4 mM (equal to 95, 1307, 112, and
836 mg Lÿ1) of Cu, Zn, Cd, and Pb, respectively
(Morrison, 1989; Sansalone et al., 1996; Boller, 1997;
Pettersson et al., 1999). Then, NaCl was added, which
resulted in final salinity levels of 0, 0.5, and 5&. After
48 h, the experiment was ended and the fresh weights of
plants were measured as well as the length of the Elodea
plants. The pH remained at 5.0 G 0.1 for the duration of
the experiment.
All plants of both species were rinsed twice in
redistilled water, once in 20 mM EDTA, and twice in
redistilled water, for half a minute each time. They
were dried at 105 C for 24 h and then weighed to
determine the plant dry weights (DW). All plant
materials from each container was wet digested in
concentrated HNO3:HClO4 (7:3, v:v). Metal concentration was measured using a flame atomic absorption
spectrophotometer (SpectrAA-100, Varian, Springvalve, Australia). Standards were added to the samples
to eliminate the interaction of the sample matrix.
Lagharosiphon major was used as a reference material
(BCR No 60 N 675, Commission of the European
communities).
2.4. Calculations and statistics
Dry weight-to-fresh weight ratios (DW/FW) were
calculated for both species. To determine the growth
rate, the increase in fresh weight of Potamogeton and the
increase in length of Elodea during the 48-h experimental period were calculated as g FW (24 h)ÿ1 and cm
(24 h)ÿ1, respectively. Metal accumulations during the
experiment were calculated by subtracting control
concentrations from the concentrations found in the
treated plants.
The effects of the fixed factors, temperature and
salinity, on Zn, Cu, Cd, and Pb concentrations in
Potamogeton and Elodea were analyzed in a general
linear model which used biomass ( g DW Lÿ1) as
a covariable, in order to sort out the effects of the
various factors on the metal concentrations in the two
plant species. Insignificant interactions with the continuous covariable were dropped from the model using
a backward elimination procedure. Analysis of the metal
accumulations was preformed in the same way. Tukey’s
honest significant difference was used as a post hoc test
(Statistica software, StatSoft Inc., ’99 edition, kernel
release 5.5 A).
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Å. Fritioff et al. / Environmental Pollution 133 (2005) 265–274
3. Results
Growth rate, measured as increase in length
(cm ! 24 hÿ1), increased significantly with increasing
temperature for Elodea (Table 1). Furthermore, Elodea
plants grew significantly longer (cm) in containers kept
at 20 C than in containers kept at colder temperatures.
The DW/FW ratio (%) was significantly higher in
Elodea subject to colder rather than warmer treatments.
The dry and fresh weights of Elodea per liter did not
differ significantly among treatments. For Potamogeton,
no significant differences in dry or fresh weights, DW/
FW ratios, or growth rates ( g FW ! 24 hÿ1) were found
among treatments. The plant fresh weights were slightly
higher for Potamogeton than for Elodea, while the plant
dry weights and DW/FW ratios were more than twice as
high for Potamogeton as for Elodea.
Because of the higher biomass (DW lÿ1) of Potamogeton than of Elodea (Table 1), total metal accumulations (mg containerÿ1) in the plant material of
Potamogeton in some cases exceeded those of Elodea
(Fig. 1). Thus, compared with Elodea, Potamogeton
removed more of the added Zn from the medium in all
treatments and more Cu and Cd from the medium in
the 5 C treatment. However, the metals added to the
nutrient solution were never depleted in any of the
treatments.
The metal concentrations in the plants at the end of
the experiment are shown in Table 2. Elodea had, with
a few exceptions in the case of Zn, higher concentrations
of heavy metals than did Potamogeton in both treated
and control plants. Except for Cu in Elodea at 5 C and
5& salinity, controls had lower metal concentrations
than did the treated plants. Concentrations of each
metal in the control plants did not differ significantly
among temperature treatments.
The metal concentrations in plants increased with
increasing temperature in most cases (Fig. 2, Tables 3 and
4). This was found to be significant in Elodea for Cu, Zn,
and Cd and in Potamogeton for Cu, Cd, and Pb, and
a tendency was found in Potamogeton for Zn. Furthermore, the concentrations of Cu, Zn, and Cd increased
with decreasing salinity: this was significant in both
species for Zn and Cd and in Elodea for Cu, and a
tendency was found in Potamogeton for Cu. Lead concentration, however, was unaffected by salinity. Analysis
of metal accumulations (not shown) revealed the same
patterns as were found for metal concentrations.
Biomass affected the metal uptake in plants, smaller
plants having higher metal concentrations and accumulations than larger plants, significantly so for Cu and Zn
in Elodea and for all the metals measured in Potamogeton (Fig. 3, Tables 3 and 4).
Several interactions were seen. Temperature and
salinity interacted both with each other and with the
biomass. Between temperature and salinity, two interactions were found (Fig. 4, Tables 3 and 4). First,
Potamogeton plants growing in medium salinity water
(0.5&) accumulated more Zn than did plants growing in
high salinity water (5&) at 5 and 20 C but not at 11 C.
Second, the temperature effect on Pb concentration in
Potamogeton was significant only when plants were
growing in medium salinity water (0.5&).
The different temperatures had stronger effects on the
metal concentrations and accumulations in plants with
lower biomass rather than in plants with higher biomass,
Table 1
Treatment averages of dry and fresh weights, DW/FW ratio (dry weight/fresh weight ratio), length, and growth rate for Elodea and Potamogeton
treated for 48 h with metal levels of 1.5 mM Cu, 20 mM Zn, 1 mM Cd, and 4 mM Pb, temperatures of 5, 11, or 20 C, and salinities of 0, 0.5, or 5&
Species
Treatment
Elodea
Potamogeton
Dry weight
( g DW Lÿ1)
Fresh weight
( g FW Lÿ1)
DW/FW ratio
(%)
Total length
(cm)
Growth rate
(cm (24 h)ÿ1)
Growth rate
( g FW (24 h)ÿ1)
Temperature ( C)
5
11
20
0.43 G 0.01 a
0.43 G 0.00 a
0.44 G 0.01 a
8.1 G 0.08 a
8.7 G 0.06 a
9.0 G 0.08 a
5.2 G 0.03 a
5.0 G 0.02 b
4.8 G 0.02 b
76.76 G 0.106 b
75.10 G 0.059 b
83.67 G 0.251 a
0.23 G 0.01 b
0.32 G 0.01 ab
0.43 G 0.01 a
n.d.
n.d.
n.d.
Salinity (&)
0
0.5
5
0.45 G 0.01 a
0.44 G 0.01 a
0.41 G 0.00 a
8.7 G 0.08 a
8.7 G 0.08 a
8.3 G 0.06 a
5.1 G 0.03 a
5.0 G 0.02 a
4.9 G 0.02 a
78.50 G 0.26 a
78.82 G 0.286 a
78.21 G 0.16 a
0.31 G 0.01 a
0.32 G 0.01 a
0.35 G 0.01 a
n.d.
n.d.
n.d.
Temperature ( C)
5
11
20
1.32 G 0.03 a
1.49 G 0.04 a
1.11 G 0.03 a
11.1 G 0.23 a
12.4 G 0.26 a
10.3 G 0.27 a
11.8 G 0.1 a
11.8 G 0.1 a
10.5 G 0.1 a
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
0.00 G 0.01 a
0.12 G 0.02 a
0.02 G 0.01 a
Salinity (&)
0
0.5
5
1.35 G 0.04 a
1.39 G 0.03 a
1.19 G 0.03 a
11.7 G 0.28 a
11.7 G 0.24 a
10.3 G 0.25 a
11.2 G 0.1 a
11.7 G 0.1 a
11.2 G 0.1 a
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
0.03 G 0.02 a
0.00 G 0.00 a
0.06 G 0.01 a
Different letters indicate significant differences within each metal and treatment (n = 21) GSE. n.d. = no data.
269
Å. Fritioff et al. / Environmental Pollution 133 (2005) 265–274
µg Cd accumulated
100
50
0
0 0.5 5
Salinity (‰)
Temperature (°C) 5
µg Zn accumulated
150
Cu
0 0.5 5
0 0.5 5
11
20
1200
800
400
0
0 0.5 5
Salinity (‰)
Temperature (°C) 5
100
50
0 0.5 5
5
Zn
1600
Cd
0
µg Pb accumulated
µg Cu accumulated
150
0 0.5 5
0 0.5 5
11
20
0 0.5 5
11
0 0.5 5
20
Pb
1000
800
600
400
200
0
0 0.5 5
5
0 0.5 5
11
0 0.5 5
20
Fig. 1. Total accumulation of Cu, Zn, Cd, and Pb in mg containerÿ1 in Elodea (black bar) and Potamogeton (white bar), treated for 48 h with metal
levels of 1.5 mM Cu, 20 mM Zn, 1 mM Cd, and 4 mM Pb, temperatures of 5, 11, or 20 C, and salinity of 0, 0.5, or 5&. Each container contained 2 L
of medium and on average 0.43 and 1.31 g of Elodea and Potamogeton, respectively (n = 7).
this being the case for Cu in both plant species and for Cd
and Pb in Potamogeton (Tables 3 and 4). Biomass also
interacted with salinity, and the concentration and
accumulation of Cu in Elodea and Zn in Potamogeton
were more affected by differences in salinity at lower
rather than higher biomasses. Furthermore, the Cd concentration and accumulation in Elodea increased with
increasing biomass when grown at 11 C, while at the
other temperatures Cd concentrations decreased with
increasing biomass (Table 3).
Table 2
Concentrations of Cu, Zn, Cd, and Pb in Elodea and Potamogeton both before (Controls) and after treatment for 48 h with 1.5 mM Cu, 20 mM Zn,
1 mM Cd, and 4 mM Pb, temperatures of 5, 11, or 20 C, and salinities of 0, 0.5, or 5& (n = 7, for controls n = 2–3, GSE)
Metal accumulation (mg( g DW)ÿ1)
Species
Temperature ( C)
Salinity (&)
Cu
Zn
Cd
Pb
Elodea
5
0
0.5
5
Control
340 G 15
311 G 24
246 G 7
271 G 81
1165 G 41
1137 G 114
548 G 37
387 G 91
145 G 11
117 G 9
38 G 4
3G2
1732 G 289
1838 G 336
1495 G 184
145 G 55
11
0
0.5
5
Control
345 G 10
354 G 15
302 G 15
116 G 12
1168 G 65
1128 G 50
626 G 62
208 G 62
259 G 43
181 G 27
85 G 19
3G1
1447 G 208
1528 G 300
2037 G 225
16 G 7
20
0
0.5
5
Control
444 G 15
406 G 30
348 G 25
154 G 19
2193 G 118
1919 G 138
1354 G 276
178 G 21
269 G 16
244 G 24
212 G 23
1G1
1771 G 152
1694 G 134
2190 G 168
53 G 33
5
0
0.5
5
Control
145 G 22
118 G 13
137 G 25
23 G 0
1381 G 122
1258 G 264
560 G 64
48 G 6
94 G 12
70 G 8
32 G 4
5G1
127 G 40
74 G 19
110 G 20
5G5
11
0
0.5
5
Control
133 G 19
111 G 22
70 G 10
17 G 3
1297 G 132
873 G 139
672 G 74
69 G 9
108 G 12
74 G 13
32 G 3
3G2
88 G 17
75 G 9
113 G 39
7G3
20
0
0.5
5
Control
193 G 60
160 G 40
142 G 26
21 G 5
1679 G 255
1311 G 194
693 G 84
61 G 6
151 G 33
100 G 18
63 G 27
2G1
151 G 40
227 G 68
162 G 42
7G0
Potamogeton
270
Å. Fritioff et al. / Environmental Pollution 133 (2005) 265–274
µg Me / g DW
Cd
Cu
400
Cu
Cd
300
200
Table 3
Analysis of variance of heavy metal concentrations in Elodea exposed
to Cu, Zn, Cd, and Pb, three different salinities, and three different
water temperatures
Response
variable
Source of
variation
df
MS
F
p
Cu
Temperature
Salinity
Biomass
Temperature
! Salinity
Temperature
! Biomass
Salinity
! Biomass
Error
2
2
1
4
17784
9548
54589
1034
15.7
8.4
48.1
0.9
!0.001***
!0.001***
!0.001***
0.4655
2
6564
5.8
0.0056**
2
4143
3.6
0.0334*
49
1136
Temperature
Salinity
Biomass
Temperature
! Salinity
Error
2
2
1
4
5282023
2357154
462852
40936
53.4
23.8
4.7
0.4
!0.001***
!0.001***
0.0351*
0.7981
53
98979
Temperature
Salinity
Biomass
Temperature
! Salinity
Temperature
! Biomass
Error
2
2
1
4
33081
57252
1139
6665
11.7
20.2
0.4
2.4
!0.001***
!0.001***
0.5288
0.0663
2
22256
7.9
0.0011**
51
2833
Temperature
Salinity
Biomass
Temperature
! Salinity
Error
2
2
1
4
343788
584252
1355437
314637
1.0
1.6
3.8
0.9
0.3889
0.2049
0.0569
0.4823
53
357601
100
0
5
11
20
5
11
20
0
0.5
5
0
0.5
5
2500
Pb
Zn
Pb
Zn
Zn
µg Me / g DW
2000
1500
1000
Cd
500
0
5
11 20
5
11
Temperature (°C)
20
0
0.5
5
0
0.5
5
Salinity (‰)
Fig. 2. Treatment averages of Cu, Zn, Cd, and Pb concentrations in
Elodea (black bar) and Potamogeton (white bar) treated for 48 h with
metal levels of 1.5 mM Cu, 20 mM Zn, 1 mM Cd, and 4 mM Pb, temperatures of 5, 11, or 20 C, and salinities of 0, 0.5, or 5& (n = 21) GSE.
4. Discussion
This research found that metal concentration and
accumulation in the two studied submersed macrophyte
species generally increased with increasing temperature
but decreased with increasing salinity (Fig. 2, Tables 3
and 4). The influences of temperature and salinity were
not always independent of each other (Fig. 4, Tables 3
and 4). Furthermore, metal concentrations and accumulations increased with decreasing biomass in most
cases (Fig. 3, Tables 3 and 4). Interactions between
temperature and biomass or salinity and biomass were
sometimes found as well (Tables 3 and 4).
Because of the higher biomass (DW lÿ1) of Potamogeton than of Elodea (Table 1), total metal accumulations (mg containerÿ1) in Potamogeton plant material
in some cases exceeded those of Elodea (Fig. 1). Thus,
compared with Elodea, Potamogeton removed more
added Zn from the medium in all treatments and more
Cu and Cd from the medium in the 5 C treatment.
In line with the first hypothesis, metal concentrations
and accumulations increased with increasing water
temperature for Cu, Zn, and Cd in Elodea (Fig. 2, Table
3) and for Cu, Zn, Cd, and Pb in Potamogeton (Fig. 2,
Table 4), independent of salinity. Furthermore, total
Pb
Plants were allowed to acclimatize to the temperatures for 10 days before
exposure to salinity treatments and heavy metals. Only single factors
(temperature and salinity), the covariable (biomass, as dry weight),
interactions between main factors (temperature and salinity), and significant interactions are shown. Error, within group variance; df, degrees
of freedom; MS, between group variance; F, ratio between MS and
Error.
*p ! 0.05, **p ! 0.01, ***p ! 0.001.
metal accumulations (mg containerÿ1) increased with
increasing temperature (Fig. 1). One likely explanation
for this is that increased temperature will increase the
biomass and thus the absorption area of the plant; this, in
turn, will increase the metal uptake. This was found to be
the case for Cd uptake by Pinus silvestris (L.) (Ekvall and
Greger, 2003). In the present study, Elodea length
increased significantly more in containers at 20 C than
in those subject to colder treatments (Table 1). However,
biomass did not differ between treatments; thus the effect
of temperature on metal uptake was direct, and an
explanation other than growth rate must be found.
Earlier investigations suggested that the extracellular
concentration increases with increasing temperature
because there is a change in the equilibrium between
the cell wall exchange sites and the metal in solution
Å. Fritioff et al. / Environmental Pollution 133 (2005) 265–274
Table 4
Analysis of variance of heavy metal concentrations in Potamogeton
exposed to Cu, Zn, Cd, and Pb, three different salinities, and three
different water temperatures
Response
variable
Source of
variation
df
MS
F
p
Cu
Temperature
Salinity
Biomass
Temperature
! Salinity
Temperature
! Biomass
Error
2
2
1
4
16893
6306
182345
1894
6.7
2.5
72.1
0.7
0.0026**
0.0926
!0.001***
0.5632
2
13555
5.4
0.0077**
51
2528
Temperature
Salinity
Biomass
Temperature
! Salinity
Salinity
! Biomass
Error
2
2
1
4
204246
1888943
4417721
216076
2.6
23.6
55.3
2.7
0.0876
!0.001***
!0.001***
0.0406*
2
510731
6.4
0.0033**
51
79944
Temperature
Salinity
Biomass
Temperature
! Salinity
Temperature
! Biomass
Error
2
2
1
4
11998
25012
42135
1033
11.3
23.6
39.7
1.0
!0.001***
!0.001***
!0.001***
0.4305
2
7696
7.2
0.0017**
51
1062
Temperature
Salinity
Biomass
Temperature
! Salinity
Temperature
! Biomass
Error
2
2
1
4
36547
1807
191946
16666
6.8
0.3
35.8
3.1
0.0024**
0.7152
!0.001***
0.0229*
2
19125
3.6
0.0354*
51
5355
Zn
Cd
Pb
Plants were allowed to acclimatize to the temperatures for 10 days
before exposure to salinity treatments and heavy metals. Only single
factors (temperature and salinity), the covariable (biomass, as dry
weight), interactions between main factors (temperature and salinity),
and significant interactions are shown. Error, within group variance;
df, degrees of freedom; MS, between group variance; F, ratio between
MS and Error.
*p ! 0.05, **p ! 0.01, ***p ! 0.001.
(Gonzalez-Davila et al., 1995; Mautsoe and Beckett,
1996). However, if this was the only factor, temperature
should affect Pb accumulation in Elodea in the same way
it affects accumulation of the other three metals, which
was not the case (Fig. 2, Table 3). Instead, it is suggested
that the absence of a temperature effect on Pb accumulation is due to a high proportion of extracellular binding
sites for Pb in Elodea. This was earlier shown by Vásquez
et al. (1999) to be the case in two aquatic bryophyte
species, Fontinalis antipyretica Hedw. and Scapania
undulata (L. Dum.). Extracellular binding of metal is
less temperature dependent than is intracellular uptake,
according to Beckett and Brown (1984), Marschner
(1995), and Mautsoe and Beckett (1996). The intracel-
271
lular uptake of Cu, Cd, and Zn might also be the fractions most affected by temperature in the present study.
Slightly increased extracellular uptake could facilitate
intracellular uptake by concentrating metals close to the
membrane. Furthermore, a higher temperature changes
the lipid composition of the plasma membrane (Lynch
and Steponkus, 1987) and thereby its fluidity, which may
facilitate both passive and active metal flux through the
membrane. In addition, increased temperature increases
both metabolism (Marschner, 1995; Nilsen and Orcutt,
1996) and protein synthesis (Nilsen and Orcutt, 1996),
and this may result in higher metal uptake at additional
uptake sites on membranes or an increased release of
molecules facilitating metal uptake.
The second hypothesis, that metal concentration and
accumulation decrease with increasing salinity, was
supported by the results for Cu, Zn, and Cd in both
species (Fig. 2, Tables 3 and 4), independent of
temperature. E. canadensis and P. natans are freshwater
species and have been found growing in the field in
salinities up to 3& and 2&, respectively (Luther, 1951).
Even at the highest salinity used in this study, 5&, no
toxic effect was observed in these species (not shown).
The likely reason for the decreased accumulation with
increasing salinity is the decreased availability of the
metals in the growth medium because of the complexes
formed between chloride and metals (Förstner, 1979).
This has, for example, been shown to depress Cd uptake
in P. pectinatus (Greger et al., 1995). In addition,
increased competition with sodium ions at uptake sites,
both on the plasma membrane and in apparent free
space in the cell walls, may account for the decreased
metal accumulation at higher salinities (Noraho and
Gaur, 1995). That salinity has no influence on Pb
accumulation (Fig. 2, Tables 3 and 4) may be because Pb
does not form a complex with chloride (Förstner, 1979),
is retained briefly in water, and has a high affinity to
bind to organic matter (Förstner, 1979) such as plant
surfaces (Vásquez et al., 1999). Therefore, despite
different salinities, the same amount of Pb will bind to
and be taken up by plants.
Biomass did not differ among treatments, but rather
varied within each treatment (Table 1), and plants with
lower biomasses had higher metal accumulations and
concentrations than did plants with higher biomasses
(Fig. 3, Tables 3 and 4). This may be because a proportionally larger absorption area in relation to the external
metal concentration may result in a dilution effect and
thus a low internal metal concentration. Ekvall and
Greger (2003) showed that this was the case for Cd in
Pinus sylvestris. Besides different amounts of biomass,
different metal accumulations may also be due to
different proportions of stems to leaves. Preliminary
results for P. natans indicate that the uptake by its leaves
is almost 10 times higher than the uptake by its stems
(Fritioff and Greger, unpublished).
272
A
2000
1000
0
0.5
0.75
Elodea biomass (g DW)
0
2
1
3
Biomass (g DW)
200
0
0.25
1000
400
Biomass (g DW)
200
2000
0
0.25 0.5 0.75
(µg Me / g DW)
(µg Me / g DW)
B
0
0
400
0
(µg Me / g DW)
(µg Me / g DW)
Å. Fritioff et al. / Environmental Pollution 133 (2005) 265–274
0
1
2
3
Potamogeton biomass (g DW)
Fig. 3. The effect of biomass on Cu, Zn, Cd, and Pb concentrations in Elodea (A) and Potamogeton (B) treated for 48 h with metal levels of 1.5 mM
Cu, 20 mM Zn, 1 mM Cd, and 4 mM Pb, temperatures of 5, 11, or 20 C, and salinities of 0, 0.5, or 5&. Only trend lines are shown; the line pattern
indicates which metal is represented: Cu (d – d – d –), Zn (––– ––– ––– –––), Cd (– – – – – –), Pb (ddd). n = 21.
The concentrations of the various metals tested were,
in almost all cases, higher in Elodea than in Potamogeton
(Fig. 2). The Zn concentration was about the same in the
two species, while Cu and Cd concentrations were about
twice as high in Elodea than in Potamogeton and the Pb
concentration was over 10 times higher in Elodea. This is
5 ºC
11 ºC
20 ºC
A
µg Pb / g DW
300
250
200
150
100
50
0
0
0.5
5
Salinity (‰)
B
0‰
0.5 ‰
5‰
µg Zn / g DW
2000
1500
1000
500
0
5
11
20
Temperature (°C )
Fig. 4. Concentrations (mg ! g ÿ1 ! DWÿ1) of Pb and Zn in
Potamogeton (A and B, respectively), showing interactions between
temperature and salinity after treatment for 48 h with 1.5 mM Cu,
20 mM Zn, 1 mM Cd, and 4 mM Pb, temperatures of 5, 11, or 20 C,
and salinity of 0, 0.5, or 5& (n = 7) GSE.
in accordance with other studies done both in laboratory-scale wetlands (Nyquist and Greger, 2003) and in
the field (Yurukova and Kochev, 1994). First, Potamogeton plants had higher biomasses than did the Elodea
plants (Table 1); as previously discussed, within
a species, plants with higher biomass will have lower
metal concentrations when exposed to the same metal
concentration in the medium. Second, Elodea had
a higher water content than did Potamogeton, as shown
by low DW:FW ratio (Table 1). This may facilitate
metal uptake, by presenting larger vacuoles for storage
and because a higher water content in the cell wall
facilitates diffusion into the apparent free space by
dilution. Other reasons for the higher metal accumulation in Elodea than in Potamogeton (Fig. 2) may be
species differences in uptake capacities and differences in
the proportion of stems to leaves. Sculthorpe (1967)
described how Elodea had a higher leaf uptake of solutes
than did Potamogeton, because of their differences in
morphology. Elodea’s high surface-to-volume ratio as
well as its high proportion of leaves relative to total
plant biomass may facilitate metal uptake by diffusion,
while the thick waxy cuticle of Potamogeton may
obstruct metal uptake.
Submersed plants often display high metal accumulations in the field (Galiulin et al., 2001). In the present
laboratory experiment, Elodea had in most cases higher
metal concentrations than did Potamogeton. In some
cases, however, Potamogeton removed more of the
added metals in total from the containers than did
Elodea. Depending on the treatment, Elodea removed at
most 70, 30, 51, and 56% and at least 13, 5, 8, and 40%
of the added Cu, Zn, Cd, and Pb, respectively, during
the 48-h treatment with elevated levels of metals.
Å. Fritioff et al. / Environmental Pollution 133 (2005) 265–274
Potamogeton removed at most 75, 64, 50, and 11% and
at least 50, 22, 14, and 5% of the added Cu, Zn, Cd, and
Pb, respectively, during the same 48-h treatment. The
average weight of plant material in each container was
0.43 g DW for Elodea and 1.3 g DW for Potamogeton
(Table 1). Thus, the high removal might depend on
a high plant biomass per volume of solution. Both metal
concentration in plant material and total uptake were
highly dependent on plant biomass; therefore, biomass
in combination with metal load was important for
overall metal removal during the experiment. In fact,
Potamogeton removed at least twice as much Zn as did
Elodea in this experiment. However, Nyquist and
Greger (2003) found the opposite in a laboratory-scale
wetland experiment. They reported that Elodea removed
64% of the total amount of added Zn, while Potamogeton removed just 23%. The present study found that
Elodea and Potamogeton did accumulate heavy metals,
although the quantity depended on the biomass, indicating that Elodea and Potamogeton can both remove
metals from stormwater during storm events.
5. Conclusions
Although Elodea and Potamogeton are most effective
at accumulating heavy metals discharged in stormwater
at high temperatures, they have still good heavy metal
accumulation capacity down to at least 5 C. They may
therefore be useful in treating stormwater in temperate
climates. Since plant biomass is low in early spring,
metal accumulation per unit of biomass can be expected
to be comparatively high, because of generally higher
metal accumulation when a small, compared to a large,
biomass is exposed to a given amount of metal in the
water. Increased salinity resulting from road deicing will
reduce metal accumulation in both Elodea and Potamogeton more than temperature will. Low salinity levels in
the summer lower Cu, Zn, and Cd accumulation by
plants, while salinity peaks in the winter may reduce the
accumulations by plants by over 50%. Unlike the other
metals, Pb accumulation is generally unaffected or very
little affected by temperature and/or salinity, and will
thus remain relatively stable throughout all seasons and
possible salinity levels. Although during the experiment
Potamogeton accumulated more Zn and sometimes
more Cu and Cd from the medium than did Elodea,
Potamogeton more often had a lower metal concentration than did Elodea; this is possibly because Potamogeton had a higher biomass per liter than did Elodea.
Based on our results, we suggest that since in the field
Elodea often occurs in more dense stands, grows faster,
and often has higher metal concentrations than does
Potamogeton, it is the better of the two tested species for
use in stormwater treatment facilities. However, further
in situ investigations are needed, and studies of the
273
storage capacity of heavy metals and the fate of metals
accumulated in biomass decomposing over the winter
merit further investigation.
Acknowledgements
We gratefully thank the Swedish Council for Forestry
and Agricultural Research for financing this study.
Thanks are also due to Stefan Dahlgren for collecting P.
natans and to Patrik Dinnetz for help with the statistical
calculations.
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