Journal of Experimental Botany, Vol. 57, No. 11, pp. 2661–2671, 2006
doi:10.1093/jxb/erl029 Advance Access publication 4 July, 2006
RESEARCH PAPER
The response of nutrient assimilation and biochemical
composition of Arctic seaweeds to a nutrient input in
summer
Francisco J. L. Gordillo*, José Aguilera and Carlos Jiménez
Departamento de Ecologı´a, Facultad de Ciencias, Universidad de Málaga, Campus Teatinos, E-29071 Málaga, Spain
Received 18 February 2006; Accepted 17 April 2006
Abstract
Twenty-one species of macroalgae (four Chlorophyta,
eight Rhodophyta, and nine Phaeophyta) from the
Kongsfjord (Norwegian Arctic) were examined for their
response to nutrient enrichment (nitrate and phosphate) in the summer period. The enzymatic activities
related to nutrient assimilation, external carbonic anhydrase (CAext, EC 4.2.1.1), nitrate reductase (NR, EC
1.6.6.1), and alkaline phosphatase (AP, EC 3.1.3.1), as
well as the biochemical composition (total C and N,
soluble carbohydrates, soluble proteins, and pigments)
were measured. CAext activity was present in all
species, and showed a general decrease after nutrient
enrichment. Inversely, NR activity increased in most
of the species examined. Changes in pigment ratios
pointed to the implication of light harvesting system in the acclimation strategy. Despite enzymatic
and pigmentary response, the Arctic seaweeds can
be regarded as not being N-limited even in summer,
as shown by the slight effect of nutrient enrichment
on biochemical composition. The exception being the
nitrophilic species Monostroma arcticum and, to a
lesser extent, Acrosiphonia sp. For the rest of the
species studied, changes in total internal C and N,
soluble proteins, soluble carbohydrates, pigment content, and the internal pool of inorganic N were recorded
only for particular species and no general pattern
was shown. Acclimation to unexpected nutrient input
seemed to ensure the maintenance of a stable biomass
composition, rather than an optimized use of the newly
available resource (except for the nitrophilic species).
This indicates a high degree of resilience of the algal
community to a disruption in the natural nutrient
availability pattern.
Key words: Carbon, carbonic anhydrase, chlorophyll, eutrophication, nitrate reductase, nitrogen, seaweeds.
Introduction
The Arctic coastal environment is characterized by relatively constant and near-freezing water temperatures
(typically 2 to +4 8C) and strong seasonal variations in
light and nutrient availability (Hanelt et al., 2001; Hop
et al., 2002). The West Spitsbergen Current (WSC) has a
strong Atlantic character and brings relatively warm and
nutrient-rich waters to Kongsfjord (0 to +6 8C; Hanelt et al.,
2004), supporting a relatively rich macroalgal community
of around 50 species (Hop et al., 2002; Wiencke
et al., 2004). In Kongsfjord both boreal and Arctic species
are present, and their balance is influenced by oceanographic conditions (Atlantic versus Arctic) and glacial
inputs. The influx of Atlantic water and melting of glaciers
in this region has been linked to climate change (Svendsen
et al., 2002) and thus algal communities of Kongsfjord
function as a climate indicator at a local scale (Hop
et al., 2002).
Two of the most relevant global phenomena that
threaten ecosystems are the increase in atmospheric CO2
concentration (Ramanathan, 1998) and the increased load
of nitrogen in both the terrestrial and aquatic environment
(Galloway, 1998). Changing the availability of these nutrients
will force a change in the acclimation strategies of the
organisms. Seawater nitrate concentration in Kongsfjord
is generally highest in winter and spring (c. 10 lmol l1),
but it drops rapidly following the sea-ice break up due to
production of fast-growing microalgae, so that it becomes
depleted from early summer to winter. Soluble reactive
phosphorus (SRP) follows a similar pattern with maximum
* To whom correspondence should be addressed. E-mail: [email protected]
ª The Author [2006]. Published by Oxford University Press [on behalf of the Society for Experimental Biology]. All rights reserved.
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2662 Gordillo et al.
concentrations of c. 1 lmol l1 (Aguilera et al., 2002).
Nitrogen has long been recognized to limit macroalgal
productivity (Lapointe and O’Conell, 1989). Nutrient uptake and assimilation characteristics of macroalgae from
temperate waters have been shown to vary among populations and species in ways that optimize survival and
growth under local nutrient supply conditions (Wheeler
and Weidner, 1983; Hernández et al., 1993; Gordillo
et al., 2001a). Arctic species are also reported to show
nutritional strategies that allow them to cope with the long
periods of darkness in winter and nutrient depletion in
summer (Korb and Gerard, 2000a, b). Kelps living in the
Arctic rely on stored photosynthates accumulated during
the summer ice-free N-depleted period, supporting new
growth during ice-covered N-sufficient winter. Traditionally, it was thought that this seasonal growth pattern
was a direct consequence of N availability (Chapman
and Lindley, 1980) as well as light availability (Henley
and Dunton, 1997). However, it has become apparent
that, at least in some species, this pattern is under the
control of an endogenous free-running circannual rhythm
entrained by a critical minimum daylength in autumn
(Lüning, 1991; Schaffelke and Lüning, 1994), suggesting
that the addition of nitrate-N to summer N-limited kelps
would have only a marginal effect on growth and biochemical composition, presumably due to the prevailing
internal clock (Henley and Dunton, 1997). As far as it is
known this has never been tested at the community level
in the Arctic, and it is still unknown whether the limited
effect of N enrichment is only restricted to the few kelp
species studied, or is a common characteristic in Arctic
macroalgae.
Among the metabolic paths that are suspected to be
most influenced by changing the nutritional conditions
of the environment are the main nutrient assimilatory enzymes for CO2, nitrate and phosphate, as well as the lightharvesting strategy. External carbonic anhydrase (CAext)
and nitrate reductase (NR) are well known as key regulatory
enzymes for the C and N assimilation pathways, respectively. Carbonic anhydrases are involved in the conversion of HCO
3 to CO2, the only form of inorganic C used by
Rubisco (Raven, 1991), as part of the carbon-concentrating
mechanisms (CCMs). Many species shows CA activity
due to an enzyme in the plasma membrane, referred to as
external CA (CAext) independent from the internal CA
which is present also in algae lacking a CCM. CAext is
supressed by high CO2 levels, and enhanced by inorganic
nitrogen enrichment (Jiménez del Rı́o et al., 1995), so that
changes in CAext activity due to nutrient enrichment are
presumed to have a relevant (yet unknown) role in the
acclimation strategy.
Nitrate reductase is usually regarded as the limiting
step in nitrate assimilation, and is regulated by the external
levels of nitrate (among other factors, such as light). The
use of NR in an ecological context is particularly relevant
for marine environments where nitrogen is often limiting,
providing relevant information about the physiological
N-status of the organisms (Hernández et al., 1993). The
aim of this study was to provide an insight into whether
an occasional input of nutrient (nitrate and phosphate)
by Atlantic water during a period of external nutrient
depletion would onset acclimation strategies that would
lead to physiological disturbance, in representative species
of the Arctic macroalgal community.
Materials and methods
Plant material and experimental set-up
Twenty-one species of macroalgae were collected from the
Kongsfjord at Spitsbergen, Norwegian Arctic (78855‘ N, 11856’ E)
during July 2002 at depths of 2–6 m; four Chlorophyta, eight
Rhodophyta, and nine Phaeophyta (Table 1). The bathymetric
and habitat location for each species are described in Wiencke
et al. (2004) for most of the species studied. Visually healthy thalli,
free from macroscopic epibiota were selected and incubated in
2.0 l aquaria for 72 h immediately after collection. Seawater was
changed daily, and one set was enriched with 10 lM NO
3 and 1 lM
PO3
4 at the moment of water change (duplicate aquaria per set).
Incubations were performed at 5 8C in continuous white fluorescent
light of 30 lmol m2 s1. This irradiance was chosen according to
the average daily irradiance recorded at depths of 1–5 m for this
time of the year (Bischof et al., 2002). The aquaria were aerated at
0.5 l min1. The enzymatic methods described below were applied
to fresh material taken directly from the incubation aquaria. For
species with differentiated tissue, the most active part of the thallus was chosen. Biochemical composition was analysed from lyophilized material stored at 80 8C.
External carbonic anhydrase assay
The external carbonic anhydrase (CAext, EC 4.2.1.1) activity
was measured potentiometrically, according to Haglund et al.
(1992). The assay was carried out at 0–2 8C determining the time
taken for a linear drop in pH in the range 8.5 to 7.5 in a 3 ml volume
cuvette containing a buffer (50 mM TRIS, 25 mM ascorbic acid, and
5 mM EDTA). Small pieces of algae weighing 150–250 mg FW were
washed with distilled water and placed in the cuvette. The reaction
was started by adding 1 ml of ice-cold CO2-saturated distilled water.
One unit of relative enzymatic activity (REA) was defined as
(t0/tc)–1, where t0 and tc are the time taken for the pH change in the
absence and the presence of the alga, respectively. Typically, 5–6
independent replicates were assayed.
Nitrate reductase assay
Nitrate reductase (NR, EC 1.6.6.1) activity of fresh material was
measured following the in situ method according to Corzo and
Niell (1991), so that the enzyme is assayed in its original cellular
location. The assay medium contained a buffer (0.2 M H2 PO
4 and
1 mM EDTA; pH 8), a compound able to permeabilize the membrane (0.1% propanol v/v), nitrate in excess (50 mM NaNO3), and
a source of reducing power (10 lM glucose). The reaction was
carried out by placing 30–70 mg FW of alga in a test tube containing
2 ml of the assay medium. The incubation time was 30 min at
a temperature of 5 8C. The assay medium was bubbled with N2 gas
for 2 min before and 2 min after placing the algal sample in order
to remove the dissolved O2, that competes with NO
3 in its reduction
to NO
2 : The incubation was performed in darkness to prevent
+
further reduction of NO
2 to NH4 : The activity is measured as the
Nutrient assimilation in Arctic seaweeds
2663
Table 1. List of species from the Kongsfjord examined, and their fresh weight to dry weight ratio (FW:DW)
Phylum
Name
Chlorophyta
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
Rhodophyta
Phaeophyta
a
Acrosiphonia sp.
Chaetomorpha melagonium
Monostroma arcticum
Prasiola crispa
Ceramium strictum
Devaleraea ramentaceaa
Odonthalia dentata
Palmaria palmata
Phycodrys rubens
Polysiphonia arctica
Ptilota plumosa
Rhodomela lycopodioides
Alaria esculenta
Chorda tomentosa
Chordaria flagelliformis
Desmarestia aculeata
Fucus distichus
Laminaria saccharina
Laminaria solidungulaa
Scytosiphon lomentaria
Sphacelaria plumosa
Authority
FW:DW
C. Agardh
(F. Weber et Mohr) Kützing
Wittrock
(Lightfoot) Kützing
(Kützing) Harvey
(L.) Guiry
(L.) Lyngb.
(L.) Kuntze
(L.) Batt.
J. Agardh
(L.) C. Agardh
(L.) C. Agardh
(L.) Grev.
Lyngb.
(O.F.Muel.) C.Agardh
(L.) Lamour.
L.
(L.) L.
J. Agardh
(Lyngb.) Lynk
Lyngb.
4
7.3
3.4
2.6
6.1
6.3
4
6.6
4.1
5
3.6
6
4
13.5
7.2
5.3
4.8
6.2
6.5
13.7
4.5
Arctic endemic species.
rate of NO
2 produced. The NO2 in the assay medium was quantified spectrophotometrically according to Snell and Snell (1949).
The observed activity is a potential measure of the NRA of the cell
under the conditions prior to the assay. Typically, three independent
replicates were assayed.
Alkaline phosphatase assay
The alkaline phosphatase (AP, EC 3.1.3.1 ) activity (APA) was
assayed in 4–6 replicate samples of 50–100 mg FW using the
artificial substrate p-nitrophenol phosphate (pNPP). This compound
is cleaved by enzymatic hydrolysis rendering Pi and the coloured compound p-nitrophenyl (pNP) determined by absorbance at
410 nm. The procedure followed the method adapted for marine
macrophytes by Hernández and Whitton (1996). The assay medium
consisted of 2 ml of 500 lM pNPP (which enables maximum
velocity of the enzymatic reaction) and 500 lM HEPES-NaOH
buffer, pH 8.3, dissolved in P-free artificial seawater of 33 psu. The
samples were incubated for 30 min at room temperature. Enzymatic
activity was expressed as lmol pNP released g1 FW h1.
Carbon and nitrogen content
Total internal C and N were measured from lyophilized tissue
samples using a C:H:N elemental analyser (Perkin-Elmer 2400CHN).
Internal content of inorganic N (nitrate+nitrite) was estimated by
grinding to powder lyophilized material (3–4 mg), and adding 10 ml
deionized water. The extracts were incubated at 30 8C for 30 min,
and then filtered (Whatman GF/C). Nitrate was determined in the
aqueous fraction by using an automated autoanalyser (Traacs 800,
BranLuebbe) using the method provided by the manufacturer.
Biochemical composition
Soluble carbohydrates, soluble proteins, phycobiliproteins, chlorophyll a, chlorophyll b, and total carotenoids were determined from
material lyophilized right after collection from the incubation aquaria.
Soluble carbohydrates were estimated colorimetrically based on
Kochert (1978) using phenol–sulphuric acid for colour development
and glucose as standard.
Total soluble proteins from crude extracts (phosphate buffer,
pH 6.5) were determined using a commercial protein assay (Bio-Rad),
based on Bradford (1976). Protein content was determined spectrophotometrically at 595 nm and concentrations were calculated compared with a standard of bovine serum albumin. Phycoerythrin and
phycocyanin were determined from the same extracts using the
equations given by Beer and Eshel (1985) for spectrophotometrical
quantification. Chlorophyll a, chlorophyll b, and total carotenoids
were extracted in N, N dimethyl-formamide overnight (room temperature) after rehydrating lyophilized thalli. For spectrophotometric
quantification the equations given by Wellburn (1994) were used.
Tests performed in this laboratory demonstrated that the lyophilization process does not alter the absorbance characteristics of pigments,
so that determination was not significantly affected (t test, P <0.01).
Data analyses
Replicate measurements (n=2–6 independent thalli) were tested for
significance of differences (P <0.05) promoted by nutrient enrichment for each species using t tests. In addition, phylum-grouped
species were tested for significance of differences by two-way
ANOVAs followed by Tukey’s paired tests (P <0.05). All tests
were performed using the SigmaStat 2.03 statistical software
(SPSS Inc.).
All variables measured for the 21 species studied (Tables 2–5),
except chlorophyll b and phycoerythrin, were included in two principal component analyses (PCA), for control and enriched conditions, respectively. The PCA is one of the best-known ordination
methods. Mathematically, PCA consists of an eigenanalysis of a
covariance or correlation matrix calculated on the original measurement data. Graphically, it can be described as a rotation of a swarm
of data points in multidimensional space so that the longest axis
(the axis with the greatest variance) is the first PCA axis, the second
longest axis perpendicular to the first is the second PCA axis, and
so forth. Thus the first few PCA axes represent the greatest amount
of variation in the data set and usually contain some patterns of
significance (for clarity, only the first two axes for each analysis
is shown). Each new axis is a linear combination of the variables
measured, i.e. each variable contributes a certain amount (called
loading factor, Table 7) to the definition of each new axis. The
analyses were performed using the software MVSP 3.12 (Kovach
Computer Services).
2664 Gordillo et al.
Table 2. Activity of the assimilating enzymes external carbonic anhydrase (CAext, REA g1 FW), nitrate reductase (NRA, lmol NO
2
g1 FW h1), and alkaline phosphatase (APA, lmol PNP g1 FW h1)
Standard deviation in brackets (n=3–5); significant differences indicated by an asterisk (P <0.05).
Phylum
Chlorophyta
Rhodophyta
Phaeophyta
Name
CAext
Acrosiphonia sp.
Chaetomorpha melagonium
Monostroma arcticum
Prasiola crispa
Ceramium strictum
Devaleraea ramentacea
Odonthalia dentata
Palmaria palmata
Phycodrys rubens
Polysiphonia arctica
Ptilota plumosa
Rhodomela lycopodioides
Alaria esculenta
Chorda tomentosa
Chordaria flagelliformis
Desmarestia aculeata
Fucus distichus
Laminaria saccharina
Laminaria solidungula
Scytosiphon lomentaria
Sphacelaria plumosa
NRA
APA
Control
Enriched
Control
Enriched
Control
Enriched
17.2
9.7
33.4
59.0
15.4
3.6
11.9
3.9
24.9
11.6
14.0
17.2
26.8
32.0
14.7
18.8
8.4
23.6
21.8
14.4
25.1
12.3
15.8
32.2
46.6
16.4
4.0
7.4
3.1
13.8
7.3
10.8
6.4
14.5
27.6
13.7
7.8
8.8
9.5
8.0
15.3
24.7
2.67
5.79
5.16
8.30
3.54
1.16
8.48
1.48
2.19
2.42
4.18
2.19
2.34
1.53
1.03
2.27
1.22
2.20
3.44
1.10
2.45
3.81
3.97
11.95
8.94
4.95
2.92
10.14
2.70
1.57
5.37
3.92
3.14
11.01
0.74
3.35
1.97
4.53
2.35
11.94
1.66
3.53
0.39
0.29
0.47
0.49
1.45
1.14
2.76
0.75
2.79
2.69
2.34
2.67
1.30
0.22
0.31
0.22
1.35
0.15
0.14
0.53
2.45
2.57
0.29
0.32
0.89
1.88
0.79
2.06
0.59
2.33
1.69
1.59
2.58
0.57
0.43
0.34
0.43
0.74
0.21
0.08
0.46
3.20
(1.4)
(2.3)
(0.7)
(0.8)
(0.8)
(1.4)
(3.2)
(1.5)
(3.7)
(0.0)
(0.7)
(1.6)
(1.3)
(5.2)
(0.9)
(2.19)
(3.1)
(0.8)
(0.6)
(2.4)
(0.6)
(0.2)*
(6.0)
(2.3)
(3.3)*
(1.2)
(0.4)
(2.3)
(1.4)
(0.1)*
(0.1)*
(2.2)
(1.4)*
(1.1)*
(3.9)
(1.3)
(1.3)*
(1.2)
(1.4)*
(0.9)*
(3.9)
(0.1)
(0.48)
(0.80)
(0.96)
(0.47)
(0.39)
(0.64)
(0.51)
(0.31)
(0.49)
(0.61)
(0.54)
(0.53)
(0.49)
(0.04)
(0.39)
(0.28)
(0.50)
(0.38)
(1.70)
(0.32)
(0.06)
(0.91)
(0.55)*
(1.43)*
(0.83)
(0.66)*
(0.50)*
(0.31)*
(1.11)
(0.48)
(1.60)*
(0.35)
(0.59)
(2.35)*
(0.02)*
(1.06)*
(0.44)
(0.35)*
(0.32)
(1.17)*
(0.47)
(0.35)
(0.02)
(0.03)
(0.25)
(0.06)
(0.05)
(0.16)
(0.40)
(0.13)
(0.83)
(0.22)
(0.35)
(0.49)
(0.73)
(0.02)
(0.06)
(0.02)
(0.27)
(0.03)
(0.01)
(0.05)
(0.32)
(0.32)*
(0.03)
(0.06)
(0.08)*
(0.05)*
(0.08)*
(0.46)
(0.05)
(0.92)
(0.22)*
(0.21)*
(0.32)
(0.23)
(0.02)*
(0.02)
(0.02)*
(0.03)*
(0.05)
(0.01)*
(0.07)
(0.20)*
Table 3. Biomass elemental composition as total C (% DW), total N (%DW), and atomic C:N ratio, as well as internal (NO
3 +NO2 )
1
concentration (mmol l internal water)
Standard deviation in brackets (n=2–5); significant differences indicated by an asterisk (P <0.05).
Phylum
Name
Chlorophyta Acrosiphonia sp.
Chaetomorpha melagonium
Monostroma arcticum
Prasiola crispa
Rhodophyta Ceramium strictum
Devaleraea ramentacea
Odonthalia dentata
Palmaria palmata
Phycodrys rubens
Polysiphonia arctica
Ptilota plumosa
Rhodomela lycopodioides
Phaeophyta Alaria esculenta
Chorda tomentosa
Chordaria flagelliformis
Desmarestia aculeata
Fucus distichus
Laminaria saccharina
Laminaria solidungula
Scytosiphon lomentaria
Sphacelaria plumosa
Biomass C
Biomass N
Internal NO
3 +NO2
C:N ratio
Control
Enriched
Control
Enriched
Control
Enriched
Control
Enriched
20.1
29.7
24.8
30.6
25.5
25.5
35.3
27.5
30.1
12.0
24.5
27.0
34.6
16.0
26.5
37.2
31.3
26.2
25.4
10.5
25.6
17.9
32.9
26.3
31.7
28.8
33.8
31.0
32.5
26.3
30.5
27.9
20.1
32.9
17.3
22.2
36.0
31.0
24.0
30.2
13.1
29.6
2.27
3.54
1.49
3.09
3.00
2.66
2.49
1.82
3.40
1.31
2.49
1.95
1.76
1.46
1.76
2.29
2.18
1.25
1.46
0.88
2.89
2.08
3.65
2.20
4.73
2.92
2.57
2.88
1.61
3.33
2.98
3.35
2.28
1.25
1.39
1.70
2.36
1.70
0.65
1.48
1.06
2.16
10.3
9.8
19.4
8.3
9.9
11.1
16.6
17.8
10.3
10.8
11.5
11.4
23.0
12.8
17.6
19.0
18.5
24.5
20.5
13.9
14.3
10.0
10.5
14.5
7.8
9.6
15.7
11.3
23.8
9.4
12.0
11.0
10.3
30.6
14.6
15.3
17.8
21.3
42.9
24.2
14.4
16.2
27.54
80.77
3.31
3.73
26.06
53.41
11.27
7.80
11.66
1.92
6.78
1.13
1.93
0.99
2.86
1.87
7.15
2.85
2.70
2.74
4.54
42.71
62.80
13.79
3.11
11.96
41.11
9.73
2.83
14.73
1.47
3.11
1.35
1.96
0.91
6.86
1.58
1.58
1.77
1.03
3.14
4.03
(2.8)
(0.2)
(0.5)
(0.6)
(3.5)
(4.3)
(1.5)
(3.0)
(2.1)
(0.5)
(0.6)
(0.7)
(2.3)
(1.1)
(4.6)
(2.8)
(2.5)
(1.5)
(1.4)
(1.6)
(2.7)
(0.3)
(0.9)*
(0.1)*
(1.6)
(1.3)
(0.3)*
(3.3)
(2.0)
(0.6)*
(2.5)*
(2.5)
(3.3)*
(1.1)
(0.5)
(3.9)
(1.0)
(3.3)
(2.6)
(2.4)*
(4.4)
(0.4)*
Results
The experiments were carried out during the Arctic summer month of July. At that time of the year both ambient
nitrate and phosphate concentrations were close to the
detection limit (0.4 lM nitrate and 0.5 lM SRP). Ammonium levels were below detection limits (<0.1 lM). CO2
(0.02)
(0.10)
(0.04)
(1.70)
(0.00)
(0.12)
(0.22)
(0.37)
(0.05)
(0.12)
(0.18)
(1.16)
(0.07)
(0.06)
(0.33)
(0.15)
(0.86)
(0.07)
(0.22)
(0.12)
(1.14)
(0.03)
(0.72)
(0.16)*
(0.15)
(0.82)
(0.52)
(0.62)
(0.30)
(0.52)
(0.38)*
(0.42)*
(0.13)
(0.07)*
(0.05)
(0.32)
(0.02)
(0.08)
(0.02)*
(0.20)
(0.36)
(0.33)
(1.5)
(0.3)
(0.2)
(0.1)
(1.4)
(1.4)
(0.8)
(1.7)
(0.9)
(1.4)
(0.5)
(0.7)
(0.6)
(0.1)
(0.2)
(0.2)
(8.6)
(0.1)
(0.9)
(0.2)
(1.3)
(0.0)
(2.1)
(0.9)*
(0.1)*
(1.1)
(3.4)
(3.2)
(3.0)*
(1.7)
(0.5)
(0.9)
(1.1)
(0.7)*
(0.1)*
(0.2)*
(0.3)*
(3.3)
(6.1)*
(1.2)*
(0.1)
(2.7)
(4.05)
(8.14)
(0.30)
(1.05)
(0.85)
(3.10)
(0.05)
(2.12)
(0.30)
(0.62)
(0.61)
(0.06)
(0.38)
(0.03)
(0.36)
(0.39)
(1.04)
(1.42)
(1.30)
(0.25)
(0.43)
(7.58)*
(9.80)
(0.59)*
(0.52)
(1.72)*
(3.40)*
(0.17)*
(0.02)*
(1.81)*
(0.24)
(1.74)*
(0.07)*
(0.21)
(0.24)
(0.24)*
(0.05)
(0.20)*
(0.35)
(0.28)
(0.27)
(0.28)
concentration in the seawater from Kongsfjord was calculated by using the CO2sys program (Lewis and Wallace,
1998). The calculated values were 110 lM CO2
3 , 2067 lM
,
and
20
lM
CO
.
Under
these
chemical
conditions,
HCO
2
3
all the 21 species examined showed significant CAext
activity (Table 2). On average, green algae showed the
Nutrient assimilation in Arctic seaweeds
Table 4. Tissue content of soluble carbohydrates (mg Glc g
1
FW), soluble protein (mg g
1
2665
1
FW) and phycoerythrin (lg g
FW)
Standard deviation in brackets (n=2–4); significant differences indicated by an asterisk (P <0.05). nd, not determined.
Phylum
Chlorophyta
Rhodophyta
Phaeophyta
Name
Acrosiphonia sp.
Chaetomorpha melagonium
Monostroma arcticum
Prasiola crispa
Ceramium strictum
Devaleraea ramentacea
Odonthalia dentata
Palmaria palmata
Phycodrys rubens
Polysiphonia arctica
Ptilota plumosa
Rhodomela lycopodioides
Alaria esculenta
Chorda tomentosa
Chordaria flagelliformis
Desmarestia aculeata
Fucus distichus
Laminaria saccharina
Laminaria solidungula
Scytosiphon lomentaria
Sphacelaria plumosa
Soluble carbohydrates
Soluble proteins
Control
Enriched
Control
Enriched
14.1
25.0
15.0
8.1
14.0
17.6
22.6
36.1
32.0
13.2
15.0
39.4
45.7
6.9
23.0
19.9
31.8
18.7
10.0
2.2
41.3
12.5
39.7
20.4
8.7
7.3
19.7
18.3
31.9
33.5
20.1
23.6
28.2
44.8
6.4
12.7
26.0
33.9
7.0
8.0
1.5
38.7
2.37
6.79
1.34
nd
3.96
9.02
2.04
8.17
5.95
1.50
10.80
nd
2.05
0.52
3.98
8.24
9.07
0.40
1.08
0.35
1.51
0.84
5.91
1.71
nd
2.68
8.70
1.97
9.46
5.11
1.61
7.13
nd
4.59
0.63
0.83
3.86
18.63
0.51
1.24
0.39
0.76
(3.3)
(2.7)
(1.7)
(1.2)
(3.9)
(1.3)
(2.4)
(1.0)
(4.0)
(2.5)
(4.5)
(4.4)
(1.2)
(0.4)
(2.8)
(0.4)
(3.1)
(3.8)
(0.4)
(0.1)
(8.3)
(1.8)
(2.1)*
(3.9)
(2.1)
(1.4)
(2.5)
(3.3)
(0.1)*
(2.7)
(0.8)*
(4.8)
(8.0)
(1.3)
(0.6)
(3.3)*
(2.7)*
(2.7)
(0.7)*
(0.4)*
(0.3)*
(14.4)
(0.54)
(0.61)
(0.60)
(0.41)
(0.59)
(0.23)
(0.46)
(0.97)
(0.38)
(0.04)
(0.64)
(0.05)
(0.42)
(0.53)
(2.63)
(0.01)
(0.02)
(0.06)
(0.37)
Phycoerythrin
Control
Enriched
14
249
152
126
702
153
92
nd
29
135
180
60
427
60
36
nd
(0.12)*
(0.40)
(0.25)
(0.49)*
(0.31)
(0.48)
(0.98)
(0.66)
(0.41)
(0.57)*
(7)
(51)
(59)
(67)
(325)
(65)
(65)
(13)
(47)*
(100)
(24)
(135)
(24)
(5)
(0.15)*
(0.19)
(0.02)*
(0.20)*
(0.67)*
(0.05)*
(0.09)*
(0.13)
(0.02)*
Table 5. Pigment content (lg g1 FW) and the ratio accessory pigments:chlorophyll a
Standard deviation in brackets (n=2–4), and significant differences indicated by an asterisk (P <0.05).
Phylum
Chlorophyta
Rhodophyta
Phaeophyta
a
Name
Acrosiphonia sp.
Chaetomorpha melagonium
Monostroma arcticum
Prasiola crispa
Ceramium strictum
Devaleraea ramentacea
Odonthalia dentata
Palmaria palmata
Phycodrys rubens
Polysiphonia arctica
Ptilota plumosa
Rhodomela lycopodioides
Alaria esculenta
Chorda tomentosa
Chordaria flagelliformis
Desmarestia aculeata
Fucus distichus
Laminaria saccharina
Laminaria solidungula
Scytosiphon lomentaria
Sphacelaria plumosa
Chlorophyll a
Chlorophyll b
Total carotenoids
Accessory pigments:Chl. aa
Control
Enriched
Control
Enriched
Control
Enriched
Control
Enriched
588
1920
2578
3589
559
129
538
1012
918
1134
721
979
1399
1338
1595
1190
1345
403
1072
1517
1510
912
1354
1511
3083
885
330
1076
689
1239
1319
726
999
632
614
2773
1340
1165
543
777
794
1915
478
1377
2000
901
776
1130
1261
761
178
520
737
1011
69
11
57
189
141
135
82
193
271
243
507
348
327
105
152
410
487
195
390
356
1211
152
60
133
135
200
194
95
182
165
175
824
454
321
144
189
105
728
1.12
0.99
1.06
0.53
0.12
0.08
0.11
0.19
0.15
0.12
0.11
0.20
0.19
0.18
0.32
0.29
0.24
0.26
0.14
0.27
0.32
1.07
1.12
1.07
0.64
0.17
0.18
0.12
0.20
0.16
0.15
0.13
0.18
0.26
0.29
0.30
0.34
0.28
0.27
0.24
0.13
0.38
(75)
(290)
(333)
(98)
(35)
(17)
(46)
(7)
(22)
(19)
(62)
(13)
(200)
(191)
(61)
(19)
(52)
(106)
(66)
(94)
(268)
(50)*
(208)*
(181)*
(84)*
(56)*
(27)*
(92)*
(5)*
(45)*
(22)*
(13)
(24)
(90)*
(88)*
(76)*
(56)*
(36)*
(143)
(48)*
(49)*
(8)*
(27)
(217)
(218)
(31)
(33)*
(47)
(59)*
(26)*
(56)
(68)
(32)
(39)
(3)
(2)
(6)
(7)
(38)
(5)
(4)
(2)
(36)
(4)
(8)
(3)
(4)
(12)
(13)
(11)
(13)
(62)
(51)
(2)*
(47)*
(18)*
(12)*
(13)*
(5)*
(31)
(5)*
(20)
(0)*
(2)*
(3)*
(86)*
(67)
(15)
(4)*
(8)*
(7)*
(44)*
(Chl. b+carotenoids):Chl.a for Chlorophyta; carotenoids:Chl. a for Rhodophyta and Phaeophyta.
highest activity, followed by the brown algae, and the red
algae, respectively. The lowest values were obtained in
the red algae P. palmata and D. ramentacea (3.1 and 3.6
REA g1 FW, respectively) while the maximum was found
in the green alga P. crispa (59 REA g1 FW). Nine out of
the 21 species studied (two green, three red, and four
brown) significantly decreased their CAext activity when
incubated in nutrient-enriched seawater, so that decreased
CAext cannot be ascribed to any particular group (Table 6).
None of the species tested showed a significant increase.
This effect in CAext was accompanied by a general increase in NR activity (NRA, Tables 2, 6). Nitrate reductase
1
activity ranged from 0.7 to 12 lmol NO
FW h1.
2 g
Under control conditions (non-enriched), the highest values
2666 Gordillo et al.
Table 6. Statistical significance of differences produced by
nutrient enrichment in each variable measured (after standardization)
Species were grouped by phylum or considered altogether. Asterisk,
significant (P <0.05); (–), not significant for Tukey’s tests (phylumgrouped species), or two-way ANOVA tests (all species together). The
direction of significant changes are indicated by arrows (", higher in
enriched medium; #, lower in enriched medium).
Chlorophyte Rhodophyte Phaeophyte All
CAext
NRA
APA
Total internal C
Total internal N
Internal C:N atomic ratio
Internal NO
3 + NO2
Soluble carbohydrates
Soluble proteins
Phycoerithrin
Chlorophyll a
Chlorophyll b
Total carotenoids
Accessory
pigments:Chlorophyll a
–
–
–
–
–
–
–
–
–
–
–
–
–
*#
*"
–
–
–
–
*#
–
–
*#
*"
*#
*"
–
–
–
*"
–
*#
–
*#
*"
–
–
–
–
–
–
–
–
–
*"
–
–
*"
–
*"
corresponded in general to green algae, and the lowest
to brown algae; however, N enriched kelps A. esculenta
and L. solidungula, showed the overall highest values
1
(above 11 lmol NO
FW h1) together with the green
2 g
M. arcticum. Twelve out of the 21 species examined significantly increased NRA under nutrient enrichment, while
only Chorda tomentosa significantly decreased (Table 2).
The highest percentage of induction was found in
M. arcticum, D. ramentacea, P. arctica, A. esculenta,
C. flagelliformis, F. distichus, and L. solidungula with
NRA values more than double the values registered in
non-enriched conditions. The results for APA, showed a
significant increase in six species and a significant decrease in five species (Table 2). Values ranged from 0.08 to
3.20 lmol pNP g1 FW h1, the red algae showing
highest activity.
Values for total internal C (Table 3) were similar for
the three algal groups, averaging 26% DW. The lowest
values were measured in the red P. arctica (12% DW),
and the brown C. tomentosa (16% DW) and S. lomentaria
(10.5% DW) for control conditions; while the highest
corresponded to the red O. dentata (35% DW) and the
brown A. esculenta (35% DW) and D. aculeata (37%
DW), also for control conditions. Nutrient enrichment did
promote some significant but discrete changes in general
terms, with six species increasing and two species decreasing. Values under enriched conditions ranged from
13% DW in S. lomentaria to 36% DW in D. aculeata.
When considered by phylum, no significant changes were
recorded for any group (Table 6). Total internal N averaged
2.2% DW under control conditions and 2.3 under en-
riched conditions. Significant changes were only detected
in five out of the 21 species studied. Total N content was
slightly lower in brown algae (1.8% and 1.5% DW on
average) than in green (2.6% and 3.2% DW on average)
and red algae (2.4% and 2.7% DW on average), in control
and enriched conditions, respectively. When considered
by phylum, no significant changes were recorded for
any group (Table 6). Changes in internal C and N were
further evaluated by calculating the C:N ratio (Table 3).
Values ranged from 8 in the green P. crispa to 42 in
nutrient-enriched L. saccharina. The average group value
was higher in brown (18 and 22) than in green (12 and 11)
and in red (12 and 13) algae for control and enriched
conditions, respectively. No general pattern of significant
change after nutrient enrichment was found, with five
species showing higher C:N values and four species
showing lower values with respect to the control. The
Laminariales were a special case. The three Laminariales
species examined showed increased C:N values, resulting
in the significant increase observed when considering the
whole brown algae group (Table 6).
Nitrogen in the cell may be present as part of the biomass (organic) or stored in inorganic form as an internal
reservoir for later use. Table 3 shows the concentrations
for inorganic N (nitrate+nitrite) measured in the water
contained in the thalli (the latter being the difference
between FW and DW; Table 1). Stored inorganic N ranged
from 1 mM in the brown C. tomentosa and enriched
L. solidungula to 81 mM in the green C. melagonium. If
winter inorganic N had been 10, that corresponded to
accumulation factors from 3100 to 38100 (assuming
external concentration to be 10 lM inorganic N). On
average, green algae showed the highest capacity to store
inorganic N (29 mM) compared with red (15 mM) and
brown (3 mM). In N-enriched thalli, averaged differences
between groups were slightly higher (30 mM in green,
11 mM in red, and 2.5 mM in brown algae). Only five
species increased the stored amount of inorganic N after
nutrient enrichment, the green Acrosiphonia sp. and
M. arcticum, the red P. rubens, and R. lycopodioides, and
the brown C. flageliformis, while up to seven species
showed significant reduction in their internal pool of
inorganic N.
Soluble carbohydrate ranged from 1.5 to 45.7 mg
Glc g1 FW, both values being recorded in brown algae
(Table 4). Nutrient enrichment promoted a different pattern for each group. Soluble carbohydrates increased in
green algae (22% on average), changed little in red algae,
and decreased in brown algae (15% on average). Soluble
proteins ranged from 0.35 to 18 mg g1 FW, the lowest
values corresponding to brown algae, and the highest to
red algae and the brown F. distichus and D. aculeata.
Although by groups, changes in soluble protein were not
significant (Table 6), four species showed increased protein content, including the fucoid and the three laminarians,
Nutrient assimilation in Arctic seaweeds
while six others showed a significant decrease (Table 4).
The phycobiliproteins phycoerythrin (Table 4), and phycocyanin (not shown) were measured in red algae. Phycoerythrin ranged from 14 to 700 lg g1 FW, but no significant
variations could be ascribed to nutrient enrichment.
Chlorophyll a content was highest in green algae
(1942 lg g1 FW on average) and lowest in red algae
(828 lg g1 FW on average; Table 5). Chlorophyll a was
highly sensitive to nutrient enrichment, which promoted
significant variations in 18 out of the 21 species studied.
However, there was no general pattern, with nine species
increasing and nine species decreasing their chlorophyll
a content. The most consistent trend was found in red
algae with an average increase of 38% (Table 6). Chlorophyll b in green algae showed no specific trend, with two
out of four decreasing their values after nutrient enrichment
(Table 5). Total carotenoids were highly variable, ranging
from 11 lg g1 FW in the red D. ramentacea to more
than 1200 lg g1 FW in the green P. crispa. Similarly to
chlorophyll a, total carotenoids significantly varied after
nutrient enrichment in most species (Table 5), but no general trend was found, with nine species showing an increase
and six a decrease. The most regular pattern appeared
again in red algae (Table 6) with a significant increase of
30% (on average). The ratio of accessory pigments to
chlorophyll a was calculated and is shown in Table 5. Slight
increases were recorded for all three groups, but it was
significant only for brown algae (Table 6).
Figure 1 shows the results of the principal components
analyses (PCAs) performed on data for non-enriched
(upper panel) and enriched thalli (lower panel). In the
analysis for non-enriched thalli, the three algal phyla were
clearly separated in non-overlapping regions of the twodimensional space formed by axis I and axis II. The separation of each phylum generated mainly in axis I, while
axis II (and axis I to a lesser extent) served as a discriminant between species of the same phylum. Axis I was
mainly defined by CAext activity, chlorophyll a, and
total carotenoids; NRA, soluble carbohydrates and proteins, and the ratio accessory pigments:chlorophyll a
were significant but had lower loading factors (Table 7).
Axis II was most influenced by variables directly related
to nitrogen; the main component was internal N followed
by stored nitrate, the C:N ratio, and NRA. Both axes
together explained 54% of the total variance of data.
In nutrient-enriched thalli, red and green algae occupied
a region of the axis I–axis II space similar to the control
situation; however, brown algae no longer occupied the
middle region between red and green algae, but expanded
mainly along axis I. Moreover, four out of nine brown
species fell within the regions delimited by red (three
species) and green (one species). The Laminariales shifted
to the left hand side of the brown region (species 13, 18,
and 19; Table 1). Regarding the variance along axis I,
the pigment content contributed with the highest loading
2667
Fig. 1. Principal components analyses performed using the 12 variables
measured in all the 21 species studied, for both non-enriched (Control,
upper panel) and nutrient-enriched thalli (lower panel). Species numbered
as in Table 1. Curved lines were arbitrarily drawn to delimit regions for
each phylum. Loading factors and percentage of variance for each axis are
shown in Table 7. (filled squares) Chlorophytes; (open circles)
Rhodophytes; (filled triangles) Phaeophytes.
factor (Table 7) followed by CAext, internal N, the C:N
ratio, and the ratio accessory pigments:chlorophyll a.
Axis II was mainly influenced by variables directly related
to C and N composition of the biomass, such as total
internal carbon, carbohydrates, proteins, and total internal
N. The relative position of species along axis II changed
little upon nutrient enrichment.
Discussion
The presence of CAext has been hypothesized to relate
to the habitat in which populations develops (Giordano
and Maberly, 1989). Although no clear relationship
has yet been established (Mercado et al., 1998), it is generally assumed that the presence of CAext has ecological
2668 Gordillo et al.
Table 7. Loading factors for each variable of principal
components analyses shown in Fig. 1
The percentage of variance corresponding to each axis is also given.
Significant loading factors are indicated by an asterisk (P <0.05).
Control
CAext
NRA
APA
Total internal C
Total internal N
Internal C:N atomic ratio
Internal NO
3 +NO2
Soluble carbohydrates
Soluble proteins
Chlorophyll a
Total carotenoids
Accessory
pigments:Chlorophyll a
Percentage of variance
Cumulative percentage
Enriched
Axis I
Axis II
Axis I
Axis II
0.414*
0.243*
–0.240*
–0.010
–0.034
0.004
–0.086
–0.258*
–0.328*
0.463*
0.464*
0.316*
0.019
0.335*
0.153*
0.293*
0.573*
–0.346*
0.399*
0.234*
0.258*
0.099
0.112*
0.166*
0.424*
0.148*
0.089
0.027
0.352*
–0.301*
0.091
–0.036
–0.238*
0.463*
0.451*
0.302*
–0.159*
0.010
0.163*
0.442*
0.373*
–0.215*
0.296*
0.514*
0.450*
–0.071
–0.082
0.040
31.075
31.075
22.866
53.941
29.377
29.377
21.291
50.668
relevance. A particular case is the high CAext value
found for P. crispa. This species lives on land away
from seawater and its high CAext can be ascribed to
a need to convert CO2 to HCO
3 ; providing the substrate
for a presumed HCO
3 transporter. This mechanism
was earlier suggested by Raven et al. (1982) and Badger
(1987). In general, CAext seems to play a major role in
the Arctic seaweeds community as evidenced from PCAs.
The CAext loading factors for axis I (the most relevant) in
both control and enriched situations were among the
highest, together with pigment composition. Theoretically,
the impact of low temperature on the photosynthesis
of marine macrophytes is predicted to favour diffusive
CO2 entry rather than a CO2-concentrating mechanism
(Raven et al., 2002); however, the average value of CAext
was much higher for Arctic species (19 REA g1 FW )
than those reported for macroalgae from more temperate
waters, such as those from Australia (6.4 REA g1 FW;
Graham and Smillie, 1976), Scotland (5.5 REA g1 FW;
Giordano and Maberly, 1989), and Southern Spain
(3.6 REA g1 FW; Mercado et al., 1998). We propose to
call this ‘the Arctic paradox’ of the Ci uptake system. In
principle, Arctic seawater has a higher concentration of
dissolved CO2 than temperate waters, because of lower
water temperature and lower salinity. Even though it
is a well-known phenomenon that a higher concentration
of CO2 represses CA activity, CAext is difficult to relate
to the ability to use HCO
3 (Giordano and Maberly, 1989;
Mercado et al., 1998), so that the high CAext values
found in the present work may not be related to HCO
3
versus CO2-only users. The explanation for this paradox
could be that high levels of CAext are present in Arctic
species as part of a general strategy to cope with low
temperatures. This strategy consists of a general increase in
the cellular level of enzymes involved in photosynthesis,
according to Davison (1987), and is invoked to operate in
Arctic macroalgae (Korb and Gerard, 2000b). Furthermore,
increased CAext as an acclimation to low temperature has
recently been observed in macroalgae from a temperate
environment (FJL Gordillo, unpublished results).
The general decrease in CAext activity following
nutrient addition (Table 6) contradicts previous findings
that N-supply enhances CAext activity (Jiménez del Rı́o
et al., 1995). These authors ascribe the increase to Nlimitation prior to N addition. This may not be the case
for Arctic macroalgae as discussed below. On the other
hand, they added N in the form of NH4+ instead of NO
3:
The latter requires more energy in order to be incorporated
into new biomass (eight extra electrons for each atom of
N). CAext is involved in energy-costly CCMs, and in a
nitrate-enriched situation, energetic demand may be enhanced by competitive mechanisms such as nitrate assimilation, and presumably, macromolecule biosynthesis
may take place, decreasing the energetic investment into
C assimilation (Gordillo et al., 2001c). This is consistent
with the general increase in NRA observed (Tables 2, 6).
The evidence that CAext plays a major (yet unknown)
role in Arctic communities needs further and more detailed
investigations, that fall beyond the scope of the present
study.
Nitrate reductase acitvity (NRA) seems to be directly
enhanced by nitrate addition, with relatively small feedback
from the N status of the cell. Species showing the greatest
increase in NRA (above 2-fold the values under control
conditions) already had full internal inorganic N pools;
the latter showed no response or even a decrease to N
enrichment. Therefore, NRA can be considered to respond
to external levels of nitrate regardless of stored N. The
green algae M. arcticum and Acrosiphonia sp. (to a lesser
extent) were the exception, but did show a marked nitrophilic response pattern. Increased NRA, protein synthesis
and stored N, leading to lower C:N points towards optimum
utilization of readily available external N. This behaviour
is characteristic of nitrophilic metabolism in other green
macroalgae such as Ulva (Gordillo et al., 2001c).
Efficient use of low ambient irradiance for C-fixation
requires a high concentration of pigments (Falkowski and
Raven, 1997), and the maintenance of metabolic rates at
near-freezing temperatures requires a high concentration of
enzymatic proteins (Machalek et al., 1996), all of which contain nitrogen. Survival under these conditions, even with
slow growth, must require relatively high levels of nitrogenous nutrients, even in those species without a nitrophilic
metabolism. Despite the widespread enzymatic response
(lower CAext and higher NRA) to nutrient enrichment, biomass composition was not proportionally affected. According to these results, a community facing a summer input
of nutrients would have, on average, 20% less CAext
Nutrient assimilation in Arctic seaweeds
activity, and could assimilate 82% more N through NRA.
However, biomass C and N composition did not vary
beyond 8% and 4% on average, respectively, when all 21
species are considered together. The overall C:N ratio
was only affected by 7%. The maintenance of a reasonably
constant C:N ratio is a quite common priority for algae
when acclimating to external nutritional disturbance. Different regulatory mechanisms have been proposed; for
instance, organic carbon release has been shown to maintain
the metabolic integrity of the cell (Fogg, 1983; Ormerod,
1983). According to Wood and Van Valen (1990), organic
carbon release would protect the photosynthetic apparatus
from an overload of products that cannot be used for
growth or stored. In the green seaweed U. rigida, this
mechanism seems to be repressed in response to high
environmental CO2 levels; thus maintaining the internal
C:N balance (Gordillo et al., 2001c). It is unknown whether
Arctic seaweeds are able to regulate the release of organic
carbon, but they seem to maintain their metabolic integrity
through a stable C:N ratio rather than the accumulation
of N-rich compounds.
It is well documented that kelps from seasonally low
N environments accumulate internal N-reserves that support growth when external supply is low. Arctic kelp
populations seem to survive, at least in part, due to their
ability to store photosynthates during the N-depleted summer, using them for growth during the following winter
when nutrients become available. Traditionally, it was
thought that this seasonal growth pattern was a consequence
of N availability (Chapman and Lindley, 1980). However,
at least in some species, this pattern is known to be under
the control of an endogenous free-running circannual
rhythm entrained by a critical minimum daylength in the
autumn (Lüning, 1991; Schaffelke and Lüning, 1994;
Henley and Dunton, 1997). Henley and Dunton (1997)
reported changes in the biochemical composition of
L. solidungula (an Arctic endemic species) as an effect
of light availability and the interaction light3N rather
than the addition of N alone. These authors suggested
that the addition of N would only have a marginal effect
on growth and biochemical composition, presumably due
to the prevailing internal clock. Kelps studied here showed
no accumulation of N after nitrate addition. Neither total
internal N nor stored inorganic N increased, although
some degree of acclimation was observed. The increase
in the light-harvesting antennae size (as derived from
pigment ratio) and soluble protein suggest an enhanced
photosynthetic ability. This is further supported by significantly higher C:N values under N-enriched conditions.
Lack of N accumulation and enhanced photosynthetic
ability would be in agreement with the internal clock prevailing over external nutrient conditions, which in summer
would promote active photosynthesis over nutrient use,
as mentioned above. This acclimation strategy could also
be observed by principal component analysis. Under
2669
control conditions all brown species occupied a narrow
region in axis I between the red and the green group
(Fig. 1), mainly due to their pigment content and CAext
activity (Table 7). After nutrient addition, the brown group
spreads over axis I, overlapping with the red and green
region. The three Laminariales are now placed in the
most negative side of axis I for the brown region (together
with F. distichus and S. lomentaria) and well away from
S. plumosa and C. flagelliformis and the green nitrophilic
species. The variables with the most significant loading
factors along axis I are similar to control conditions, except
that internal N and C:N become highly relevant variables
after N enrichment.
The significant increase in pigment content observed
in the red algal group (Table 7) may be responsible for
its displacement towards the positive side of axis I in the
PCA analysis for enriched thalli with respect to control
conditions (Fig. 1). This displacement would also implicate
pigments, and hence photosynthesis, in their acclimation
to nutrient enrichment, as supported by the overlap observed for this group with the laminarian side of the
brown region. The pigment content of Arctic seaweeds is
known to be higher in winter than in summer (Aguilera
et al., 2002). This pattern may be speculated to be a
consequence of extreme low light and/or nutrient sufficiency in winter, the opposite occurring in summer. If
low pigmentation in summer is a consequence of N
limitation, it is expected that nutrient enrichment would
promote pigment synthesis. This seems to be the case
for most red species tested here, but is not a general trend
of the whole community (Table 6). The discrete effect of
N enrichment on pigment synthesis agrees with previous
findings according to which, pigments in Arctic seaweeds
are inversely related to light availability, increasing even
in summer periods when water becomes turbid by ice
melting (Aguilera et al., 2002). However, it was found that
the increase in the ratio of accessory pigments:chlorophyll
a is a general response to nutrient enrichment, indicating
that external nutrient level influences light harvesting
strategies. The increase in the ratio is generally related to
bigger antenna size. The result of this acclimatory response
is a decrease in photosynthesis per chlorophyll a (Gordillo
et al., 2001b). This makes sense when taking into account
that Arctic seaweeds need a strong co-ordination between
nutrient availability (normally occurring in winter) and
light availability during the summer.
PCA analysis confirmed that the effects of nutrient
enrichment on biomass composition play a marginal role.
Loading factors for total C, total N, internal inorganic
N pool, soluble carbohydrates, and soluble proteins, are
generally significant in axis II (which explains less
variance), rather than axis I in both analyses (control and
enriched conditions; Table 7). Furthermore, changes in the
relative position of species along axis II following nutrient enrichment were only discrete.
2670 Gordillo et al.
From the evidence shown here, it is inferred that
Arctic macroalgae in Kongsfjord are not N-limited, at
least during the first half of the summer. This is further
supported by the accumulation factor for inorganic N (the
difference between internal and external concentration)
averaging 31200, which is in agreement with previously
reported values for N-replete brown algae from more
temperate regions (Gordillo et al., 2001a).
In conclusion, the minimal effect of nutrient enrichment
previously reported on Laminariales has now been confirmed at the whole community level. This may well
indicate that other groups present in the Arctic could
have adapted by developing similar endogenous strategies.
Acclimation to summer nutrient input seems to involve
mechanisms that allow for the maintenance of a stable
biomass composition, rather than a rapid assimilation of
the newly available resource (except in the nitrophilic
species). This indicates a high degree of resilience of the
algal community to a disruption in the natural nutrient
availability pattern. As far as is known, there is still a
lack of evidence of progressive eutrophication in the West
coast of Svalbard; however, the fact that the Spitsbergen
current brings rich Atlantic waters from the south makes
eutrophication a likely phenomenon. The ecological
balance in Kongsfjord is influenced by oceanographic
conditions (Atlantic versus Arctic) (Svendsen et al.,
2002; Hop et al., 2002). Biodiversity could be compromised since fast-growing nitrophilic species are likely to
show enhanced growth and outcompete resilient slowgrowing species.
Acknowledgements
This work was financed by a European LSF grant (AWI-69/2001)
and was performed at the Ny Ålesund International Research and
Monitoring Facility on Spitsbergen (Svalbard, Norway). The authors
are indebted to the Norwegian Polar Institute, the Alfred Wegener
Institute for Polar and Marine Research (Germany), and to Kingsbay
AS. Francisco JL Gordillo and José Aguilera were financed by the
‘Ramón y Cajal’ programme of the Spanish Ministry of Education
and Science.
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