Oecologia (1997) 109:69–73
© Springer-Verlag 1997
&roles:Josep Peñuelas · Marc Estiarte
Trends in plant carbon concentration and plant demand
for N throughout this century
&misc:Received: 12 November 1995 / Accepted: 17 May 1996
&p.1:Abstract Atmospheric CO2 concentration has increased
by 25% over the preindustrial level. A parallel increase
in C concentration and decreases in N concentration and
δ13C of plants grown throughout this century have been
observed in plant specimens stored in herbaria. We tested
our previous results in a study of 12 more species collected in the western Mediterranean throughout this century (1920–1930, 1945–1955, and 1985–1990) and tree
rings of Quercus pubescens from the same area. These
changes were accompanied by apparent increases in condensed tannin concentration. A decreasing trend in δ15N
both in herbarium material and tree rings was also found,
indicating that ecosystems might cope with higher plant
N demand by decreasing N losses and increasing N fixation and mineralization. These results may contribute to
a better understanding of the effects of global change on
carbon and nitrogen cycling.
&kwd:Key words Atmospheric CO2 concentration · δ15N ·
δ13C · Herbarium specimens · Nitrogen&bdy:
Introduction
The long-term effects of atmospheric CO2 increases on
vegetation growing throughout this century have been
studied by the examination of plants stored in herbaria
(Woodward 1987; Körner 1988; Peñuelas and Matamala
1990; Peñuelas and Azcón-Bieto 1992), or tree rings
(LaMarche et al. 1984; Graumlich 1991). This historical
plant material integrates long-term environmental conditions, and therefore partly overcomes the limitations of
classical, mostly artificial, short-term experimentation in
controlled small-scale environments (Eamus and Jarvis
1989). Although there are some problems arriving at reliable conclusions when using herbarium material, mainly due to lack of control or knowledge of growth condiJ. Peñuelas (✉) · M. Estiarte
Centre de Recerca Ecològica i Aplicacions Forestals,
Facultat de Ciències, Universitat Autònoma, 08193 Bellaterra,
Barcelona, Spain&/fn-block:
tions and possible chemical changes during preservation,
most of the results from herbarium material (increases in
C concentration, and decreases in stomatal density, δ13C,
N and other element concentrations) mirror experimental
results obtained by exposing plants to different CO2 concentrations under controlled conditions (Woodward and
Bazzaz 1988; Polley et al. 1993).
We first aimed to corroborate our previous results on
herbarium specimens (Peñuelas and Matamala 1990;
Peñuelas and Azcón-Bieto 1992) by studying 12 more
species collected in the western Mediterranean region
and stored in herbaria throughout this century
(1920–1930, 1945–1955, and 1985–1990, when atmospheric CO2 concentrations were 300, 310 and 350 µmol
mol–1 respectively). Moreover, as the environmental conditions that produce carbon excess would also be expected to affect the amount of carbon-based secondary compounds (CBSC) in plant tissues (Herms and Mattson
1992; Peñuelas et al. 1996), we also investigated changes
in tannins as representative of CBSC. Finally, to further
investigate the interaction between nitrogen cycling and
the growth processes driven by increased carbon availability, we used the natural leaf abundances of 13C and
15N as indicators of long-term plant responses to changes
in availabilities of water, carbon and nitrogen.
Materials and methods
We studied the following species of Mediterranean trees, shrubs
and herbs: Fagus silvatica L., Fraxinus excelsior L., Quercus ilex
L., Arbutus unedo L., Laurus nobilis L., Hedera helix L., Ruscus
aculeatus L., Cistus albidus L., Salvia officinalis L., Borrago officinalis L., Amaranthus retroflexus L., and Chenopodium ambrosoides L. Leaves had been stored in the herbaria of the Departaments
de Botànica de les Facultats de Farmàcia i de Biologia de la Universitat de Barcelona. The herbarium specimens studied were collected mainly in wild areas near Barcelona (Catalonia). The drying
procedure was similar for all specimens (herbarium press).
Species collected in the same locations and seasons were sampled throughout the studied periods and leaves from the same terminal position of the herbarium specimen were analysed to avoid,
as much as possible, variability due to leaf age and position and
light environment of the collected herbarium specimens. When it
70
O E C O L O G I A 109 (1997) © Springer-Verlag
was not possible to use specimens from the same location throughout the century, the closest possible location (always in the same
west Mediterranean area) was sampled. Three to four leaves from
different herbarium specimens were analysed for each species and
time period.
Carbon and nitrogen leaf concentrations were analysed with a
Carlo Erba NA1500 analyser, using the standard configuration for
those determinations. Condensed tannins were assayed by the vanillin method (Broadhurst and Jones 1978) and the proanthocyanidin method (Porter et al. 1986). Oxidised phenolics (quinones)
were estimated by browning assay both in the soluble (absorbance
at 400 nm) and in the insoluble (L, a and b colorimetric parameters) fractions (Amiot et al. 1992). The δ15N and δ13C ratios of
herbarium leaves were measured on a SIRA Series II isotope ratio
mass spectrometer (VG Isotech, Middlewich, UK) operated in direct-inlet continous-flow mode after combustion of the samples in
an elemental analyser (NA1500, Series 1, Carlo Erba Instrumentazione, Milano, Italy). The reference CO 2, calibrated against standard Pee Dee belemnite (PDB) was obtained from Oztech (Dallas,
Tex., USA). A system check of analysis was achieved with interspersed working standards of cellulose, atropine and urea (Sigma,
St. Louis, Mo.,USA). Data on δ18O composition of the samples
were used for correction of δ13C. The accuracy of the measurement was ±0.1‰ for δ13C and 0.2–0.5‰ for δ15N. The δ13C and
δ15N of Q. pubescens tree rings from the same Mediterranean area
were also analysed. ∆13C was calculated as (δair –δplant)/(1+δplant)
after Farquhar et al. (1989) in order to concentrate on 13C biologi-
Table 1 Leaf C and N concentration (% dry weight), δ13C
(‰), and δ15N (‰) of herbarium-stored leaves of 12 species
of trees, shrubs, and herbs from
the western Mediterranean region, collected over the current
century. Values are averages±SEM (n=3 or 4)&/tbl.c:&
cal changes and avoid the effect of progressive decrease of δ13C in
the free atmosphere throughout this century.
As only a small number of samples (3–4) were obtained for
each species and period, changes relative to current values for
each species were also used in the data analysis to normalize the
variability due to species genetic differences and focus on the environmental variability. Statistical analyses used a one-way ANOVA, calculating the minimum significant difference and using
Tukey’s significant difference between means at the 0.05 level for
the periods 1920–1930, 1945–1955 and 1985–1990. All analyses
were performed using SYSTAT 5.2 (SYSTAT Inc., Evanston, Ill.,
USA) and StatView 4.5 (Abacus Concepts Inc., Berkeley, Calif.,
USA) statistical programme packages.
Results and discussion
Throughout the century there was a general decreasing
trend of leaf N concentration and an increasing trend of
leaf C concentration in most of the C3 plant species studied (Table 1). The overall average leaf carbon concentration for C3 plants was 1.9% lower and the overall average nitrogen concentration 17% higher in the first part of
this century than in the current decade (Tables 1 and 2).
Species
Period
N (%DW)
C(%DW)
δ13C (‰)
δ15N (‰)
Fagus silvatica
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
3.79 (0.06)
3.22 (0.16)
2.95 (0.14)
3.22 (0.11)
2.69 (0.02)
3.68 (0.18)
1.58 (0.15)
1.55 (0.10)
1.46 (0.04)
1.76 (0.15)
1.62 (0.14)
0.81 (0.12)
3.39 (0.57)
3.20 (0.49)
1.94 (0.08)
2.00 (0.14)
2.10 (0.17)
1.65 (0.15)
1.32 (0.14)
1.94 (0.22)
2.04 (0.14)
1.62 (0.18)
1.34 (0.14)
1.81 (0.18)
2.53 (0.30)
2.57 (0.49)
2.40 (0.22)
2.43 (0.13)
2.15 (0.12)
2.65 (0.16)
3.12 (0.14)
4.70 (0.56)
2.81 (0.25)
2.89 (0.36)
3.86 (0.42)
4.67 (0.57)
46.84 (0.19)
46.79 (0.13)
48.13 (0.15)
42.95 (0.36)
42.56 (0.15)
43.07 (0.14)
46.49 (0.13)
47.08 (0.29)
47.85 (0.17)
47.66 (0.73)
48.97 (0.11)
48.16 (0.07)
48.69 (0.17)
49.87 (0.09)
49.22 (0.13)
44.50 (0.13)
46.98 (0.18)
47.19 (0.19)
48.40 (0.32)
49.13 (0.26)
46.57 (0.11)
44.33 (0.17)
45.65 (0.05)
45.82 (0.10)
46.92 (0.51)
47.79 (0.31)
46.08 (0.27)
35.11 (0.71)
34.46 (0.62)
36.23 (0.27)
36.32 (0.13)
37.19 (0.18)
40.62 (0.19)
34.44 (0.23)
35.42 (0.35)
35.99 (0.24)
−25.20 (0.18)
−25.77 (0.14)
−27.55 (0.28)
−24.63 (0.18)
−25.90 (0.11)
−29.15 (0.21)
−23.51 (0.10)
−25.58 (0.18)
−27.65 (0.06)
−26.90 (0.19)
−26.00 (0.11)
−25.86 (0.18)
−26.22 (0.15)
−26.00 (0.26)
−27.69 (0.76)
−27.05 (0.11)
−26.77 (0.19)
−27.52 (0.28)
−27.86 (0.37)
−25.36 (0.18)
−28.59 (0.16)
−27.48 (0.15)
−27.75 (0.20)
−29.12 (0.12)
−26.73 (0.23)
−26.61 (0.27)
−29.02 (0.38)
−25.41 (0.15)
−27.26 (0.19)
−29.41 (0.30)
−25.61 (0.18)
−26.43 (0.11)
−27.74 (0.15)
−11.09 (0.05)
−10.89 (0.08)
−12.43 (0.12)
0.3 (0.20)
−0.5 (0.30)
−5.1 (1.81)
−0.5 (0.22)
−2.3 (0.43)
−2.3 (1.80)
0.5 (2.68)
−2.1 (1.42)
1.3 (0.22)
−4.3 (0.42)
−0.9 (1.30)
−9.0 (2.90)
6.5 (1.33)
1.3 (0.46)
0.0 (1.58)
4.8 (1.84)
4.5 (1.45)
2.9 (0.20)
−2.0 (1.40)
0.5 (0.22)
2.4 (0.22)
1.5 (1.50)
−1.0 (0.23)
−6.7 (1.67)
2.2 (1.64)
2.8 (1.56)
2.6 (2.29)
5.2 (1.74)
2.6 (0.46)
7.0 (2.20)
9.2 (1.84)
9.8 (2.27)
1.0 (1.05)
8.6 (0.49)
8.6 (1.87)
13.5 (2.68)
Fraxinus excelsior
Quercus ilex
Arbutus unedo
Laurus robilis
Hedera helix
Ruscus aculeatus
Cistus albidus
Salvia officinalis
Borrago officinalis
Chenopodium ambrosoides
C4 Amaranthus retroflexus
&/tbl.:
71
O E C O L O G I A 109 (1997) © Springer-Verlag
The increase in carbon concentration and the decrease in
nitrogen concentration throughout this century parallel
an increase in condensed tannins measured with the vanillin assay, although the changes were not clear when
measured with the proanthocyanidin assay, which detects
slightly different attributes of condensed tannins (Waterman and Mole 1994) (Tables 3 and 4). A change in tannin concentration, as representative of CBSC, could have
implications for plant biotic interactions through modifications in herbivory and decomposition (Herms and
Mattson 1992; Bazzaz 1990). However, these increases
in phenolic content paralleled decreases in browning (indicative of oxidised phenolic compounds) both in extractable and non-extractable material (Tables 3 and 4).
Thus, the tannin changes could be partly due to storage
degradation of phenolic compounds.
A decreasing trend in δ13C and δ15N was also found
throughout the century both in herbarium material of
most of the studied species (Table 1 and Fig. 1) and in Q.
pubescens tree rings [0.016‰ δ13C per year (r2=0.95,
n=6) and 0.014‰ δ15N per year (r2=0.83, n=6)]. The C4
plant studied, Amaranthus retroflexus, showed increasing
Table 2 Percentage change relative to the present in leaf C and
leaf Nconcentration (measured on a dry weight basis) and in C/N
ratio of herbarium-stored leaves of the C3 species of trees, shrubs
and herbs from western Mediterranean, collected over the current
century. Values are averages of overall species means for
1920–1930 and 1945–1955 samples, and 95% confidence limits
are given in parentheses&/tbl.c:&
Concentration change
relative to 1985–90
1920–1930
1945–1955
N
C
C/N
17% (14) *
–1.9% (0.8) *
–14% (10) *
12% (13) NS
–0.2% (0.9) NS
–13% (9) *
Significance of the difference from average 1985–1990 values: NS
not significant, *P<0.05&/tbl.:
Table 3 Condensed tannin leaf
concentration and browning of
herbarium-stored leaves of six
C3 species of trees and shrubs
(herbs had no detectable concentrations) from the western
Mediterranean region. Condensed tannin leaf concentration
was assayed by vanillin and proanthocyanidin procedures, and
measured as mg catechin gdw−1
and absorbance units at
550 nm gdw−1 respectively.
Browning (indicative of oxidised phenolic compounds) was
measured both in the soluble
and insoluble fractions as absorbance units at 400 nm gdw−1
and lightness L units (from 0,
black, to 100, white) respectively. Values are averages and SEM
(n=3 or 4), except for most
browning measures which required all the material available
&/tbl.c:
trends in C and decreasing trends in δ13C (similarly to C3
plants), but both N and δ15N increased throughout the
century (Table 1).
These results fit with the expected direct and indirect
(warmer and drier conditions in the western Mediterranean region in the 1980s; Burgueño 1989) effects of increased CO2 concentration and show ecologically trascendent changes in plant C and N cycling. These results
are reinforced by agreement with previous results from
herbarium specimens (Peñuelas and Matamala 1990;
Peñuelas and Azcón-Bieto 1992; Woodward 1987) and
experimental results (Woodward and Bazzaz 1988; Polley et al. 1995; Ehleringer et al. 1993). The only exception is the increase in ∆13C (Fig. 1). The δ13C values for
1985–1990 found here were more negative than in the 13
species previously studied by Peñuelas and Azcón-Bieto
(1992), and this made ∆13C increase instead of decrease.
Table 4 Percentage change relative to the present in leaf condensed tannin concentration (assayed by vanillin and proanthocyanidin procedures and measured on a dry weight basis) and in
browning (measured both in the soluble and insoluble fractions
and indicative of oxidised phenolic compounds) of herbariumstored leaves of six C3 species of trees and shrubs (herbs had no
detectable concentrations) from the western Mediterranean. Values
are averages of overall species means for 1920–1930 and
1945–1955 samples, and 95% confidence limits are given in parentheses&/tbl.c:&
Concentration change
relative to 1985–1990
1920–1930
1945–1955
Condensed tannin (vanillin)
Condensed tannin
(proanthocyanidin)
Soluble browning
Insoluble browning
−56% (12) *
–5% (10) NS
−43.5% (19) *
−32.5% (9) *
35% (30) *
10.4% (9.5) *
15% (18) NS
2.5% (8.6) NS
Significance of the difference from average 1985–1990 values: NS
not significant, *P<0.05
&/tbl.:
Species
Period
CT
(vanillin)
CT
(proanthocyanidin)
Soluble
browning
Insoluble
browning
Fagus silvatica
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
1920-30
1945-55
1985-90
8.06 (1.44)
17.40 (2.13)
15.78 (1.58)
10.95 (1.01)
9.44 (1.04)
18.21 (1.08)
24.33 (3.43)
25.91 (2.07)
73.40 (2.14)
12.91 (1.23)
7.80 (0.70)
33.63 (2.03)
4.48 (0.54)
5.21 (0.43)
7.62 (0.80)
5.01 (1.15)
18.11 (2.93)
35.41 (2.71)
186.92 (2.80)
165.88 (45.8)
192.06 (11.3)
85.88 (4.82)
56.89 (0.28)
82.70 (2.35)
100.99 (9.21)
54.51 (1.19)
106.53 (4.18)
110.27 (36.7)
71.33 (23.7)
100.69 (23.7)
2.68
1.73
1.96
1.31
1.22
1.09 (0.01)
1.41
1.85
1.19
1.80
1.56
0.92 (0.03)
1.07 (0.02)
1.87
2.01
1.41
0.93
1.13
32.36
37.27
41.88
41.68
44.27
48.99 (1.26)
30.70
36.73
45.24
38.67
41.73
47.54 (3.59)
55.99 (0.43)
57.43
56.28
33.12
38.15
45.05
Quercus ilex
Arbutus unedo
Laurus nobilis
Ruscus aculeatus
&/tbl.:
Cistus albidus
78.47 (3.08)
67.67 (1.37)
112.09 (0.60)
72
O E C O L O G I A 109 (1997) © Springer-Verlag
Fig. 1 Changes in δ15N (‰), δ13C (‰) and ∆13C (‰) of herbarium-stored leaves of 11 species of C3 trees, shrubs, and herbs from
the western Mediterranean region. Averages of overall means for
1920–1930, 1945–1955, and 1985–1990 samples. Error bars represent 95% confidence limits. Different letters (a and b) stand for
significantly different means. Atmospheric CO2 concentrations for
the three periods were 300, 310 and 350 µmol mol–1 respectively&ig.c:/f
We have no clear explanation for that change, which implies lower water use efficiency (WUE; Farqhuar et al.
1989). However, parallel increases in ∆13C and CO2 concentration have also been found in several studies (Polley
et al. 1995; Ehleringer et al. 1993; Beerling and Woodward 1995).
Together with increased CO2 and climate change
there are other factors affecting plant C and N metabolism, for instance genetics, age and position of leaf, light
environment, source of nutrients, or soil characteristics
(Herms and Mattson 1992; Jones et al. 1991). We tried to
normalize most of these factors in sampling and data
treatment but there are several remaining possible causes
of the carbon and nitrogen changes. For example, soil
weathering and long-term patterns of forest product utilization by humans, which constitute a continuous drain of
nutrients from ecosystems, could also cause the decline
in N concentration through the stimulation of nutrient
leaching from soils. However, the δ15N results are inconsistent with the hypothesis that these factors have a main
role in plant trends found throughout the century in the
Mediterranean zone studied.
As the δ15N value of a plant sample is primarily determined by the isotope ratio of the nitrogen source, decreasing leaf δ15N during this century would indicate:
1. Lower nitrogen losses in ecosystems, because all major pathways of nitrogen loss (denitrification, ammonia
volatilization, and nitrate leaching) are thought to cause
15N enrichment of the remaining nitrogen (Shearer and
Kohl 1986; Durka et al. 1994; Schulze et al. 1994)
2. Use of increasingly mineralized N, because mineralization processes discriminate against the heavy isotope
in every transformation (Shearer and Kohl 1986; Durka
et al. 1994; Schulze et al. 1994; Mariotti et al. 1982);
and/or
3. A larger proportion of N coming from fixation now
than in preceeding decades, because δ15N declines during incorporation of atmospheric N2, which has a lower
15N concentration than soil N (Shearer and Kohl 1986;
Durka et al. 1994; Schulze et al. 1994; Armstrong et al.
1994)
Decreased water availability could deplete δ15N
slightly more in nitrogen-fixing plants (Ledgard 1989).
These δ13C and δ15N results could also indicate increased secondary metabolism because compounds in
plant tissues that are the most depleted in 13C and 15N
should be products from the end of metabolic chains
(Gebauer and Schulze 1991). However, as mentioned
above, this possible increase in secondary metabolites
was not clear in the material studied.
The decreases in leaf N concentrations are consistent
with the current view that N limitations will constrain
any positive growth response of natural plants to elevated
CO2 (Shaver et al. 1992). However, data presented here
on δ15N indicate that ecosystems tend to cope with increasing plant N demands by decreasing ecosystem N
losses and increasing N fixation and mineralization. N
deposition in the studied area is low (3 kg ha–1 year–1
NH4-N and 3 kg ha–1 year–1 NO3-N for the 1985–1990
period; A. Avila and F. Roda personal communication).
In spite of the possible limitations of using herbarium
material and the great changes in Mediterranean landscapes that are likely to have altered growing conditions,
the results reported here add a new indication that the environmental changes induced by human activities, such
as increased atmospheric CO2 or changes in land use or
climate, are currently influencing C and N cycles by increasing plant C concentration and N demand. However,
confirmation is needed from new studies with larger
numbers of samples and species and a wider range of
biomes.
&p.2:Acknowledgements We thank Dr. J. Vallés and Departaments de
Botànica de la Facultat de Farmàcia i de la Facultat de Biologia,
Universitat de Barcelona, for supplying the herbarium specimens,
and CICYT, INIA (Spanish Government) and CIRIT (Catalan
O E C O L O G I A 109 (1997) © Springer-Verlag
Government) for financial support. We thank Isidre Casals and Pilar Fernández (Servei d’Anàlisi, UB), Isotope Services Inc. (Los
Alamos, New Mexico) and Larry Giles (Duke University) for
technical assistance. We also thank Dr. J. Terradas, Dr. F. Rodà
and M. Boada for providing Quercus pubescens trunks.
References
Amiot MJ, Tacchini M, Aubert S, Nicolas J (1992) Phenolic composition and browning of various apple cultivars at maturity. J
Food Sci 57:958
Armstrong EL, Pate JS, Unkovich MJ (1994) Nitrogen balance of
field pea crops in south west Australia, studied using the 15N
abundance technique. Aust J Plant Physiol 21:533–549
Bazzaz FA (1990) The response of natural ecosystems to the rising
CO2 levels. Annu Rev Ecol Syst 21:167–96
Beerling DJ, Woodward FI. (1995) Leaf stable carbon isotope
composition records increased water use efficiency of C3
plants in esponse to atmospheric CO2 enrichment. Funct Ecol
9:394–401
Broadhurst RB, Jones WT (1978) Analysis of condensed tannins
using acidified vanillin. J Sci Food Agric 29:788–794
Burgueño J (1989) Aplicacions d’un index de disparitat consecutiva a sèries pluviomètriques. MSc dissertation, University of
Barcelona
Durka W, Schulze ED, Gebauer G, Voerkellus S (1994) Effects of
forest decline on uptake and leaching of deposited nitrate determined from 15N and 18O measurements. Nature 372:
765–767
Eamus D, Jarvis PG (1989) The direct effect of increase in the
global atmospheric CO2 concentration on natural and commercial temperate trees and forests. Adv Ecol Res 19:1–55
Ehleringer JR, Hall AE, Farqhuar GD (eds) (1993) Stable isotopes
and plant carbon-water relations. Academic Press, San Diego
Farquhar GD, Ehleringer JR, Hubick KT (1989) Carbon isotope
discrimination and photosynthesis. Annu Rev Plant Physiol
Plant Mol Biol 40:503–537
Gebauer G, Schulze ED (1991) Carbon and nitrogen isotope ratios
in different compartments of a healthy and a declining Picea
abies forest in the Fichtelgebirge, NE Bavaria. Oecologia 87:
198–207
Graumlich LJ (1991) Subalpine tree growth, climate, and increasing CO2:an assessment of recent growth trends. Ecology 71:
1–11
Herms DA, Mattson WJ. (1992). The dilemma of plants: to grow
or to defend. Q Rev Biol 67:283–335
Jones JB, Wolf B, Mills HA (1991) Plant analysis handbook. Micro-Macro, Athens, Georgia
73
Körner C (1988) Does global increase of CO2 alter stomatal density? Flora 181:253–257
LaMarche VC, Graybill DA, Fritts HC, Rose MR (1984) Increasing atmospheric carbon dioxide: tree ring evidence for growth
enhancement in natural vegetation. Science 225:1019–1021
Ledgard SF (1989) Nutrition, moisture and rhizobial strain influence isotopic fractionation during N2 fixation in pasture legumes. Soil Biol Biochem 21:65–68
Mariotti A, Mariotti F, Champigny ML, Amarger N, Moyse A
(1982) Nitrogen isotope fractionation associated with nitrate
reductase activity and uptake of NO3– by pearl millet. Plant
Physiol 69:880–884
Peñuelas J, Azcón-Bieto J (1992) Changes in leaf ∆13C of herbarium plant species during the last 3 centuries of CO2 increase.
Plant Cell Environ 5:485–489
Peñuelas J, Matamala R (1990) Changes in N and S leaf content,
stomatal density and specific leaf area of 14 plant species during the last three centuries of CO2 increase. J Exp Bot 230:
1119–1124
Peñuelas J, Estiarte M, Kimball BA, Idso SB, Pinter PJ, Wall GW,
Garcia RL, Hansaker DJ, LaMorte RL, Hendrix DL (1996)
Variable responses of plant phenolic concentration to CO 2 enrichment. J Exp Bot, in press
Polley HW, Johnson HB, Mayeux HS, Malone SR (1993) Physiology and growth of wheat across a subambient carbon dioxide
gradient. Ann Bot 71:347–356
Polley HW, Johnson HB, Mayeux HS (1995) Nitrogen and water
requirements of C3 plants grown at glacial to present carbon
dioxide concentrations. Funct Ecol 9:86–96
Porter LJ, Hrstich LN, Chan BC (1986) The conversion of procyanidins and prodelphinidins to cyanidin and delphinidin. Phytochemistry 25:223–230
Schulze ED, Chapin FS III, Gebauer G (1994) Nitrogen nutrition
and isotope differences among life forms at the northern treeline of Alaska. Oecologia 100:406–412
Shaver GR, Billings WD, Chapin FS III (1992) Global change and
the carbon balance of artic ecosystems. BioScience 42:
433–441
Shearer G, Kohl DH (1986) N2-fixation in field settings:estimations based on natural 15N abundance. Aust J Plant Physiol 13:
699–756
Waterman PG, Mole SM (1994) Analysis of plant phenolic metabolites. Blackwell, Oxford
Woodward FI (1987) Stomatal numbers are sensitive to increases
in CO2 from pre-industrial levels. Nature 327:617–618
Woodward FI, Bazzaz FA (1988) The responses of stomatal density to CO2 partial pressure. J Exp Bot 209:1771–1781