Directory UMM :Data Elmu:jurnal:T:Tree Physiology:Vol15.1995:

Tree Physiology 15, 351--359
© 1995 Heron Publishing----Victoria, Canada

Responses of foliar gas exchange to long-term elevated CO2
concentrations in mature loblolly pine trees
SHANYIN LIU1 and ROBERT O. TESKEY1,2
1

Warnell School of Forest Resources, The University of Georgia, Athens, GA 30602, USA

2

Author to whom correspondence should be addressed

Received October 13, 1994

Summary Branches of field-grown mature loblolly pine
(Pinus taeda L.) trees were exposed for 2 years (1992 and 1993)
to ambient or elevated CO2 concentrations (ambient + 165
µmol mol −1 or ambient + 330 µmol mol −1 CO2). Exposure to
elevated CO2 concentrations enhanced rates of net photosynthesis (Pn) by 53--111% compared to Pn of foliage exposed to

ambient CO2. At the same CO2 measurement concentration, the
ratio of intercellular to atmospheric CO2 concentration (Ci/Ca)
and stomatal conductance to water vapor did not differ among
foliage grown in an ambient or enriched CO2 concentration.
Analysis of the relationship between Pn and Ci indicated no
significant change in carboxylation efficiency of ribulose-1,5bisphosphate carboxylase/oxygenase during growth in elevated CO2 concentrations. Based on estimates derived from
Pn /Ci curves, there were no apparent treatment differences in
dark respiration, CO2 compensation point or Pn at the mean Ci.
In 1992, foliage in the three CO2 treatments yielded similar
estimates of CO2-saturated Pn (Pmax), whereas in 1993, estimates of Pmax were higher for branches grown in elevated CO2
than in ambient CO2. We conclude that field-grown loblolly
pine trees do not exhibit downward acclimation of leaf-level
photosynthesis in their long-term response to elevated CO2
concentrations.
Keywords: acclimation, intercellular CO2, net photosynthesis,
Pinus taeda.

Introduction
Global atmospheric CO2 concentration has been increasing
since the beginning of the industrial revolution in the mid-18th

century (Neftel et al. 1985, Keeling and Whorf 1991) and is
predicted to double at some time in the mid- to late 21st
century (Keeling et al. 1989). Because this potential increase
may have both direct and indirect effects on vegetation, studies
of the ecological effects of rising atmospheric CO2 and associated global climate change constitute a major research challenge (Taylor and Grotch 1990).
Forests range across some 30% of the Earth’s land surface
(Myers 1993) and are estimated to account for approximately

70% of terrestrial carbon assimilation (Waring and Schlesinger
1985). Model predictions (Kohlmaier et al. 1987, Houghton et
al. 1987, Tans et al. 1990) and eddy correlation observations
(Wofsy et al. 1993) suggest that forests may constitute a
potential sink for the increasing atmospheric CO2 concentration. Because estimates of many of the components in the
global carbon budget have not been validated (Houghton et al.
1983, Sundquist 1993), information on the reaction of trees
and forests to an increase in atmospheric CO2 concentration is
needed to assess the ecological consequences of an increasing
atmospheric CO2 concentration. Although there have been
many studies on this subject, many questions still remain to be
answered (Eamus and Jarvis 1989, Graham et al. 1990). In

particular, the responses of older trees, i.e., beyond the seedling and sapling stages, have not been well characterized.
Older trees have many distinct morphological, phenological
and physiological differences from seedlings and saplings
(Cregg et al. 1989). These differences raise concerns about
whether information gathered from seedlings or saplings can
adequately characterize the responses of mature trees to increasing atmospheric CO2 concentrations (Barton et al. 1993).
We have assessed the effects of elevated CO2 concentrations
on the gas exchange characteristics of foliage of mature trees
growing in field conditions. The project was begun in 1992 in
a 21-year-old plantation of loblolly pine (Pinus taeda L.) using
the branch chamber technique (Teskey et al. 1991). The branch
chamber technique has the disadvantage that the response of a
single branch may not be the same as that of a whole tree when
exposed to elevated CO2 concentrations; however, we believe
that it may be of value for comparative studies of the effects of
long-term CO2 enrichment on physiological and developmental processes in foliage of large trees. An alternative to this
technique would be to enclose whole trees in large open-top
chambers, but this approach has the disadvantage that it usually results in a large change in the microclimate around the
tree. Open air exposure systems are another option but are
expensive to operate, which limits replication. In contrast,

branch chambers create less alteration of the microclimate than
whole-tree chambers, and treatments can easily be replicated
because of the small size and low cost of the chambers.

352

LIU AND TESKEY

Materials and methods
Study site and plant material
The research was conducted in 1992 and 1993. The study site
was located in the Whitehall Forest, an experimental forest of
the University of Georgia, situated about 3 km south of Athens,
Georgia (33°57′ N, 83°19′ W). The site is occupied by a
plantation of 21-year-old mature loblolly pine trees, with a
spacing of approximately 3 × 3 m. Site index of the study area
on a 25-year basis is 16.5 m (Bailey et al. 1985), with a low to
medium soil fertility; nitrogen concentration of current-year,
first-flush foliage is approximately 1.2% in late August to early
October. The soil on the site is in the Pacolet series, approximately 0.9 m deep, well drained, and with a sandy loam texture. The average annual precipitation is 1257 mm, and the

mean annual air temperature is 16.5 °C, with mean summer
and winter temperatures of 25.4 and 7.2 °C, respectively.
Ambient CO2 concentrations at the site, monitored during the
past several years, are consistent with worldwide seasonal
averages, and range between 330 and 340 µmol mol −1 during
the day and between 420 and 440 µmol mol −1 at night.
Experimental design
The experiment was conducted on upper mid-canopy branches
of six healthy trees of average diameter at breast height and
canopy size. Each tree was surrounded by four scaffolding
towers linked by walkways to hold experimental apparatus and
to provide access to branches. On each of the six trees, three
main branches were chosen from the middle portion of the
crown. Each branch was enclosed in a chamber and randomly
assigned to receive one of three CO2 treatments: fumigation
with ambient air (about 330 µmol mol −1 CO2, control), fumigation with air containing 495 µmol mol −1 CO2 (ambient air +
165 µmol mol −1 CO2), and fumigation with air containing 660
µmol mol −1 CO2 (ambient air + 330 µmol mol −1 CO2), for a
total of 18 branch chambers.
The design of the branch chamber has been described elsewhere (Teskey et al. 1991). Briefly, each chamber was cylindrical and consisted of a rigid supporting frame made of

aluminum alloy covered with a transparent polyvinylchloride
plastic film (Livingstone Coating Corp., Columbia, SC), with
dimensions of 1.5 m in length and 0.5 m in diameter. There was
a 1.5-m long nylon zipper sewn to the plastic covering to allow
easy access to the foliage in the chamber. The chamber was
open at the bottom and there was a plenum at the top that had
a series of holes to help distribute the air uniformly across the
chamber. Each chamber was fastened at the top and bottom to
the towers and walkways. Each chamber had one blower
(Model AF-8, American Fan Co., Fairfield, OH) mounted on
the walkway below it, and air was continuously blown into the
chamber. Because of the relatively high rate of air exchange
(10 times per min), the temperature and relative humidity
within the chambers remained close to ambient. The temperature differed by 0.3 °C and relative humidity by 0.7% inside
and outside the chamber, and PAR within the chambers was
lowered by approximately 10% (Teskey et al. 1991).
Elevated CO2 was generated from liquid CO2 (Airco, Mur-

ray, NJ), which was vaporized and injected into the air stream
as it exited from the blower. Control of CO2 concentrations in

the branch chambers was realized by a combination of mass
flow controllers and rotometers. To determine CO2 concentrations, air samples were measured twice hourly by means of a
microcomputer-controlled switching system sequentially
routed to an infrared gas analyzer (IRGA) (Model 6252, LiCor Inc., Lincoln, NE).
Beginning in March 1992, the branches were exposed to the
CO2 treatments for 24 h per day, 7 days per week for the
duration of the project. To avoid water stress, the irrigation
system was turned on when soil water was at or below −0.05
MPa and turned off at field capacity.
Gas exchange measurements
Gas exchange data were collected with a closed-loop LI-6200
portable photosynthesis system (Li-Cor Inc.) with a 250-ml
leaf cuvette. Values of net photosynthesis (Pn), stomatal conductance (gs) and intercellular CO2 concentration (Ci), on a
total needle surface area basis, were calculated according to
von Caemmerer and Farquhar (1981). Measurements were
conducted at ambient conditions between 1000 and 1700 h
Eastern Standard Time (EST) at saturating sunlight (PAR
1000--2000 µmol m −2 s −1) from late August to mid-October in
both 1992 and 1993. Each measurement took approximately
20--50 s. Usually, the change during the measurement period

in relative humidity was ≤ 2% and in leaf temperature ≤ 2 °C.
All the measurements were made inside the branch chambers
on attached, south-facing, current-year foliage from the middle
portion of the first flush developed under the experimental
conditions. The CO2 concentrations at which measurements
were made were 330, 495 and 660 µmol mol −1 CO2 for the
ambient CO2 treatment (control), 330 and 495 µmol mol −1 CO2
for the 1.5× ambient CO2 treatment, and 330 and 660 µmol
mol −1 CO2 for the 2.0× ambient CO2 treatment. For each
branch chamber, four to six measurements were made at each
measurement CO2 concentration using different sets of fascicles. Measurements of the three treatments on each tree were
completed on the same day.
Measurements of Pn /Ci curves were also made with an
LI-6200 portable photosynthesis system. The measuring procedure was similar to that described by Davis et al. (1987) and
McDermitt et al. (1989). Beginning with the highest CO2
concentration, photosynthesis was measured at seven to 11
different CO2 concentrations. Concentrations of CO2 higher
than those of the exposure treatments were obtained by breathing into the cuvette, and lower concentrations were achieved
by either scrubbing CO2 from the system with soda lime or
allowing foliage to assimilate CO2. The increased humidity in

the cuvette resulting from breathing was brought down to the
ambient relative humidity before measurement of Pn. At each
measurement CO2 concentration, data were logged as soon as
equilibration was reached and the CO2 concentration started to
drop steadily. To minimize the heat load within the cuvette, a
small external fan was used to ventilate the outside of the
cuvette. On average, leaf temperatures in the cuvette increased
by ≤ 2 °C over the course of a complete sequence of measure-

GAS EXCHANGE OF LOBLOLLY PINE IN ELEVATED CO2

ments. To avoid overestimation of carboxylation efficiency
(Pn /Ci curve initial slope), cuvette chamber leaks were mathematically corrected using the Li-Cor protocol (Application
Note No. 103, 1991). Measurement of each Pn/Ci curve took
approximately 20 min. Two to three Pn/Ci curves were measured on each branch using different sets of fascicles.
Data analysis
The initial slopes of the Pn/Ci curves (dPn/dCi) were calculated
by linear regression of the first several points (Ci below 200-250 µmol mol −1). The whole Pn /Ci curves were then fitted to
the empirical nonlinear model:
Pn = Pmax1 − (1 − Rd / Pmax )(1 − C i / Γ) ,




(1)

where Pmax, the asymptotic level, represents the predicted CO2saturated Pn, Rd is the rate of dark respiration at zero CO2, and
Γ is the CO2 compensation point (Taylor and Gunderson
1988). The fitting was carried out by the Marquardt-Levenberg
iterative least-square method available in SigmaPlot software
(Jandel Scientific, San Rafael, CA).
Relative limitation of stomatal conductance on photosynthesis (Ls) was estimated according to the equation:
Ls = (1 − Pi / Pa)100 ,

(2)

where Pi is the estimated Pn at Ci corresponding to the CO2
concentration in a specific ambient air (Ca), and Pa is the
estimated Pn when Ci = Ca, the rate at which stomatal resistance
to CO2 diffusion is assumed to be zero (Farquhar and Sharkey
1982). Both Pi and Pa were derived using Equation 1.

Treatment differences in foliar gas exchange data, Pn/Ci
curve initial slopes and estimates of other parameters were
determined by analysis of variance (ANOVA) with SAS software (SAS Institute Inc., Cary, NC). The experiment was
treated as a randomized complete block design, with individual trees as blocks and individual branches receiving specific
treatments as experimental units. The basic linear additive
model was:
yijk = µ + τ i + βj + εij + δijk ,

(3)

where yijk is the value of the kth measurement for the ith branch
on the jth tree, µ is the overall mean, τi is the branch (i.e., CO2
treatment) effect where i = 1, 2 or 3, βj is the tree (i.e., block)
effect where j = 1, 2, 3, 4, 5 or 6, εij is the experimental error
(= (τβ)ij, treatments × trees), and δijk is the sampling error.
Least significant difference (LSD) pairwise comparisons
were made to separate the means of the parameters when
ANOVAs indicated significant treatment effects.
Results
Net photosynthetic rate (Pn) of current-year, first-flush foliage
of mature loblolly pine trees consistently responded positively
to elevated growth CO2 concentrations in both 1992 and 1993
(P < 0.0002 for 1992, Figure 1A; P < 0.0001 for 1993, Figure

353

1B). Doubling of the ambient CO2 concentration (660 µmol
mol −1) increased Pn by 92% in 1992 and by 111% in 1993,
whereas enhancement of Pn for foliage grown at 495 µmol
mol −1 CO2 was 61and 53% in 1992 and 1993, respectively. A
comparison of the results for 1992 and 1993 indicated that
there was no reduction in the enhancement of Pn with exposure
time for branches grown at high concentrations of CO2.
Stomatal conductance, gs, was not significantly affected by
elevated CO2. In 1993, foliage of the branches in the three CO2
treatments had almost identical gs (P < 0.6594, Figure 1D).
Similarly, in 1992, the treatment difference was not statistically significant (P < 0.3692), although gs was somewhat
higher for foliage of branches grown at 495 µmol mol −1 CO2
and lower for foliage grown at 660 µmol mol −1 CO2 compared
with foliage of branches grown under ambient conditions (Figure 1C). The ratio of intercellular CO2 concentration to atmospheric CO2 concentration (Ci/Ca) was unaffected by the CO2
enrichment treatments in both years (P < 0.4229 for 1992,
Figure 1E; P < 0.3363 for 1993, Figure 1F).
Net photosynthesis tended to be lower in foliage of branches
grown at elevated CO2 concentrations than in foliage of
branches grown at 330 µmol mol −1 CO2 when measured at the
same atmospheric CO2 concentration (Figure 2). Stomatal
conductance and Ci also tended to be slightly less in branches
grown at 660 µmol mol −1 CO2 than in branches grown at 330
µmol mol −1 CO2. Nevertheless, results of ANOVAs for the two
years of measurements indicated that there were no statistically significant differences in Pn, gs or Ci among the three
treatments when measured at the same CO2 concentration
(Figure 2A--F, P-values ranged from 0.2391 to 0.9609).
Elevated CO2 concentrations during growth had little effect
on the relationship between Ci and Pn (Figure 3). Foliage in the
three treatments had similar Pn/Ci function curves in both 1992
and 1993. Analysis of initial slopes of the Pn/Ci curves showed
that, although the initial slope tended to be slightly lower for
branches grown at elevated CO2 concentrations, the treatment
differences were not significant (Table 1).
Carbon dioxide-saturated Pn (Pmax), foliar dark respiration at
zero CO2 (Rd), CO2 compensation point (Γ) and Pn at Ci values
corresponding to atmospheric CO2 concentrations of 330, 495
and 660 µmol mol −1 were estimated from the individual Pn/Ci
curves using Equation 1 (Table 2). There were no effects of the
treatments on Pmax of current-year, first-flush foliage of
branches in 1992. In contrast, a significant difference in Pmax
was detected among treatments in 1993. Compared with the
branches grown at ambient CO2 concentration, Pmax increased
by 7 and 21% in branches grown at 495 and 660 µmol mol −1
CO2, respectively. There were no differences in Rd and Γ
among the three CO2 treatments. The estimated average Pn at a
given Ci had a tendency to decrease with increasing treatment
CO2 concentration, but the differences among treatments were
consistently nonsignificant for both 1992 and 1993.
Although relative stomatal limitation of Pn (Ls) tended to be
slightly higher for foliage of branches grown at elevated CO2
concentrations (Table 2), there were no significant differences
in Ls among treatments except in foliage grown at 660 µmol
mol −1 CO2, where Ls was significantly higher than in foliage

354

LIU AND TESKEY

Figure 1. Rate of net photosynthesis (Pn),
stomatal conductance (gs) and the ratio of
intercellular to atmospheric CO2 (Ci/Ca)
during late August to mid-October of
1992 and 1993 for current-year, first-flush
foliage of loblolly pine tree branches
grown at 330, 495 or 660 µmol mol −1
CO2. The measurements were made at
saturating PAR (≥ 1000 µmol m − 2 s − 1)
and each respective growth CO2 concentration. Values for Pn, gs and Ci/Ca ratio are
means ± MSE.

grown at 330 µmol mol −1 CO2 in 1993 (28.4% versus 22.4%,
P < 0.0435). When treatments were compared for Ls at their
respective growth Ca, however, the differences were consistently significant in 1992 and 1993. Relative limitation of
stomatal conductance on photosynthesis was lower in foliage
grown at the elevated CO2 concentrations compared with Ls in
foliage grown at the ambient CO2 concentration. Compared
with Ls of the control (330 µmol mol −1 CO2) treatment, Ls was
23 and 31% lower in the 495 and 660 µmol mol −1 CO2
treatments, respectively, in 1992, and in 1993, the decline in Ls
was 14 and 29% in the 495 and 660 µmol mol −1 CO2 treatments, respectively.

Discussion
Short-term CO2 enrichment typically enhances leaf net photosynthesis in trees (Eamus and Jarvis 1989); however, this

short-term leaf-level photosynthetic enhancement may not be
maintained in trees during long-term exposure to elevated CO2
concentrations (Gunderson and Wullschleger 1994). We found
that branches of mature loblolly pine trees exhibited increased
Pn in current-year, first-flush foliage in response to long-term
elevated exposures to 495 or 660 µmol mol −1 CO2 (Figures 1A
and 1B). We conclude that there was no reduction in the
enhancement of photosynthetic capacity of the foliage grown
at elevated CO2 over the 2-year study, because there was no
difference in Pn between CO2 treatments when measured at the
same CO2 concentrations (Figures 2A and 2B). These results
suggest that there was no downward photosynthetic acclimation of Pn during two growing seasons. This result is consistent
with other CO2-enrichment studies conducted in the field or in
large pots (Sage 1994).
Curves of Pn as a function of intercellular CO2 concentrations (Pn /Ci curves) have been used to assess photosynthetic

GAS EXCHANGE OF LOBLOLLY PINE IN ELEVATED CO2

355

Figure 2. Comparative rates of net photosynthesis (Pn), intercellular CO2 concentrations (Ci) and stomatal conductance (gs)
for current-year, first-flush foliage of loblolly pine tree branches grown and measured at 330, 495 or 660 µmol mol − 1 CO2
in 1992 and 1993. The measurements
were made at saturating PAR (≥ 1000
µmol m −2 s −1) during late August to midOctober. For foliage grown at 330 µmol
mol − 1 CO2, gas exchange was measured
at CO2 concentrations of 330, 495 and 660
µmol mol −1 CO2. For foliage grown at
495 µmol mol − 1 CO2, measurements were
made at 330 and 495 µmol mol − 1 CO2.
For foliage grown at 660 µmol mol − 1
CO2, measurements were made at 330 and
660 µmol mol − 1 CO2. Values for Pn, Ci
and gs are means ± MSE.

acclimation and to identify the mechanisms contributing to
acclimation (Harley et al. 1992, Gunderson and Wullschleger
1994, Sage 1994). In the Pn/Ci curve, Pn is theoretically limited
by carboxylation efficiency (amount, activity and kinetic properties) of Rubisco at low Ci, by ribulose-1,5-bisphosphate
(RuBP) regeneration at intermediate Ci, and by inorganic phosphate (Pi) regeneration capacity at high Ci (von Caemmerer
and Farquhar 1981, Sharkey 1985, Harley and Sharkey 1991).
We observed no significant change in Rubisco carboxylation
efficiency in foliage grown at elevated CO2 (Table 1). Also,
there was no reduction in CO2-saturated Pn (Pmax) or Pn at
average Ci values, further demonstrating that long-term elevated CO2 fumigation of branches does not induce down-regulation of photosynthetic capacity. In 1993, estimates of Pmax
were higher for branches grown at elevated CO2 than for

branches grown at ambient CO2, perhaps reflecting a gradual
increase in Pi regeneration capacity for growth during longterm exposure to elevated CO2 (Makino 1994, Sage 1994).
Positive acclimation of photosynthesis has been observed in
mature Picea sitchensis (Bong.) Carr. (Barton et al. 1993) and
nonwoody plants (Campbell et al. 1988, Arp and Drake 1991)
grown in the field.
The absence of downward photosynthetic acclimation is
similar to the findings of the other few field studies on woody
species (Idso and Kimball 1991, Barton et al. 1993, Dufrêne et
al. 1993, Gunderson et al. 1993), but contrasts with responses
in many experiments with potted plants (see reviews by Eamus
and Jarvis 1989 and Sage 1994). Field-grown trees have a
large, if not unlimited, rooting volume, which represents a
large sink for photosynthates. The fact that only a portion of

356

LIU AND TESKEY

Figure 3. Representative response curves
of net photosynthesis (Pn) to intercellular
CO2 concentration (Ci) for current-year,
first-flush foliage of loblolly pine
branches grown at 330, 495 or 660 µmol
mol − 1 CO2 in 1992 and 1993. The measurements were made at ambient temperature and saturating PAR (≥ 1000 µmol
m − 2 s − 1) in early October in one tree.

Table 1. Slopes of the initial linear portion of the Pn/Ci curves for
current-year, first-flush foliage of loblolly pine branches grown at 330,
495 or 660 µmol mol − 1 CO2 , derived from the linear regression of the
first several points with Ci below 200--250 µmol mol −1. Measurements were taken from late August to mid October for both years
(1992 and 1993). Values presented are means ± SE (n = 9--13). The
unit for the initial slopes is mol m −2 s − 1. The level of significance (P)
is given in the last row.
Treatment [CO2]

330
495
660
P

Initial slope
1992

1993

0.0276 ± 0.0014
0.0258 ± 0.0016
0.0250 ± 0.0023
0.4031

0.0283 ± 0.0017
0.0253 ± 0.0010
0.0249 ± 0.0021
0.4279

the tree crown was exposed to elevated CO2 concentrations in
this study may be another reason for the large sink strength in
our trees. In contrast, root growth is restricted in many pot
experiments, and this may reduce sink demand for photosynthates thereby causing down-regulation of Pn in the potted
plants (Ehret and Joliffe 1985, Arp 1991, Stitt 1991, Thomas
and Strain 1991). The pot-size effect could also be attributed
to limited nutrient supply (Conroy et al. 1986, Fetcher et al.
1988, Norby and O’Neill 1991, Silvola and Ahlholm 1992,
Tissue et al. 1993, El Kohen and Mousseau 1994, Nobel et al.
1994, Thomas et al. 1994) and the release of inhibitory signals
(e.g., ABA) in response to pot-associated physical stresses
(Sage 1994).
Stomatal response to CO2 is a common phenomenon (Morison 1987, Mott 1990) and stomatal conductance (gs) in many
plants decreases in response to increasing atmospheric CO2
concentrations (Oberbauer et al. 1985, Tolley and Strain 1985,
Fetcher et al. 1988, Eamus et al. 1993, Thomas et al. 1994). In
our study, however, elevated CO2 treatments did not alter foliar
gs in mature loblolly pine trees (Figures 1C and 1D). In loblolly
pine seedlings, Tolley and Strain (1985) also found that there

was no difference in gs between CO2 treatments when plants
were grown at high irradiances (1000 µmol m −2 s −1). Similar
results have been reported in Pinus contorta Dougl. ex Loud.
(Higginbotham et al. 1985), Pinus radiata D. Don (Conroy et
al. 1986, 1988), Malus domestica Borkh, Quercus prinus L.
and Quercus robur L. (Bunce 1992), Fagus sylvatica L. (Dufrêne et al. 1993), and Liriodendron tulipifera L. and Quercus
alba L. (Gunderson et al. 1993). However, using the same
branch bag technique, Barton et al. (1993) found that, in
mature P. sitchensis, gs was higher in shoots grown in an
elevated CO2 concentration than in shoots grown in ambient
CO2. Although the mechanisms by which CO2 affects stomatal
aperture and gs are not known (Morison 1987, Mott 1990),
there may be interspecific variation in CO2 sensitivity of gs
(Eamus and Jarvis 1989). In our study, lack of stomatal sensitivity to CO2 might be associated with environmental conditions during growth. The branches receiving the CO2
treatments were grown at full PAR with an adequate supply of
soil water, and stomatal sensitivity to CO2 may be reduced at
high PAR in well-watered plants (Eamus and Jarvis 1989).
The ratio of intercellular to atmospheric CO2 (Ci/Ca) is
directly linked to changes in the relationship between gs and
the biochemical characteristics of Pn (Ball and Berry 1982) and
may be a useful parameter for evaluating possible stomatal
acclimation to long-term CO2 exposure (Sage 1994), although
no clear mechanistic basis for this relationship has been demonstrated (Mott 1990). The Ci/Ca ratio was constant during the
2 years of elevated CO2 exposure (Figures 1E and 1F), indicating that there was no stomatal acclimation to long-term elevated CO2 in mature loblolly pine trees. Results of the
treatment comparison of gs and Ci supported this conclusion,
i.e., when measured at the same Ca concentrations, no differences in gs and Ci between CO2 treatments were observed
(Figures 2C--F). However, elevated CO2 concentrations during
growth decreased the stomatal limitation to photosynthesis (Ls)
(Table 2), reflecting unchanged rates of gas phase diffusion of
CO2 and an increase in biochemical limitations with increasing
Ci. This result is similar to those observed in cotton (Thomas

GAS EXCHANGE OF LOBLOLLY PINE IN ELEVATED CO2

357

Table 2. Foliar gas exchange parameters for branches of mature loblolly pine grown at 330, 495 or 660 µmol mol − 1 CO2 in branch chambers,
estimated from response curves of net photosynthesis (Pn) to intercellular CO2 concentrations (Ci) measured in late August to mid-October of 1992
and 1993. Values are means ± SE for each treatment (n = 9--13). The fitting model used was Equation 1. The Ls value was calculated using
Equation 2. Parameters are defined in the text. The level of significance (P) is given in the last column. Units are µmol m − 2 s − 1 for Pmax , Rd and
Pn, µmol mol −1 for Γ, and Ls is a percentage.
Parameter

Treatment CO2 concentration (µmol mol − 1)

P<

330

495

660

1992
Pmax
Rd
Γ
Pn at Ci = 220
Pn at Ci = 330
Pn at Ci = 420
Ls at Ca = 330
Ls at Ca = 495
Ls at Ca = 660
Ls at growth Ca

13.95 ± 0.94
−3.20 ± 0.25
80.2 ± 4.5
4.08 ± 0.19
6.36 ± 0.22
7.80 ± 0.26
36.0 ± 1.3
27.3 ± 1.3
24.3 ± 1.5
36.0 ± 1.3 a1

14.04 ± 0.85
−2.80 ± 0.32
75.5 ± 7.9
4.07 ± 0.29
6.30 ± 0.33
7.73 ± 0.37
36.2 ± 2.0
27.8 ± 1.5
25.1 ± 1.6
27.8 ± 1.5 b

13.96 ± 1.10
−2.92 ± 0.53
72.2 ± 9.3
3.81 ± 0.19
5.93 ± 0.31
7.28 ± 0.39
36.1 ± 1.8
27.6 ± 1.5
24.9 ± 1.8
24.9 ± 1.8 b

0.3442
0.9387
0.7629
0.6402
0.6685
0.7288
0.9024
0.7472
0.5503
0.0006

1993
Pmax
Rd
Γ
Pn at Ci = 200
Pn at Ci = 290
Pn at Ci = 400
Ls at Ca = 330
Ls at Ca = 495
Ls at Ca = 660
Ls at growth Ca

12.21 ± 1.41 a
−3.29 ± 0.76
69.4 ± 11.9
3.97 ± 0.33
5.88 ± 0.33
7.57 ± 0.37
40.0 ± 3.5
31.3 ± 2.7
22.4 ± 2.5 a
40.0 ± 3.5 a

13.04 ± 1.29 ab
−2.31 ± 0.40
67.4 ± 10.6
3.40 ± 0.27
5.12 ± 0.29
6.76 ± 0.34
41.4 ± 3.2
34.4 ± 2.9
26.4 ± 2.8 ab
34.4 ± 2.9 ab

14.76 ± 1.38 b
−3.17 ± 1.01
70.1 ± 11.5
3.5 ± 0.22
5.59 ± 0.31
7.61 ± 0.53
43.9 ± 3.2
36.6 ± 1.8
28.4 ± 1.3 b
28.4 ± 1.3 b

0.0313
0.4679
0.7809
0.3604
0.1898
0.0997
0.3177
0.0630
0.0435
0.0214

1

Values followed by different letters in the same row are significantly different (P < 0.05).

and Strain 1991) and P. radiata (Whitehead et al. 1992).
To conclude, current-year, first-flush foliage of branches
exposed to ambient + 165 or ambient + 330 µmol mol −1 CO2
concentrations in branch chambers for 2 years had greatly
enhanced Pn compared with Pn of foliage exposed to ambient
CO2 concentration. Various foliar gas exchange measurements
and the modeling results of Pn/Ci curves indicated that there
was no reduction in photosynthetic capacity in response to
elevated CO2 in the field-grown mature loblolly pine trees
during the 2-year study. We also demonstrated that field-grown
trees do not experience the downward photosynthetic acclimation typically observed in long-term elevated CO2 experiments
with potted plants. Although there is need for caution when
extrapolating data derived from the branch chamber technique
to whole trees, we note that our findings are consistent with
other long-term whole-plant field studies (Idso and Kimball
1991, Gunderson et al. 1993, Drake et al. 1994).
Acknowledgments
This research was supported by the USDA Forest Service Southern
Global Change Program.
References
Arp, W.J. 1991. Effects of source--sink relations on photosynthetic
acclimation to elevated CO2. Plant Cell Environ. 14:869--875.

Arp, W.J. and B.G. Drake. 1991. Increased photosynthetic capacity of
Scirpus olneyi after 4 years of exposure to elevated CO2. Plant Cell
Environ. 14:1003--1006.
Bailey, R.L., G.E. Grider, J.W. Rheney and L.V. Pienaar. 1985. Stand
structure and yields for site-prepared loblolly plantations in the
piedmont and upper coastal plain of Alabama, Georgia, and South
Carolina. The University of Georgia, Agricultural Experimental
Station Bulletin 328, 118 p.
Ball, J.T. and J.A. Berry. 1982. The Ci/Cs ratio: a basis for predicting
stomatal control of photosynthesis. Carnegie Inst. Washington Yearbook 81:88--92.
Barton, C.V.M., H.S.J. Lee and P.G. Jarvis. 1993. A branch bag and
CO2 control system for long-term CO2 enrichment of mature Sitka
spruce (Picea sitchensis (Bong.) Carr.). Plant Cell Environ.
16:1139--1148.
Bunce, J.A. 1992. Stomatal conductance, photosynthesis and respiration of temperate deciduous tree seedlings grown outdoors at an
elevated concentration of carbon dioxide. Plant Cell Environ.
15:541--549.
Campbell, W.J., L.H. Allen and G. Bowes. 1988. Effects of CO2
concentration on Rubisco activity, amount, and photosynthesis in
soybean leaves. Plant Physiol. 88:1310--1316.
Conroy, J.P., M. Kuppers, B. Kuppers, J. Virgona and E.W.R. Barlow.
1988. The influence of CO2 enrichment, phosphorus deficiency and
water stress on growth, conductance and water use of Pinus radiata
D. Don. Plant Cell Environ. 11:91--98.

358

LIU AND TESKEY

Conroy, J.P., R.M. Smillie, M. Kuppers, D.I. Bevege and E.W. Barlow.
1986. Chlorophyll a fluorescence and photosynthetic and growth
responses of Pinus radiata to phosphorus deficiency, drought stress,
and high CO2. Plant Physiol. 81:423--429.
Cregg, B.M., J.E. Halpin, P.M. Dougherty and R.O. Teskey. 1989.
Comparative physiology and morphology of seedling and mature
forest trees. In Air Pollution Effects on Forest Vegetation. Eds. R.D.
Noble, J.L. Martin and K.F. Jensen. USDA Forest Service, Northeastern Forest Experiment Station, Broomall, PA, pp 111--118.
Davis, J.E., T.J. Arkebauer, J.M. Norman and J.R. Brandle. 1987.
Rapid field measurement of the assimilation rate versus internal
CO2 concentration relationship in green ash (Fraxinus pennsylvanica Marsh.): the influence of light intensity. Tree Physiol.
3:387--392.
Drake, B.G., G.J. Peresta and E. Beugeling. 1994. No decline in effect
of elevated atmospheric CO2 on net ecosystem CO2 exchange
(NCE), biomass production and tissue nitrogen concentration [N]
of a C3 plant community in a Chesapeake Bay wetland after 7 years.
Suppl. Bull. Ecol. Soc. Am. 75(abstr.):57.
Dufrêne, E., J.-Y. Pontailler and B. Saugier. 1993. A branch bag
technique for simultaneous CO2 enrichment and assimilation measurements on beech (Fagus sylvatica L.). Plant Cell Environ.
16:1131--1138.
Eamus, D., C.A. Berryman and G.A. Duff. 1993. Assimilation, stomatal conductance, specific leaf area and chlorophyll responses to
elevated CO2 of Maranthes corymbosa, a tropical monsoon rain
forest species. Aust. J. Plant Physiol. 20:741--755.
Eamus, D. and P.G. Jarvis. 1989. The direct effects of increase in the
global atmospheric CO2 concentration on natural and commercial
temperate trees and forests. Adv. Ecol. Res. 19:1--55.
Ehret, D.L. and P.A. Joliffe. 1985. Photosynthetic carbon dioxide
exchange of bean plants grown at elevated carbon dioxide concentrations. Can. J. Bot. 63:2026-- 2030.
El Kohen, A. and M. Mousseau. 1994. Interactive effects of elevated
CO2 and mineral nutrition on growth and CO2 exchange of sweet
chestnut seedlings (Castanea sativa). Tree Physiol. 14:679--690.
Farquhar, G.D. and T.D. Sharkey. 1982. Stomatal conductance and
photosynthesis. Annu. Rev. Plant Physiol. 33:317--345.
Fetcher, N., C.H. Jaeger, B.R. Strain and N. Sionit. 1988. Long-term
elevation of atmospheric CO2 concentration and the carbon exchange rates of saplings of Pinus taeda L. and Liquidamber styraciflua L. Tree Physiol. 4:255--262.
Graham. R.L., M.G. Turner and V.H. Dale. 1990. How increasing CO2
and climate change affect forests. BioScience 40:575--587.
Gunderson, C.A., R.J. Norby and S.D. Wullschleger. 1993. Foliar gas
exchange responses of two deciduous hardwoods during 3 years of
growth in elevated CO2: no loss of photosynthetic enhancement.
Plant Cell Environ. 16:797--807.
Gunderson, C.A. and S.D. Wullschleger. 1994. Photosynthetic acclimation in trees to rising atmospheric CO2: a broader perspective.
Photosynth. Res. 39:369--388.
Harley, P.C. and T.D. Sharkey. 1991. An improved model of C3
photosynthesis at high CO2: reversed O2 sensitivity explained by
lack of glycerate reentry into the chloroplast. Photosynth. Res.
27:169--178.
Harley, P.C., R.B. Thomas, J.F. Reynolds and B.R. Strain. 1992.
Modelling photosynthesis of cotton grown in elevated CO2. Plant
Cell Environ. 15:271--282.
Higginbotham, K.O., J.M. Mayo, S. L’Hirondelle and D.K.
Krystofiak. 1985. Physiological ecology of lodgepole pine (Pinus
contorta) in an enriched CO2 environment. Can. J. For. Res.
15:417--421.

Houghton, R.A., R.D. Boone, J.R. Fruci, J.E. Hobbie, J.M. Melillo,
C.A. Palm, B.J. Peterson, G.R. Shaver and G.M. Woodwell. 1987.
The flux of carbon from terrestrial ecosystems to the atmosphere in
1980 due to changes in land use: geographic distribution of the
global flux. Tellus 39B:122--139.
Houghton, R.A., J.E. Hobbie, J.M. Melillo, B. Moore, B.J. Peterson,
G.R. Shaver and G.M. Woodwell. 1983. Changes in the carbon
content of terrestrial biota and soils between 1860 and 1980: a net
release of CO2 to the atmosphere. Ecol. Monogr. 53:235--262.
Idso, S.B. and B.A. Kimball. 1991. Downward regulation of photosynthesis and growth at high CO2 levels. Plant Physiol. 96:990--992.
Keeling, C.D., R.B. Bacastow, A.F. Carter, S.C. Piper, T.P. Whorf, M.
Heimann, W.G. Mook and H. Roeloffzen. 1989. A three-dimensional model of atmospheric CO2 transport based on observed
winds. I. Analysis of observational data. In Aspects of Climate
Variability in Pacific and the Western Americas. Ed. D.H. Peterson.
Geophys. Monogr. 55:165--235.
Keeling, C.D. and T.P. Whorf. 1991. Atmospheric CO2----modern
record: Mauna Loa. In Trends ’91: A Compendium of Data on
Global Change. Eds. T.A. Boden, R.J. Sepanski and F.W. Stoss.
Carbon Dioxide Information Analysis Center, Oak Ridge National
Laboratory, Oak Ridge, TN, pp 12--15.
Kohlmaier, G.H., H. Bröhl, E.O. Siré and M. Plöchl. 1987. Modelling
simulation of plants and ecosystem response to present levels of
excess atmospheric CO2. Tellus 39B:155--170.
Makino, A. 1994. Biochemistry of C3-photosynthesis in high CO2. J.
Plant Res. 107:79--84.
McDermitt, D.K., J.M. Norman, J.T. Davis, T.M. Ball, T.J. Arkebauer,
J.M. Welles and S.R. Roemer. 1989. CO2 response curves can be
measured with a field-portable closed-loop photosynthesis system.
Ann. Sci. For. 46(suppl.):416-- 420.
Morison, J.I.L. 1987. Intercellular CO2 concentration and stomatal
response to CO2. In Stomatal Function. Eds. E. Zeigler, G.D.
Farquhar and I.R. Cowan. Stanford University Press, Stanford, CA,
pp 229--252.
Mott, K.A. 1990. Sensing of atmospheric CO2 by plants. Plant Cell
Environ. 13:731--737.
Myers, N. 1993. Gaia: an atlas of planet management. Anchor Books,
Doubleday, New York, NY, 272 p.
Neftel, A., E. Moor, H. Oeschger and B. Stauffer. 1985. Evidence from
polar ice cores for the increase in atmospheric CO2 in the past two
centuries. Nature 315:45--47.
Nobel, P.S., M. Cui, P.M. Miller and Y. Luo. 1994. Influences of soil
volume and an elevated CO2 level on growth and CO2 exchange for
the Crassulacean acid metabolism plant Opuntia ficus-indica.
Physiol. Plant. 90:173--180.
Norby, R.J. and E.G. O’Neill. 1991. Leaf area compensation and
nutrient interactions in CO2-enriched seedlings of yellow-poplar
(Liriodendron tulipifera L.). New Phytol. 117:515--528.
Oberbauer, S.F., B.R. Strain and N. Fetcher. 1985. Effects of CO2-enrichment on seedling physiology and growth of two tropical tree
species. Physiol. Plant. 65:352--356.
Sage, R.F. 1994. Acclimation of photosynthesis to increasing atmospheric CO2: the gas exchange perspective. Photosynth. Res.
39:351--368.
Sharkey, T.D. 1985. Photosynthesis in intact leaves of C3 plants:
physics, physiology and rate limitations. Bot. Rev. 51:53--106.
Silvola, J. and U. Ahlholm. 1992. Photosynthesis in willows (Salix ×
dasyclados) grown at different CO2 concentrations and fertilization
levels. Oecologia 91:208--213.
Stitt, M. 1991. Rising CO2 levels and their potential significance for
carbon flow in photosynthetic cells. Plant Cell Environ. 14:741-762.

GAS EXCHANGE OF LOBLOLLY PINE IN ELEVATED CO2
Sundquist, E.T. 1993. The global carbon dioxide budget. Science
259:934--941.
Tans, P.P., I.Y. Ting and T. Takahashi. 1990. Observational constraints
on the global atmospheric CO2 budget. Science 247:1431--1438.
Taylor, G.E., Jr. and C.A. Gunderson. 1988. Physiological site of
ethylene effects in carbon dioxide assimilation in Glycine max L.
Merr. Plant Physiol. 86:85--92.
Taylor, K.E. and S.L. Grotch. 1990. Observational and theoretical
studies of greenhouse climate effects. In Environmental Problems
and Solutions: Greenhouse Effect, Acid Rain, Pollution. Ed. T.N.
Veziroglu. Hemisphere Publishing Corporation, New York, NY, pp
3--16.
Teskey, R.O., P.M. Dougherty and A.E. Wiselogel. 1991. Design and
performance of branch chambers suitable for long-term ozone fumigation of foliage in large trees. J. Environ. Qual. 20:591--596.
Thomas, R.B., J.D. Lewis and B.R. Strain. 1994. Effects of leaf
nutrient status on photosynthetic capacity in loblolly pine (Pinus
taeda L.) seedlings grown in elevated atmospheric CO2. Tree
Physiol. 14:947--960.
Thomas, R.B. and B.R. Strain. 1991. Root restriction as a factor in
photosynthetic acclimation of cotton seedlings grown in elevated
carbon dioxide. Plant Physiol. 96:627--634.

359

Tissue, D.T., R.B. Thomas and B.R. Strain. 1993. Long-term effects
of elevated CO2 and nutrients on photosynthesis and Rubisco in
loblolly pine seedlings. Plant Cell Environ. 16:859--865.
Tolley, L.C. and B.R. Strain. 1985. Effects of CO2 enrichment and
water stress on gas exchange of Liquidambar styraciflua and Pinus
taeda seedlings grown under different irradiance levels. Oecologia
65:166--172.
von Caemmerer, S. and G.D. Farquhar. 1981. Some relationships
between the biochemistry of photosynthesis and the gas exchange
of leaves. Planta 153:376--387.
Waring, R.H. and W.H. Schlesinger. 1985. Forest ecosystems: concepts and management. Academic Press Inc., Orlando, FL, 340 p.
Whitehead, D., R.O. Teskey and J.F.F. Hobbs. 1992. The response of
large Pinus radiata trees to enhanced carbon dioxide concentration:
what can we learn with a Li-Cor porometer? In Proc. Ann. Meeting
N.Z. Soc. Plant Physiologists. Lincoln University, New Zealand, pp
14--16.
Wofsy, S.C., M.L. Goulden, J.W. Munger, S.-M. Fan, P.S. Bakwin,
B.C. Daube, S.L. Bassow and F.A. Bazzaz. 1993. Net exchange of
CO2 in a mid-latitude forest. Science 260:1314--1317.