Tree Physiology 28, 1421–1429
© 2008 Heron Publishing—Victoria, Canada
Interannual variation in leaf photosynthetic capacity during summer
in relation to nitrogen, leaf mass per area and climate within a Fagus
crenata crown on Naeba Mountain, Japan
ATSUHIRO IIO,1,2 AKIRA YOKOYAMA,1 MASAMITSU TAKANO,1 TETSUROU
NAKAMURA,1 HISAKAZU FUKASAWA,1 YACHIHO NOSE1 and YOSHITAKA KAKUBARI1
1
Faculty of Agriculture, University of Shizuoka, Ohya 836, Shizuoka 422-8529, Japan
2
Corresponding author ([email protected])
Received December 3, 2007; accepted March 19, 2008; published online July 1, 2008
Summary During the summers (July and August) of
2002–2005, we measured interannual variation in maximum
carboxylation rate (Vcmax ) within a Fagus crenata Blume crown
in relation to climate variables such as air temperature, daytime
vapor pressure deficit (VPD) and daily photosynthetic photon
flux, leaf nitrogen per unit area (Na) and leaf mass per unit area
(LMA). Climatic conditions in the summers of 2002–2004
differed markedly, with warm and dry atmospheric conditions
in 2002, cool, humid and cloudy conditions in 2003, and warm
clear conditions in 2004. Conditions in summer 2005 were intermediate between those of summers 2002 and 2003, and similar to recent (8-year) means. In July, marked interannual variation in Vcmax was mainly observed in leaves in the high-light environment (relative photon flux > 50%) within the crown. At
the crown top, Vcmax was about twofold higher in 2002 than in
2003, and Vcmax values in 2004 and 2005 were intermediate between those in 2002 and 2003. In August, although interannual
variation in Vcmax among the years 2003, 2004 and 2005 was
less, marked variation between 2002 and the other study years
was evident. Multiple regression analysis of Vcmax against
the climate variables revealed that VPD of the previous
10–30 days had a significant influence on variability in Vcmax.
Neither Na, LMA nor leaf CO2 conductance from the stomata to
the carboxylation site explained the variability in Vcmax. Our results indicate that the long-term climatic response of Vcmax
should be considered when estimating forest carbon gain
across the year.
Keywords: climate, interannual variability, leaf nitrogen content, maximum carboxylation rate.
Introduction
Long-term flux measurements have shown large interannual
variations in forest carbon gain associated with climatic variability. Saigusa et al. (2005) reported that annual net ecosystem production varied from 59 to 346 g C m –2 over a 9-year period in a cool-temperate deciduous forest in Japan. This
interannual variation in forest carbon gain is due not only to di-
rect climatic effects, but also to indirect climatic effects on
physiological and ecological parameters such as growing season length, stand leaf area index and leaf photosynthetic and
respiratory capacities (Hui et al. 2003). Thus, to obtain accurate, long-term estimates of forest carbon gain, one must incorporate these indirect effects of climate variability in gas exchange models. Saigusa et al. (2005) suggested that the main
indirect factor influencing interannual variations in forest carbon gain is the variability in length of the growing season, and
several other studies in different climatic zones and biomes
reached similar conclusions (e.g., Carrara et al. 2003, Barr et
al. 2004). However, changes not only in growing season length
but in other eco-physiological parameters such as leaf photosynthetic and respiratory capacities and canopy structure, are
expected in response to interannual climate variability (Grassi
et al. 2005, Kitaoka and Koike 2005, Krishnan et al. 2006).
Further, carbon flux measurements alone provide only limited
understanding of the causes of such variation, because of the
difficulty in isolating their numerous causes. To accurately estimate forest carbon gain, an understanding of the responses of
eco-physiological parameters to changes in climate variables
must be acquired at the single-leaf scale, in parallel with flux
measurements.
Leaf photosynthetic capacity is a major factor affecting forest carbon gain (e.g., Ito et al. 2005), and numerous studies
have measured spatial and seasonal variations in leaf photosynthetic capacity in relation to possible limiting factors, such
as leaf nitrogen content, leaf morphology and leaf environmental conditions (e.g., Wilson et al. 2000, Han et al. 2004,
Niinemets et al. 2004, Iio et al. 2005). However, there have
been few corresponding studies on interannual variations.
Grassi et al. (2005) showed that soil drought could cause twofold interannual variations in the maximum carboxylation rate
(Vcmax ) of two deciduous tree species in a sub-Mediterranean
climate, even within a season, by an effect on the photosynthetic apparatus. In addition, Kitaoka and Koike (2005) reported large interannual variations in leaf maximum photosynthetic rates in seedlings of four deciduous tree species in a cool
temperate climate that were related to differences in air tem-
1422
IIO ET AL.
perature and precipitation during leaf development. Thus,
interannual variation in leaf photosynthetic capacity, resulting
from year-to-year fluctuations in climate variables, may have
considerable effects on forest carbon gain.
In the region where our study was conducted, marked interannual climate variations occurred during the summer (July
and August) of three consecutive years: 2002–2004. In 2002,
summer days were mostly warm and dry, although there was
sufficient precipitation to avoid drought inhibition of photosynthesis. By contrast, in the summer of 2003, there were prolonged periods of cool, humid and cloudy weather. In the summer of 2004, there were frequent warm clear days although the
air was not as dry as in 2002. This year-to-year climate variability provided the opportunity to study the responses of leaf
photosynthetic characteristics of Fagus crenata Blume, a typical deciduous tree species in Japanese mountain areas.
In a previous study, we measured spatial variations in Vcmax
in relation to relative photosynthetic photon flux (rPPF), leaf
nitrogen per unit area (Na) and leaf mass per unit area (LMA)
within an F. crenata crown in mid-July to scale leaf photosynthetic parameters to canopy-level carbon gain (Iio et al. 2005).
In this study, we obtained long-term estimates of canopy carbon gain over 4 years (2002–2005). To analyze the effects of
interannual climatic variability on Vcmax, seasonal measurements of Vcmax were made at the crown top, especially during
the summer. Our primary objective was to characterize the
interannual variability in Vcmax in relation to climate variables
such as air temperature, daytime vapor pressure deficit and
irradiance, and to Na and LMA.
Materials and methods
Study site and plant material
The study site is located in a 70-year-old second-growth Fagus
crenata forest in a cool temperate climate zone 900 m above
sea level on the eastern slopes of Mt. Naeba, Japan (36°55′ N,
138°46′ E). A 20 × 30 m plot was set up in the forest and a
22-m-tall steel tower constructed in the center of the plot provided access to the crown of the selected tree, which was
21.5 m in height, with a diameter at breast height of 26 cm, a
height of 8 m to the lowest leaves and a crown radius of 2.0 m.
The sample tree is representative of other dominant trees in the
stand in leaf photosynthetic capacity at the crown top, leaf
phenology and crown structure parameters, such as crown projection area, tree height, diameter at breast height and vertical
distribution of leaf area (data not shown). Bud break in the
years 2002–2005 occurred on day of the year (DOY) 111,
118, 119 and 121, respectively, and defoliation was complete
on DOY 313, 303, 317 and 316, respectively. In 2005, a heavy
masting event occurred at the study site, and the selected tree
produced fruit. Climate variables, such as photosynthetic photon flux (PPF), air temperature (Tair), relative humidity and
wind speed, have been measured continuously at the tower top
since 1998. Soil volumetric water content (SWC) and soil temperature (Tsoil ) were measured near the selected tree at 25-cm
depth. All measurements were made at 5-s intervals, and data
were stored as 30-min means by a data logger (DL2e, Delta-T,
Cambridge, U.K.). In 2006, soil water potential (Ψsoil ) was
measured with a tensiometer (KDC-55, Kona, Sapporo, Japan) inserted near the SWC sensor, at 25-cm depth. The relationship between SWC and Ψsoil was determined and used to
calculate Ψsoil throughout the growing season of each study
year. Mean Tair and cumulative precipitation during the growing seasons (May to October: DOY 122–305) of the study
years were 17.0 °C and 1343.1 mm, respectively. Further details about the selected tree and stand have been described by
Iio et al. (2005) and Kubota et al. (2005).
Crown light measurements
We monitored PPF within the crown with 20 quantum sensors
(IKS-25, Koito, Tokyo, Japan) positioned horizontally to be as
representative as possible of the range of PPF gradients. The
measuring and storing intervals were the same as for the climate measurements at the tower. All sensors were calibrated
on a clear day from darkness to full sunlight against an LI-190
quantum sensor (LI-190-SA, Li-Cor, Lincoln, NE). In 2002,
within-crown PPF was measured for 20 days from July 1 to 20,
whereas from 2003 to 2005, measurements were taken continuously throughout the growing season.
Leaf gas exchange
Response curves of photosynthesis (P) to intercellular CO2
concentration (P/Ci ) were measured in situ on three to eight
leaves adjacent to the quantum sensors within the crown. The
total numbers of measurement locations were 13, 16, 12 and 6
in 2002–2005, respectively. Measurements were made in
mid-July (DOY 203, 194, 206 and 199 in 2002–2005, respectively) because mid-July represents the period after leaf maturity but precedes leaf senescence. The P/Ci curves were measured at 8–10 CO2 concentrations from 0 to 2000 µmol mol –1
with a Li-Cor LI-6400 system under controlled light, temperature and humidity conditions. The PPFs were selected to provide saturating light without causing photoinhibition; all foliage was measured at 1000 µmol m –2 s –1, except for that in the
darkest part of the crown (8 m above ground), which was measured at 700 µmol m –2 s –1. Leaf temperature (Tleaf) and leaf to
air vapor pressure deficit (LAVPD) were kept at 25 ± 0.5 °C
and < 1.2 kPa, respectively, during these measurements. To
quantify leaf internal CO2 conductance (gi ) from the P/Ci
curves (Harley et al. 1992), gas exchange and chlorophyll fluorescence were measured simultaneously with a Li-Cor
LI-6400-40 leaf chamber fluorometer.
Many measurements are needed to characterize interannual
variations in Vcmax . The one point method suggested by Wilson
et al. (2000) allowed us to estimate Vcmax from the net photosynthetic rate (Pn ) measured under ribulose-1,5-bisphosphate
(Rubisco)-limiting conditions (Ci < 250 µmol mol –1 ) and dark
respiration (Rd ). Accordingly, light-saturated Pn (Pnmax ) of five
leaves at the crown top was measured at ambient CO2 concentration at weekly or monthly intervals throughout the growing
season in 2002 and 2003, whereas in 2004 and 2005, most
measurements were taken during the summer. The LAVPD
TREE PHYSIOLOGY VOLUME 28, 2008
INTERANNUAL PHOTOSYNTHETIC VARIATIONS IN A FAGUS CRENATA CROWN
and temperature in the chamber were kept at < 1.5 kPa and
close to ambient temperature (20–30 °C), respectively.
To avoid photoinhibition and stomatal patchiness, which is
often associated with low values of stomatal conductance that
may cause erroneous estimates of Ci (Takanashi et al. 2006),
all field measurements were made in the early morning or on
cloudy days. To detect symptoms of stomatal patchiness, diurnal changes in P/Ci curves and Pnmax were measured under ambient conditions for five to ten leaves at the crown top. This
preliminary survey was carried out at the end of July for each
study year and indicated that decreases in Vcmax (which would
have been a symptom of stomatal patchiness) did not occur until stomatal conductance of H2O decreased below 150 mmol
m –2 s –1 (i.e., until LAVPD increased above 2.0 kPa; data not
shown). Therefore, all field measurements were conducted
above this threshold.
To standardize Vcmax obtained at ambient temperature to
25 °C (Vc25 ), temperature dependencies of the P/Ci curves
were measured in May and July in 2004, and July, August,
September and October in 2005 with a detached branch from
the crown top. The P/Ci curves were measured at five temperatures ranging from 10 to 35 °C with three replications. The
PPF and LAVPD were maintained at 1000 µmol m –2 s –1 and
< 2.5 kPa (and < 2.0 kPa below 30 °C), respectively.
where Tk is leaf temperature (°C). The temperature dependencies of Kc and Ko are described by:
f ( T k ) = f ( 298) e
Ha
( Tk − 298 )
298 R Tk
After the P/Ci curve measurements in mid-July, all monitored
leaves were harvested and excised at the petiole to determine
their area, dry mass and nitrogen concentration. The area of
each leaf was measured with a video area meter (DIAS,
Delta-T, Cambridge, U.K.). Each leaf was then dried at 80 °C
for 48 h and weighed to calculate LMA. Total nitrogen concentration on a dry mass basis (Nm ) was determined by gas
chromatography (GC-8A, Shimadzu, Kyoto, Japan) after
combustion with circulating O2 using an NC analyzer
(Sumigraph NC-95A, SCAS, Osaka, Japan). Values of Na
were calculated from Nm and LMA.
Photosynthetic parameters
We calculated Vcmax and maximum electron transfer rate (Jmax )
from the P/Ci curves following the procedures of Poot et al.
(1996) based on the model of Farquhar et al. (1980). Both
Vcmax and daytime respiration (Rd∗ ) were estimated from the
P/Ci curves at Ci < 250 µmol mol –1, assuming that the only
factors limiting Pn at low Ci were the amount, activity and kinetic properties of Rubisco. We estimated Jmax from the P/Ci
curves at Ci > 500 µmol mol –1, with the Rd∗ value. The CO2
compensation point in the absence of Rd∗ (Γ ∗ ) was estimated
based on Tleaf conversions according to Brooks and Farquhar
(1985) and Farquhar et al. (1980), and the Michaelis-Menten
constants for CO2 (Kc) and O2 (Ko ) according to von
Caemmerer et al. (1994). Values for Γ ∗, Kc and Ko at 25 °C
were 4.47 Pa, 40.4 Pa and 24.8 kPa, respectively. The temperature dependence of Γ∗ is described by:
Γ ∗ = 44.7 + 188
. ( T k − 25 ) + 0.036( T k − 25 )2
(1)
(2)
where ƒ(Tk ) is the value of a given parameter at leaf temperature Tk, ƒ(298) is the value of the parameter at 25 °C, Ha is activation energy where Ha(Kc) = 59.356 kJ mol –1 and Ha(Ko ) =
35.948 kJ mol –1 at 25 °C, and R is the gas constant (0.00831 kJ
mol –1 ).
The one point method estimates Vcmax by running the
Farqhuar model with only the parameters Pnmax, Ci and Rd measured at ambient CO2 concentration, with Rd assumed to be
constant at 0.5 µmol m –2 s –1. There was a strong relationship
between Vcmax estimated by the one point method and Vcmax estimated from the P/Ci curve, irrespective of season and year
(y = 0.946x + 2.60, r 2 = 0.97, P < 0.0001).
The temperature responses of Vcmax were fitted by a peaked
function described by Harley and Tenhunen (1991):
H ⎞
⎛
⎜c− a ⎟
R Tk ⎠
e⎝
Parameter( Vcmax ) =
1+ e
Leaf mass per unit area and leaf nitrogen content
1423
Δ S Tk − Hd
R Tk
(3)
where Hd is deactivation energy, c is a scaling constant and ΔS
is an entropy term (0.65 kJ K –1 mol –1 ). We converted Vcmax estimated by the one point method to Vc25 based on the temperature response curves normalized to 1 at 25 °C.
We estimated gi according to the constant J method (Harley
et al. 1992). The ratio of the CO2 partial pressure at the
carboxylation site to that in intercellular spaces (Cc /Ci ) was
calculated as:
Pn =
(Ci − Cc ) gi
A
(4)
where Cc is CO2 partial pressure at the carboxylation site and A
is atmospheric pressure. Details of the calculations have been
described by Iio et al. (2005).
Statistical analysis
The light environment within the crown was expressed as the
mean monthly PPF for the respective foliage relative to the simultaneous value of PPF above the crown (rPPF). For each
study year, the interrelationships between light environment
and leaf morphological, biochemical and physiological characteristics within the crown were analyzed by linear regression. Because some relationships between rPPF and leaf properties were curved, with inflection points at around rPPF =
30% (data not shown), we linearized these relationships by
log-transforming rPPF. To characterize interannual variations
in leaf properties within the crown, the parameters of the linear
regression were tested by covariance analysis (ANCOVA, P <
0.05). Initially, the differences in slopes were tested, and only
when there was no significant slope difference, we tested for
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
IIO ET AL.
Mean weekly climate variables during the growing seasons
(DOY 122–305) of years 2002–2005 are shown in Figure 1.
Marked interannual variations in climatic conditions were observed during the summer (July and August; DOY 183–244),
especially in July (DOY 183–213), the most pronounced differences were between 2002 and 2003. Values of Tair and VPD
in July 2002 were generally high, with mean monthly daily
photosynthetic photon flux (PPFday ), Tair, VPD and Tsoil of
30.9 mol m –2 day –1, 22.0 °C, 0.73 kPa and 16.5 °C, respectively, compared with recent 8-year (1999–2006) means of
31.3 mol m –2 day –1, 20.6 °C, 0.50 kPa and 15.3 °C, respectively. However, soil water potential (Ψsoil ) was never less than
–30 kPa (severe drought; Kubota et al. 2005) except at the end
of the month (DOY 209) because of frequent rain events. By
contrast, July 2003 was characterized by cool, cloudy humid
conditions. Mean monthly PPFday, Tair and VPD were 25.4 mol
m –2 day –1, 18.0 °C and 0.26 kPa, respectively; the lowest values of recent 8-year measurements. Rain events were frequent
(29 days in July 2003 compared with 20 days in July 2002),
and accordingly, Ψsoil and relative humidity remained high
throughout the month. July 2004 was characterized by high
PPFday and high Tair, with mean monthly PPFday, Tair, VPD and
Tsoil of 35.5 mol m –2 day –1, 21.5 °C, 0.50 kPa and 17.1 °C, respectively. Although mean monthly Tair in July 2004 was only
0.5 °C below that in July 2002, mean monthly VPD in July
2004 was 68% of that in July 2002. Climatic conditions in July
2005 were intermediate between those recorded in 2002 and
2003, with mean monthly PPFday, Tair, VPD and Tsoil of 27.2
mol m –2 day –1, 19.6 °C, 0.42 kPa and 15.6 °C, respectively. In
July 2004 and 2005, no severe soil drought occurred.
In August (DOY 214–244), interannual differences in Tair
and Tsoil were less pronounced than in July, but the atmosphere
was generally dry in 2002 and humid and cloudy in 2003.
Therefore, mean monthly VPD values were markedly lower in
August 2003 than in August 2002 (0.39 versus 0.79 kPa). No
severe soil drought occurred in August in any study year.
PPFday (mol m – 2 day – 1 )
Tair (°C)
Interannual variations in climate variables
Seasonal and interannual variations in Vc25 are shown in Figure 2. As with the climate variables, marked interannual variation in Vc25 was mainly observed in July, with the greatest variation observed between 2002 and 2003. Values of Vc25 were
1.5–2.1-fold higher in 2002 than in 2003, and Vc25 values in
2004 and 2005 were intermediate between those in 2002 and
2003. Although there was no marked interannual variation in
August Vc25 among the years 2003, 2004 and 2005, there were
50
40
July
A
August
2002
2003
2004
2005
30
20
10
0
25
B
20
15
10
5
20
C
Tsoil (°C)
Results
Interannual variation in Vc25 at the crown top and climate
variables
15
10
5
0
1.2
VPD (kPa)
an intercept difference. The Bonferroni method was used for
post-hoc multiple comparisons of slopes and intercepts. The
determinant coefficients of linear regression (r 2 ) were analyzed with SigmaPlot.
For Vcmax data measured in July and August at the crown top
(see Figure 2), mean daily PPF (PPFday ), Tair, daytime vapor
pressure deficit (VPD, daily mean between 0600 and 1800 h),
Tsoil and Ψsoil for 1, 3, 7, 10, 20 and 30 days before each Vcmax
measurement were calculated, and tested as climatic factors
causing the interannual variability in Vcmax by linear regression
analysis. To exclude possible bias due to leaf aging, each
month, data were analyzed separately. Then, effects of climate
factors on Vcmax were analyzed by multiple regression. A
stepwise method identified climate variables with inclusion
and exclusion threshold probabilities of P = 0.05.
1.0 D
0.8
0.6
0.4
0.2
0.0
0
-10
E
-20
-30
-40
Precipitation (mm)
1424
-50
250
200 F
150
100
50
0
90
120
150
180
210
240
270
300
330
DOY
Figure 1. Seasonal and interannual variations in weekly means of climatic variables, soil temperature and soil water potential at the Fagus
crenata study site on Mt. Naeba: (A) daily photosynthetic photon flux
(PPFday ) at the crown top; (B) air temperature (Tair ) at the crown top;
(C) soil temperature (Tsoil ) at 25 cm depth; (D) daytime (0600–
1800 h) vapor pressure deficit (VPD) at the crown top; (E) soil water
potential (Ψsoil ) at 25 cm depth; and (F) weekly cumulative precipitation.
TREE PHYSIOLOGY VOLUME 28, 2008
INTERANNUAL PHOTOSYNTHETIC VARIATIONS IN A FAGUS CRENATA CROWN
120
marked differences between 2002 and the three other study
years.
For Vc25 data in July and August (Figure 2), the correlations
of mean climate variables over the 1–30 days before the Vc25
measurements were analyzed by linear regression (Table 1). In
July, mean VPD over the prior 30 days (VPD30 ) had a strong
(r 2 > 0.50) and positive influence on Vc25. Mean VPD over the
prior 7, 10 (VPD10 ) and 20 days and mean Ψsoil over the prior
20 days (Ψsoil20 ) had a weak but significant influence on Vc25.
In August, not only VPD but also Ψsoil and Tair strongly influenced Vc25. Although VPD was positively correlated with Vc25
as in July, Ψsoil was negatively correlated in August in contrast
to July. Mean PPFday over the prior 10 days was weakly correlated with Vc25. Soil temperature had no significant influence
on Vc25 in either month.
Climate variables were significantly correlated with each
other. For example in August, VPD10 was significantly correlated with mean Tair (r 2 = 0.80), PPFday (r 2 = 0.51) and Ψsoil
(r 2 = 0.45) over the prior 10 days. To separate the effects of the
climate variables on Vc25, we conducted multiple-regression
analysis of Vc25 with the climate variables that were significantly correlated with Vc25 in linear regression analysis. In
both months, the best model was a simple linear regression of
VPD (VPD30 and VPD10 in July and August, respectively; regression parameters are shown in Table 1).
July
1425
August
2002
2003
2004
2005
80
60
P
Vc25 (µmol m – 2 s – 1 )
100
40
20
0
90
120
150
180
210
240
270
300
330
DOY
Figure 2. Seasonal changes in the maximum carboxylation rate standardized to 25 °C (Vc25 ) at the crown top of a Fagus crenata tree over
the years 2002–2005. Values are means of three to five replicates.
Values of Vc25 estimated both from P/Ci curves and by the one point
method are shown.
(Figure 3A, Table 2). When Na and LMA were substituted for
log(rPPF), similar results were obtained (Figures 3B and 3C).
In each case, the ranking of the steepness of the slopes of these
regressions was 2002 > 2005 > 2004 > 2003. Interannual variations in Vc25 were less pronounced under low rPPF, Na and
LMA conditions (log(rPPF) < 4.0, Na < 1.5 g m –2 and LMA <
70 g m –2 ). There were no significant interannual differences in
the slopes of the Vc25 /J25 regressions (Figure 3D). Although
significant differences in the intercepts of the Vc25 /J25 regressions were detected, the effect of the variation (i.e., difference
in Vc25 at a given J25 ) was unclear. Therefore, when the linear
regressions were analyzed by substituting J25 for Vc25, similar
Interannual variations in leaf physiological, morphological
and biochemical crown characteristics
The dependences of Vc25 on log(rPPF), Na, LMA and J25
within the crown are shown in Figure 3. Although significant
and positive linear relationships between log(rPPF) and Vc25
were observed in each study year, there were significant
interannual differences in the slopes of the regression lines
Table 1. Linear regression parameters (y = ax + b) and the coefficient of determination (r 2 ) for the maximum carboxylation rate standardized to
25 °C (Vc25 ) measured at the crown top in July and August of 2002–2005 (Figure 2) versus: air temperature (Tair ), daily photosynthetic photon
flux (PPFday ), daytime vapor pressure deficit (VPD), soil temperature (Tsoil ) and soil water potential (Ψsoil ), averaged over the previous 1, 3, 7, 10,
20 and 30 days to each Vc25 measurement. Significant linear regressions are indicated by asterisks: *, P < 0.05; **, P < 0.01; and ***, P < 0.001.
Period
(days)
Tair
a
July (n = 18)
1
2.45
3
3.45
7
2.41
10
0.32
20
0.13
30
–1.77
PPFday
b
17.3
–2.59
20.5
62.6
66.5
103
August (n = 15)
1
0.97
41.6
3
1.47
30.1
7
8.94 –137
10
9.99 –161
20
5.67 –61.8
30
4.88 –41.8
r2
0.13
0.20
0.08
0.00
0.00
0.04
0.02
0.02
0.31 *
0.55 **
0.39 *
0.35 *
VPD
b
r2
a
b
r2
–0.14
0.03
0.37
0.23
–0.68
0.58
73.5
68.1
57.9
62.3
87.8
52.1
0.02
0.00
0.01
0.00
0.03
0.02
0.34
1.96
4.27
4.17
6.08
15.7
67.2
59.6
50.0
51.5
45.3
4.00
0.01
0.13
0.29 *
0.23 *
0.23 *
0.74 ***
0.06
0.16
0.52
1.08
1.30
1.26
60.8
57.5
44.9
25.5
18.9
22.7
0.01
0.02
0.10
0.34 *
0.24
0.14
0.23
0.88
4.06
4.91
4.96
4.26
61.3
57.3
35.9
29.5
31.1
38.1
0.01
0.05
0.36 *
0.68 ***
0.60 **
0.48 **
a
Ψsoil
Tsoil
b
r2
a
b
r2
57.0
74.5
63.7
64.2
76.5
87.0
0.01
0.10
0.00
0.00
0.01
0.03
0.32
0.44
0.42
0.53
1.28
0.55
72.7
74.5
74.8
75.9
87.5
77.9
0.07
0.10
0.10
0.12
0.26 *
0.07
2.66 13.4
2.26 20.8
2.63 14.4
3.47 –0.41
3.51 0.48
2.94 12.2
0.06
0.04
0.06
0.12
0.17
0.12
–0.45
–0.49
–0.66
–0.75
–0.98
–1.08
55.6
54.4
50.4
48.8
47.9
48.1
0.28 *
0.41 *
0.66 ***
0.67 ***
0.46 **
0.28 *
a
0.74
0.44
0.34
0.30
–0.50
–1.25
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
1426
IIO ET AL.
Vc25. However, this conclusion may be associated with correlations and interactions of the climate variables themselves because VPD30 and Ψsoil20 were significantly correlated (r 2 =
0.30, P = 0.018). Severe soil drought causes stomatal and
non-stomatal limitations of photosynthesis, and a down-regulation of PSII activity, which may last for a few weeks even after re-watering (e.g., Galle and Feller 2007). In our study, there
were no marked interannual differences in Ci /Ca throughout
the crown, indicating no stomatal or non-stomatal limitation of
photosynthesis (Figure 4C). Furthermore, the maximum
quantum efficiency of PSII (Fv /Fm ) at the crown top was fairly
constant at around a non-stressed value of 0.83 (Björkman and
Demmig 1987) throughout the study (data not shown), which
supports the conclusion that there was no drought-induced
stress. In August, although there were significant correlations
of mean Ψsoil and Vc25 irrespective of the averaging period, the
slopes of the regressions were negative (i.e., Vc25 decreased
with decreasing drought severity; Table 1). At our study site,
interannual variation in Vc25 seemed to occur through a different pathway than observed in previous studies.
Multiple regression analysis showed that, in August as in
July, the preceding day’s VPD had the largest effect on Vc25.
However, PPFday and Tair also affected Vc25 (Table 1) and were
significantly correlated with each other (r 2 = 0.26~0.80, P <
0.05). However, our conclusion of a minor effect of PPFday is
results were obtained (data not shown).
Like Vc25, Na and LMA showed positive, linear relationships
with log(rPPF) in each study year (Figures 4A and 4B). There
was no significant interannual variation in the log(rPPF)/Na relationship. Although significant interannual variation in the intercept of the log(rPPF)/LMA relationships were detected,
only small differences in LMA were apparent compared with
those in the log(rPPF)/Vc25 relationship. The Ci /Ca ratio was
negatively related to log(rPPF) in each study year, whereas
Cc /Ci was unrelated to log(rPPF), except in 2003 (Figures 4C
and 4D). Although significant interannual differences in the
slope of the log(rPPF)/Ci /Ca relationship and in mean values
of Cc /Ci, were detected, especially between 2002 and 2003
(Cc /Ci; ANOVA P = 0.01), the effect of the variations was minor when log(rPPF) was greater than 4.0 (i.e., rPPF > 50%).
Discussion
For many deciduous tree species, marked interannual variability in the photosynthetic capacity of sunlit leaves is mainly
caused by changes in the severity of soil drought (Wilson et al.
2000, Leuschner et al. 2001, Grassi et al. 2005). In our study,
although Ψsoil20 had a significant influence on Vc25 in July (Table 1), multiple regression analysis showed that VPD30 was the
only climate factor explaining the interannual variability in
100
B
A
90
Vc25 (µmol m – 2 s – 1 )
80
70
60
50
40
2002
30
2003
20
2004
10
0
2005
0
1
2
3
4
0.0
0.5
1.0
1.5
2.0
2.5
Na (g m – 2 )
log(rPPF)
100
90
C
D
Vc25 (µmol m – 2 s – 1 )
80
70
60
50
40
30
20
10
0
0
20
40
60
80
–2
LMA (g m )
100
0
50
100
150
–2
200
–1
J25 (µmol m s )
TREE PHYSIOLOGY VOLUME 28, 2008
Figure 3. Dependence of the maximum
carboxylation rate standardized to 25 °C
(Vc25 ) on (A) log-transformed relative
photosynthetic photon flux (log(rPPF)),
(B) leaf nitrogen content per unit area
(Na ), (C) leaf mass per unit area (LMA)
and (D) maximum electron transport rate
standardized to 25 °C (J25 ) within the
crown of a Fagus crenata tree. Measurements were made in mid-July, and values
are means of three to eight replicates.
Lines show regression equations fitted to
the data for each year. Regression results
are listed in Table 2.
INTERANNUAL PHOTOSYNTHETIC VARIATIONS IN A FAGUS CRENATA CROWN
consistent with abundant experimental evidence that plasticity
of the photosynthetic apparatus in response to shifts in PPFday
is generally small for mature leaves (e.g., Frak et al. 2001,
Niinemets et al. 2004, Muller et al. 2005). Furthermore, neither an increase in the ratio of chlorophyll to nitrogen nor a de-
Table 2. Linear regression results (y = ax + b) for the data shown in
Figures 3 and 4. Different letters indicate significant differences in regression parameters among years (P < 0.05, ANCOVA). Abbreviations: Vc25 , maximum carboxylation rate standardized to 25 °C; rPPF,
relative photosynthetic photon flux; Na, leaf nitrogen per unit area;
LMA, leaf mass per unit area; J25 , maximum electron transport rate
standardized at 25 °C; Ci , Ca and Cc, CO2 partial pressure in the
intercellular space, in ambient air and in the chloroplast, respectively.
Year
a
r2
b
P
n
Vc25 = alog(rPPF ) + b
2002
23.40 a
2003
12.67 c
2004
14.66 bc
2005
19.25 ab
–24.73
3.14
2.12
–16.80
0.83
0.66
0.82
0.75
< 0.0001
< 0.0001
< 0.0001
< 0.0001
56
48
37
18
Vc25 = aNa + b
2002
49.10 a
2003
30.48 b
2004
35.03 b
2005
36.85 b
–27.40
–4.94
–6.11
–10.49
0.62
0.85
0.72
0.88
< 0.0001
< 0.0001
< 0.0001
< 0.0001
52
47
37
18
Vc25 = aLMA + b
2002
1.05 a
2003
0.63 c
2004
0.72 bc
2005
0.85 ab
–15.27
3.07
–0.46
–2.53
0.51
0.80
0.66
0.83
< 0.0001
< 0.0001
< 0.0001
< 0.0001
56
48
37
18
J25 = aVc25 + b
2002
1.94 a
2003
2.15 a
2004
2.15 a
2005
1.97 a
5.17 a
–13.44 c
–8.92 ab
–8.47 bc
0.89
0.88
0.85
0.90
< 0.0001
< 0.0001
< 0.0001
< 0.0001
55
45
36
18
0.01 a
0.24 a
0.24 a
–0.17 a
0.71
0.75
0.82
0.90
< 0.0001
< 0.0001
< 0.0001
< 0.0001
52
47
37
18
LMA = alog(rPPF ) + b
2002
22.28 a
–9.01 a
2003
20.19 a
0.11 ab
2004
20.32 a
3.33 b
2005
22.72 a
–16.85 c
0.70
0.79
0.81
0.88
< 0.0001
< 0.0001
< 0.0001
< 0.0001
56
48
37
18
Ci /Ca = alog(rPPF ) + b
2002
–0.07 a
2003
–0.04 c
2004
–0.04 bc
2005
–0.06 ab
0.97
0.86
0.91
0.97
0.62
0.46
0.39
0.40
< 0.0001
< 0.0001
< 0.0001
0.0052
56
48
38
18
Cc /Ci = alog(rPPF) + b
2002
0.01
2003
–0.05
2004
0.00
2005
–0.05
0.50
0.84
0.64
0.74
0.01
0.18
0.00
0.17
0.578
0.017
0.914
0.141
38
32
28
14
Na = alog(rPPF ) + b
2002
0.49 a
2003
0.42 a
2004
0.42 a
2005
0.52 a
1427
crease in the ratio of chlorophyll a to b was observed in the
cloudy summer of 2003 compared with the corresponding ratios in the fine summers of 2002 and 2004 (data not shown),
supporting this conclusion. Regarding the acclimation of the
photosynthetic apparatus to Tair, several studies (e.g., Medlyn
et al. 2002, Muller et al. 2005, Yamori et al. 2005) have shown
that Vc25 increases with decreasing Tair because the amounts of
photosynthetic components such as Na and ribulose-1,5bisphosphate (Rubisco) increase to compensate for low
Rubisco activity (Berry and Björkman 1980) or to protect
against photoinhibition, or both (Kato et al. 2003). In our
study, however, Vc25 increased with increasing Tair, implying a
minor effect of Tair.
Based on the results of multiple regression analysis, we suggest that the preceding day’s VPD is the most important climate factor accounting for interannual variability in Vc25.
However, this VPD effect cannot be easily explained because
little is known about the acclimation of the photosynthetic apparatus to growing-season VPD, especially under well-watered conditions. For Nothofagus cunninghamii (Hook.) Oerst.
seedlings in a cool temperate climate, Cunningham et al.
(2005) reported no effect of growing-season VPD on leaf photosynthetic capacity. Furthermore, for mature Fagus sylvatica L. in a cool-temperate climate, Leuschner et al. (2001)
showed that interannual differences in summer VPD have no
marked influence on leaf photosynthetic capacity. Our result
contradicts these findings even though the genus (Fagus) and
climate zone were the same.
One possible explanation for this inconsistency is the effect
of fungal infection during wet humid conditions. In early September 2003, many beech rust (Pucciniastrum fagi ) aecia
were observed on leaves in the high-light environment within
the crown. No aecia were observed in 2002, and only a few
were observed in early September 2004 and 2005 (unpublished data). During late spring, airborne basidiospores of
Pucciniastrum fagi, formed on needles of an intermediate host
plant, Tsuga spp., penetrate into young F. crenata leaves
(Kaneko and Hiratsuka 1980, Kaneko and Sahashi 1998). Although the photosynthetic responses of F. crenata to
Pucciniastrum attack have not been investigated, a decrease in
photosynthetic capacity as a result of a rust infection is well
documented in herbaceous plants (e.g., Bethenod et al. 2001)
and Norway spruce (Picea abies (L.) Karst.; Bauer et al. 2000,
Mayr et al. 2001). Thus, the observed variability in Vc25 may
reflect differences in the extent of rust infection.
Apart from climate effects, a heavy masting event—which
changes the source–sink relationship (Hoch 2005) and may affect leaf physiological, biochemical and morphological characteristics (e.g., Urban et al. 2003)—was observed on the selected tree in 2005. However, there were no marked differences in Na and LMA in the crown between 2005 and the other
study years (Figures 4A and 4B). Furthermore, the predominant effect of VPD during the summer did not change when
data for 2005 were excluded from the regression analysis (results not shown). Thus, we conclude that the contribution of
the masting event to interannual variability in Vc25 is minor.
Neither Na nor LMA, both of which are often used as predic-
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
1428
IIO ET AL.
120
2.5
B
A
100
LMA (g m – 2 )
Na (g m – 2 )
2.0
1.5
1.0
20
0.0
1.0
0
1.0
C
0.9
0.8
0.8
0.7
0.7
0.6
0.6
Cc /Ci
Ci /Ca
0.9
60
40
2002
2003
2004
2005
0.5
80
0.5
0.4
0.5
0.4
0.3
0.3
0.2
0.2
0.1
0.1
0.0
0
0.0
0
1
2
3
4
5
D
1
log(rPPF)
2
3
4
log(rPPF)
tors of spatio-temporal variation in Vc25, explained the
interannual variation in Vc25 (Figures 3B and 3C), as has been
observed for other deciduous tree species (Wilson et al. 2000,
Grassi et al. 2005). The absence of marked interannual variations in Na and LMA throughout the crown of our study tree
(Figures 4A and 4B) indicates that photosynthetic nitrogen-use efficiency (PNUE; Vc25 /Na) varied interannually. No
marked interannual variations in Ci /Ca and Cc /Ci were observed especially in the high-light environment within the
crown (Figures 4C and 4D), suggesting that leaf CO2 conductance from the stomata to the carboxylation site does not contribute to the variability in PNUE, which contrasts with the
findings for severe soil drought (Grassi and Magnani 2005).
Differences in other physiological factors such as nitrogen allocation to photosynthetic proteins and rates of Rubisco activity (e.g., Hikosaka et al. 1998) might have caused the variability in PNUE.
In conclusion, large interannual variation in Vc25 was observed during the summer in the high-light portion of the
crown, as reported at other F. crenata study sites on Naeba
Mountain (Wang et al. 2008). Statistical analysis showed that
VPD of the previous 10–30 days significantly influenced the
variability in Vc25. Although photosynthesis is affected by various climate variables on a short-term basis, our results imply
that an understanding of the long-term photosynthetic response to climate is necessary to accurately estimate forest
carbon gain. However, a 4-year study period may be too short
to identify the long-term photosynthetic response to climate
variables. Further continuous study is needed to determine the
general climatic response of the leaf properties of F. crenata.
5
Figure 4. Relationship between log-transformed relative photosynthetic photon flux
(log(rPPF)) and (A) leaf nitrogen content
per unit area (Na ), (B) leaf mass per unit
area (LMA), (C) the ratio of CO2 partial
pressure in the intercellular space to that
in ambient air (Ci /Ca ) and (D) the ratio of
CO2 partial pressure in the chloroplast to
that in the intercellular space (Cc /Ci )
within the Fagus crenata crown.
Measurements were made in mid-July,
and values are means of three to eight
replicates. Lines show linear regressions
fitted to the data for each year. Regression
results are listed in Table 2.
Acknowledgments
This study was part of a wider project evaluating the total CO2 budget
in a forest ecosystem, coordinated by the Ministry of Agriculture,
Forestry, and Fisheries of Japan, and was supported by a Grant-in-Aid
for Scientific Research (No. B 13460067, 14656062), provided by the
Japan Society for the Promotion of Science. We thank Dr. Q. Han of
the Forestry and Forest Products Research Institute (FFPRI), Assoc.
Prof. Q. Wang and Prof. H. Mizunaga of Shizuoka University for
valuable comments, and students of Shizuoka University for assistance with measurements.
References
Barr, A.G., T.A. Black, E.H. Hogg, N. Kljun, K. Morgenstern and
Z. Nesic. 2004. Interannual variability in the leaf area index of a boreal aspen–hazelnut forest in relation to net ecosystem production.
Agric. For. Meteorol. 126:237–255.
Bauer, H., K. Plattner and W. Volgger. 2000. Photosynthesis in Norway spruce seedlings infected by the needle rust Chrysomyxa
rhodedendri. Tree Physiol. 20:211–216.
Berry, J. and O. Björkman. 1980. Photosynthetic response and adaptation to temperature in higher plants. Annu. Rev. Plant. Physiol.
31:491–543.
Bethenod, O., L. Huber and H. Slimi. 2001. Photosynthetic response
of wheat to stress induced by Puccinia recondita and post-infection
drought. Photosynthetica 39:581–590.
Björkman, O. and B. Demmig. 1987. Photon yield of O2 evolution
and chlorophyll fluorescence characteristics at 77 K among vascular plants of diverse origins. Planta 170:489–504.
Brooks, A. and G.D. Farquhar. 1985. Effect of temperature on the
CO2 /O2 specificity of ribulose-1,5-bisphosphate carboxylase/
oxygenase and the rate of respiration in the light. Planta 165:
397–406.
TREE PHYSIOLOGY VOLUME 28, 2008
INTERANNUAL PHOTOSYNTHETIC VARIATIONS IN A FAGUS CRENATA CROWN
Carrara, A., A.S. Kowalski, J. Neirynck, I.A. Janssens, J.C. Yuste and
R. Ceulemans. 2003. Net ecosystem CO2 exchange of mixed forest
in Belgium over 5 years. Agric. For. Meteorol. 119:209–227.
Cunningham, S.C. 2005. Photosynthetic responses to vapor pressure
deficit in temperate and tropical evergreen rain forest trees of Australia. Oecologia 142:521–528.
Farqhuar, G.D., S. von Caemmerer and J.A. Berry. 1980. A biochemical model of photosynthetic CO2 assimilation in leaves of C3 species. Planta 149:78–90.
Frak, E., X. Le Roux, P. Millard, E. Dreyer, G. Jaouen, B. Saint-Joanis
and R. Wendler. 2001. Changes in total leaf nitrogen and partitioning of leaf nitrogen drive photosynthetic acclimation to light in
fully developed walnut leaves. Plant Cell Environ. 24:1279–1288.
Galle, A. and U. Feller. 2007. Changes of photosynthetic traits in
beech saplings (Fagus sylvatica) under severe drought stress and
during recovery. Physiol. Plant. 131:412–421.
Grassi, G. and F. Magnani. 2005. Stomatal, mesophyll conductance
and biochemical limitation to photosynthesis as affected by
drought and leaf ontogeny in ash and oak trees. Plant Cell Environ.
28:834–849.
Grassi, G., E. Vicinelli, F. Ponti, L. Cantoni and F. Magnani. 2005.
Seasonal and interannual variability of photosynthetic capacity in
relation to leaf nitrogen in a deciduous forest plantation in northern
Italy. Tree Physiol. 25:349–360.
Han, Q., T. Kawasaki, T. Nakano and Y. Chiba. 2004. Spatial and seasonal variability of temperature responses of biochemical photosynthesis parameters and leaf nitrogen content within a Pinus
densiflora crown. Tree Physiol. 24:737–744.
Harley, P.C. and J.D. Tenhunen. 1991. Modeling the photosynthesis
response of C3 leaves to environmental factors. In Modeling Crop
Photosynthesis—From Biochemistry to Canopy. Eds. K.J. Boote
and R.S. Loomis. ASA, Madison, WI, pp 17–39.
Harley, P.C., F. Loreto, G.D. Marco and T.D. Sharkey. 1992. Theoretical considerations when estimating the mesophyll conductance to
CO2 flux by analysis of the response of photosynthesis to CO2.
Plant Physiol. 98:1429–1436.
Hikosaka, K., Y.H. Hanba, T. Hirose and I. Terashima. 1998. Photosynthetic nitrogen-use efficiency in leaves of woody and herbaceous species. Funct. Ecol. 12:896–905.
Hoch, G. 2005. Fruit-bearing branchlets are carbon autonomous in
mature broad-leaved temperate forest trees. Plant Cell Environ.
28:651–659.
Hui, D., Y. Luo and G. Katul. 2003. Partitioning interannual variability in net ecosystem exchange between climatic variability and
functional change. Tree Physiol. 23:433–442.
Iio, A., H. Fukasawa, Y. Nose, S. Kato and Y. Kakubari. 2005. Vertical, horizontal and azimuthal variations in leaf photosynthetic characteristics within a Fagus crenata crown in relation to light
acclimation. Tree Physiol. 25:525–536.
Ito, A., N. Saigusa, S. Murayama and S. Yamamoto. 2005. Modeling
of gross and net carbon dioxide exchange over a cool-temperate deciduous broad-leaved forest in Japan: Analysis of seasonal and
interannual change. Agric. For. Meteorol. 134:122–134.
Kaneko, S. and N. Hiratsuka. 1980. Fungi inhabiting on fagaceous
trees II. Host alternation of the beech rust, Pucciniastrum fagi.
Trans. Mycol. Soc. Japan 21:417–421.
Kaneko, S. and N. Sahashi. 1998. Bunarin wo hagukumu kinrui.
Bunitsu sogo, Tokyo, 229 p.
Kato, M.C., K. Hikosaka, N. Hirotsu, A. Makino and T. Hirose. 2003.
The excess light energy that is neither utilized in photosynthesis
nor dissipated by photoprotective mechanisms determines the rate
of photoinactivation in photosystem II. Plant Cell Physiol. 44:
318–325.
1429
Kitaoka, S. and T. Koike. 2005. Seasonal and yearly variations in light
use and nitrogen use by seedlings of four deciduous broad-leaved
tree species invading larch plantations. Tree Physiol. 25:467–475.
Krishnan, P., T.A. Black, N.J. Grant, A.G. Barr, E.H. Hogg,
R.S. Jassal and K. Morgenstern. 2006. Impact of changing soil
moisture distribution on net ecosystem productivity of a boreal aspen forest during and following drought. Agric. For. Meteorol.
139:208–223.
Kubota, M., J. Tenhunen, R. Zimmermann, M. Schmidt, S. Adiku and
Y. Kakubari. 2005. Influences of environmental factors on the radial profile of sap flux density in Fagus crenata growing at different elevations in the Naeba Mountains, Japan. Tree Physiol. 25:
545–556.
Leuschner, C., K. Backes, D. Hertel, F. Schipka, U. Schmitt,
O. Terborg and M. Runge. 2001. Drought responses at leaf, stem
and fine root levels of competitive Fagus sylvatica L. and Quercus
petraea (Matt) Liebl. trees in dry and wet years. For. Ecol. Manage.
149:33–46.
Mayr, S., C. Siller, M. Kriss, W. Oberhuber and H. Bauer. 2001. Photosynthesis in rust-infected adult Norway spruce in the field. New
Phytol. 151:683–689.
Medlyn, B.E., D. Loustau and S. Delzon. 2002. Temperature response
of parameters of a biochemically based model of photosynthesis. I.
Seasonal changes in mature maritime pine (Pinus pinaster Ait.).
Plant Cell Environ. 25:1155–1165.
Muller, O., K. Hikosaka and T. Hirose. 2005. Seasonal changes in
light and temperature affect the balance between light harvesting
and light utilisation components of photosynthesis in an evergreen
understory shrub. Oecologia 143:501–508.
Niimemets, Ü., O. Kull and J.D. Tenhunen. 2004. Within-canopy
variation in the rate of development of photosynthetic capacity is
proportional to integrated quantum flux density in temperate deciduous trees. Plant Cell Environ. 27:293–313.
Poot, P., J. Pilon and T.L. Pons. 1996. Photosynthetic characteristics
of leaves of male-sterile and hermaphrodite sex types of Plantago
lanceolata grown under conditions of contrasting nitrogen and
light availabilities. Physiol. Plant. 98:780–790.
Saigusa, N., S. Yamamoto, S. Murayama and H. Kondo. 2005. Interannual variability of carbon budget components in an AsiaFlux forest site estimated by long-term flux measurements. Agric. For.
Meteorol. 134:4–16.
Takanashi, S., Y. Kosugi, N. Matsuo, M. Tani and N. Ohete. 2006.
Patchy stomatal behavior in broad-leaved trees grown in different
habitats. Tree Physiol. 26:1565–1578.
Urban, L., X. Le Roux, H. Sinoquet, S. Jaffuel and M. Jannoyer. 2003.
A biochemical model of photosynthesis for mango leaves: evidence for the effect of fruit on photosynthetic capacity of nearby
leaves. Tree Physiol. 23:289–300.
von Caemmerer, S., J.R. Evans, G.S. Hudson and T.J. Andrews. 1994.
The kinetics of ribulose-1, 5-bisphosphate carboxylase/oxygenase
in vivo inferred from measurements of photosynthesis in leaves of
transgenic tobacco. Planta 195:88–97.
Wang, Q., A. Iio, J. Tenhunen and Y. Kakubari. 2008. Annual and seasonal variations in photosynthetic capacity of Fagus crenata along
elevation gradients in the Naeba Mountains, Japan. Tree Physiol.
28:277–285.
Wilson, K.B., D.D. Baldocchi and P.J. Hanson. 2000. Spatial and seasonal variability of photosynthetic parameters and their relationship to leaf nitrogen in a deciduous forest. Tree Physiol. 20:
565–578.
Yamori, W., K. Noguchi and I. Terashima. 2005. Temperature acclimation of photosynthesis in spinach leaves: analysis of photosynthesis components and temperature dependences of photosynthesis
partial reactions. Plant Cell Environ. 28:536–547.
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com