2.6 Statistical analysis
One-way ANOVA was conducted to test the differences in variables between
short-term and multi-year elevation of [CO2] at each
observation time, and the differences in parameters measured at predawn
and other time points, and the differences in values among growth
periods. Two-way ANOVA was performed to investigate the effects of
[CO2] increase treatments, observation time points
and their interactions on PSII efficiency. Pearson correlation was
applied to inquiry the relationship between parameters of PSII
efficiency. Statistical analysis was completed in IBM SPSS 21.0 (IBM
Crop., Armonk, NY, USA).
3. Results
Overall, compared with short-term
effect
(one growing season) of abrupt
increase of [CO2] by
200 μmol mol-1(AI200), constant increase of [CO2]
(four generations) by 200 μmol mol-1 (CI)
significantly increased the maximum photochemical efficiency of PSII
(Fv/Fm) at jointing
stage
(Table
2, Fig. 2a), while this
promotive effect was not observed at
heading and grain filling stages (Table 2, Fig. 2b, c). Stepwise
increase of [CO2] (four generations) by 160 μmol
mol-1 (SI) dramatically improved
Fv/Fm than short-term (one growing
season) abrupt increase of [CO2] by 160 μmol
mol-1 (AI160) during three growth
periods (Table 2, Fig. 2d-f). Moreover, the probability that trapped
exciton moves an electron further than QA-(ψo), electron transport quantum yield
(φEo) and performance index
(PIABS) under SI
were all prominently higher than that
under AI160 at grain
filling stage (Table 2, Figs. 4f, 5f, 6f), but under CI there was only
higher PIABS than under AI200 at
jointing stage with no significant difference in ψo and
in φEo (Table 2, Fig 6a).
The Fv/Fm, ψo,
φEo and PIABS all showed significant
diurnal variation, and the p -values of them among observation
time points were all < 0.001 (Table 2).
Fv/Fm gradually decreased to the minimum
value at midday, and then increased progressively back to predawn level
(Fig. 2). That was opposite to the changing trend of PPFD in a day
(Fig. 1).
Fv/Fm declined with the increase of PPFD
(p < 0.001) and air temperature (p <
0.1) over three growth stages (Fig. 3).
The daily trend
of
ψo and
φEo both showed the shape of letter “M”, with the two
peak values both appeared at 7:00 and 17:00 at jointing and heading
stages, and at 9:00 and 15:00 during grain filling stage in all
treatments (Fig. 4, 5). The PIABS changed during three
phases: started rising from predawn to 7:00, then decreased to minimum
value at midday, and then rising again (Fig. 6). There was no obvious
correlation between the diurnal change of ψo,
φEo and PIABS and PPFD or air
temperature (correlation coefficient was not shown).
Significant effects of the interaction between [CO2]
increase treatments and observation time points on
Fv/Fm, φEo and
PIABS were detected, indicating that effect of SI and
AI160 on
Fv/Fm,
φEo and PIABS were influenced by
observation time in a day (Table 2). SI promoted
Fv/Fm by reducing midday depression at
jointing stage (Fig. 2d). The relative change (RC) of
Fv/Fm was 6.5% from predawn to 11:00
under SI, lower than 8.7% under AI160, and
Fv/Fm recovered to 93.0%, 95.4% and
97.2% of that predawn observation at 13:00, 15:00 and 17:00, all were
higher than that under AI160 which were 91.2%, 94.3%
and 96.5% (Table 3). When it came to heading stage, SI only reduced the
down-regulation of Fv/Fm at midday, with
the RC of Fv/Fm from predawn to 11:00
being approximately 2/3 of that in
AI160 (Table 3). The
relative levels of PIABS between AI160and SI fluctuated over time at jointing and heading stage, which may
result in an overall insignificant difference between SI and
AI160 (Fig. 6d, e). At grain filling stage, SI improved
Fv/Fm, ψo,
φEo and PIABS at all observation time in
a day, overall increased by 1.0%, 4.6%, 5.5% and
12.2%, respectively (Fig. 2-5). No
significant interaction was found between effects of
AI200 and CI and observation time points (Table 2).
The largest midday depression of Fv/Fmoccurred at heading stage, reduced by 9.0%-14.1% relative to predawn
value (Table 3). Fv/Fm under all
treatments rebounded to 99% of predawn value until 19:00 when PPFD was
0.4 μmol m-2s-1 during jointing
stage, and was fully restored to predawn level at 19:00 during heading
stage when the PPFD dropped to 0 (Table 3). At grain filling stage,
Fv/Fm under AI200 and
AI160 measured at 17:00 showed no significant difference
compared with those observed at predawn, while
Fv/Fm under CI and SI were still 1%
lower than the predawn values (Table 3).
There was almost no significant difference between the midday
ψo and the predawn value at jointing and heading stages,
with the maximum RC being 13.2% - 14.6% and 9.5% - 12.2% across all
treatments, respectively (Table S1). While at grain filling stage, all
ψo were dramatically greater than those measured at
predawn, and the highest RC was 21.5% - 28.8%
(Table
S1), and the φEo in a day was obviously higher than that
at predawn, except at 13:00 (Table S2). From predawn to 7:00,
PIABS under AI200 and CI increased 11%,
but AI200 and SI caused no marked change at jointing
stage, and RC at heading and grain filling stages was enhanced, while CI
and SI led to lower RC than AI200 and
AI160, respectively (Table S3). The largest midday
attenuation of PIABS was up to 63.4% on average, which
occurred during jointing period (Table S3).
The φEo was
positively correlated with
ψo and
Fv/Fm at jointing and heading stages
under all treatments, while there was insignificant correlation with
Fv/Fm at grain filling stage under
AI200 and SI (Table 4). Correlation coefficient between
φEo and
ψo was greater than that between φEo and
Fv/Fm (Table 4). Moreover,
PIABS was found to be positively correlated with
Fv/Fm, φEo and
ψo and those three correlations were weaker than the
former in sequence, except that no significant correlation was tested
between PIABS and ψo under
AI160 during heading stage (Table 4).
The predawn Fv/Fm at jointing stage was
significantly higher than that at heading and grain filling stages, but
no significant difference was observed between the latter two stages
(Table 5). The predawn Fv/Fm under CI
and SI were correspondingly higher than those under
AI200 and AI160 during grain filling
stage (Fig. 2c, f). Moreover,
ψo,
φEo and PIABS measured at predawn
reduced dramatically with the advance of growth periods in all
treatments (Table 5). The reduction
of Fv/Fm, ψo,
φEo and PIABS from jointing to grain
filling stage were 2.6%, 19.9%, 21.9% and 51.9% under CI, all were
lower than those under
AI200,
which were 3.3%, 21.4%, 24.1% and 55.7%, respectively (Table 5). SI
caused less decrease (2.8%, 20.9%, 23.3%, 54.8%) than
AI160 (3.2%, 26.3%, 28.6%, 63.4%) in
Fv/Fm, ψo,
φEo and PIABS, but slightly more than
those under CI (Table 5).