Recently, an increasing demand has been noted for new natural substitute sweeteners
for sucrose and synthetic sweeteners such as saccharine and aspartame. Stevioside
is a sweetener of plant origin possessing a 200-350 times higher sweetening
property than sucrose. Stevioside comes from the South American perennial shrub
Stevia rebaudiana Bertoni and it is the most abundant among the nine
known sweet glycosides of the plant (Crammer and Ikan, 1986).
As reviewed by Geuns (2003), stevioside is safe when
used as a sweetener and there are no side effects like mutagenicity, carcinogenicity
Leaves of Stevia rebaudiana Bertoni have been used for the last 20 years
in countries of South America and Southeast Asia as a low-calorie sugar substitute
(Young, 2002). Their sweetness is due to the glycosides
stevioside, steviolbioside and rebaudiosides A, B, C, D, E, F and dulcoside
A (Brandle et al., 2002). Stevioside is 300 times
sweeter than sugar but has a bitter aftertaste (Brandle et
al., 1992). The sweetness of rebaudiosides increases with increasing
amount of sugar units bonded to the aglycon (steviol). However, their content
in the plant material decreases at the same time (Brandle
and Rosa, 1992; Brandle et al., 1992; Tolstikov
and Kovylyaeva, 2007).
However, the photosynthetic rate and the impact factors of Stevia
rebaudiana grown in China in autumn rarely been studied. Therefore,
the goal of the present study was to determine the diurnal changes of
net photosynthetic rate and the impact factors of Stevia rebaudiana
of Chinese region.
MATERIALS AND METHODS
The research was conducted in greenhouse with thick soil layer, Plant
Science and Technology College, Qingdao Agricultural University (36°15`N,
120°20`E), where the climate is temperate. The annual average temperature
is 16°C, August was the highest temperature month during the period,
which was average 25.1°C and the lowest temperature was in January,
which was averaged -1.2°C. The mean annual precipitation is 775.6
mm and most of which falling between July to September. The organic matter
containing of the soil is 1.24%, total nitrogen is 1.04%, available nitrogen,
available phosphorus and available potassium are 86.54, 24.58 and 85.72
mg kg-1, respectively.
No. 1 Qingtian (Stevia rebaudiana Bertoni) was used in all experimentation.
The photosynthetic characters of leaves in Stevia rebaudiana
were studied with LI-6400 portable photosynthesis system (made by LI-COR
Biosciences, USA) in middle September, 2007. Five plants were repetitive
measured and 2 healthy functional leaves at the same leaf position were
randomly selected every time and measured 3 times every leaf. The result
of each test was average to get the mean value.
The measurement index were: net photosynthetic rate/(Pn, μmol CO2/m2/sec),
transpiration rate/(Tr, mmol H2O/m2/sec), leaf to
air vapour pressure deficient/(Vpd, kPa), leaf temperature/ (Tleaf,°C),
photosynthetically active radiation/(PAR, μmol mol-1),
Intercellular CO2 contain/ (Ci, μmol mol-1),
air CO2 contain/(Ca, μmol mol-1), stomatal
conductance/(Cs, mmol/m2/sec), air relative humidity/(RH, %).
Diurnal Changes of Net Photosynthetic Rate and Transpiration Rate
of Stevia rebaudiana in Greenhouse in Autumn
Diurnal variation of photosynthetic rate (Pn) (Fig.
1) presented a bimodal curve and obvious midday depression phenomenon
occurred. The first peak value was the highest, which occurred at about
12:00 am, the net photosynthetic rate was 13.4 μmol CO2/m2/sec,
thereafter Pn reduced quickly and to the minimum value was 10.8 μmol
CO2/m2/sec and the second peak value occurred at
14:00 pm, the net photosynthetic rate was 13.4 μmol CO2/m2/sec
and decreased after then. The first peak was 90% higher than the second
peak by comparing the results, there was obvious midday depression of
photosynthesis. Pn became negative after 8:00 pm. The transpiration rate
followed closely the Pn course during the day. It presented a bimodal
||Diurnal changes of photosynthetic rate in leaves of
|| Diurnal changes of photosynthetic available radiation
and air temperature and transpiration rate and relative humidity
||Diurnal changes of intercellular CO2 and
stomatal constraint in leaves of Stevia rebaudiana
Diurnal Changes of the Main Environmental Factors
Plants photosynthesis appears complex concentration profiles affected
by many factors. Figure 2 shows that the change of PAR
is a single-peak curve, the highest PAR is at 1:00 pm in a day and then
decreased gradually after-words in autumn; Leaf temperature increased
first and the highest (36.91°C) appeared at about 1:00 pm, then dropped;
The relative humidity was higher at 7:00 am, 2:00 and 8:00 pm than other
time. The Vpd followed closely the PAR course during the day; however
its peak appeared at 2:00 pm.
Stomatal movement is important for plants to exchange gas and water with environment.
It has very close relation with photosynthesis, respiration and transpiration.
According to the opinion of Sharkey and Ogawa (1987).
If Gs and Ci decreased as Pn decreased and cause a decrease in stomatal limitation
value (Ls), the stomatal limitation is dominant; If Ci increase as Gs and Ls
decrease, nonstomatal limitation is the dominant one. Figure 3
and 4 show that Ci and Ls decreased and Gs increased as Pn
decreased. The results showed that diurnal change in net photosynthetic rate
(Pn) of Stevia rebaudiana leaves was a typical bimodal curve determinately
regulated by stomatal conductance.
||Diurnal changes of stomatal conductance and transpiration
rate in leaves of Stevia rebaudiana
|| Path analysis among photosynthetic rate and main factors
Diurnal Changes of Stomatal Conductance and Transpiration Rate in
Leaves of Stevia rebaudiana
Figure 4 show that diurnal variation of stomatal
conductance (Gs) was a double peak curve, the peak (55.8 mmol/m2/sec)
of Gs appeared at 1:00 pm and the second crest value existed at about
3:00 pm, it was only 31.1 mmol/m2/sec. Its pattern was the
same with Pn.
Analysis of the Quantitative Relationship among Pn and Main Environmental
The effect of environment factors on Pn was not isolated or individual
but a result of integrate all factors affected. To make the further analysis
of the quantitative relationship between Pn and main environmental factors,
result was further investigated by making use of the regression analysis.
The regression equation between Pn and its influence factors was:
Pn = 3.6885+95.14803Gs-0.04062Ci-0.21166T+0.03332Cs-0.2308RH-0.00042PAR+3.532Vpd-1.1953Tr
R = 0.9927* (R0.05 = 0.9853)
The results approve that the regression equation is significant and the
correlation is close, so it can be used in the practical operation. Regression
analysis showed that there was a significant regression relationship between
the Pn and its influence factors.
Table 1 shows that factors have ordinal influence on
the diurnal change of Pn in Stevia rebaudiana: Vpd, Gs, Cs, PAR, RH, Tr,
T and Ci. The Tleaf, Tr, PAR and Gs had indirect positive effect on the
Pn via affecting Vpd. While Ci, Ca, RH had indirect negative effect on
the Pn via affecting Vpd. So the Vpd is the most effective factor to Pn
and the other factors affect the Pn via the Vpd.
The results showed that under natural conditions (control), diurnal course
of net photosynthetic rate (Pn) in Stevia rebaudiana leaves presented
two peaks and daily variation of stomatal conductanc (Gs) and transpiration
rate (Tr) appeared two peaks; Pn increased with enhanced photosynthetically
active radiation and stabilization when photosynthetically active radiation
reaches certain level from sunrise. At 12:00 am, Pn appeared the peak when the
environmental factors formed the most proper combination. However, PAR and temperature
increased gradually when much higher light density and the high temperature
inhibited Pn at 1:00 pm. It was not highest leaf temperature when the PAR was
maximum because Tr and RH increased with PAR and they cause leaf temperature
not to be too high. So midday depression of photosynthesis was not closely related
with temperature in greenhouse. The main cause of midday depression of photosynthesis
was due to higher light intensity which caused photorespiration increased and
Pn decreased (Heber et al., 1987).
A physiological process characteristic of crop photosynthesis is a complex
process. Pn is closely related with chlorophyll content, leaf thickness and
leaf maturity and light intensity, temperature, relative humidity and moisture
content in the soil play important roles, too. Pn in leaves of Stevia rebaudiana
leaves varied in day time, it was double peak curve and there was obvious midday
depression of photosynthesis. Midday depression is the character of C3
plants. The lowest valley between 2 peaks is the result of high photorespiration
under high light intensity and high temperature. It is also the reason that
protective stomatal conductance and intercellular CO2 concentration
decrease. The function of stomata in plants is controlling gas exchange and
modulating water balance, istomatal aperture and stomatal resistance play important
roles in water status and CO2 assimilation. Generally speaking, the
Ci and Pn decrease as stomatal aperture decreases or the stomatal resistance
increase and reducing stomatal opening status led transpiration rate decrease
and water loss reduction; Only a few was reported non-stomatal regulation sometimes
(Morison, 1987; Schulze et al.,
1987; Bahrun et al., 2002).
The results show that Vpd, whose coefficient of determination is the highest,
is a main factor affect to Pn. midday depression of photosynthesis was not closely
related with temperature in greenhouse. The main cause of midday depression
of photosynthesis was due to higher light intensity which inhibited photosynthesis
and promoted photorespiration while the correlativity of temperature and midday
depression of photosynthesis was small. It is in accordance with Raschke` theory
(Pandey et al., 2003) that if Vpd was maintain
constant the change of temperature at a certain range does not led to the obvious
change of photosynthesis. In fact, the effect of temperature on photosynthesis
is mediated via Vpd. Bisides, temperature can cause stomatal conductance and
transpiration rate which has direct or indirect effect on photosynthesis (Raschke
and Rseman, 1988).