Saddleback Journal of Biology - Saddleback College

Spring 2010 Biology 3B Paper
Hills, CA). Located on site were four Pasco GLX
data loggers with carbon dioxide probes and
photosynthesis tanks provided by Saddleback
College Department of Biology. A photosynthesis
tank is a two chambered tank in which the inner
portion can be sealed off, via a rubber stopper, to
create a sealed, isolated environment.
The four probes were set up and calibrated,
prior to each testing, to verify that all equipment was
functioning properly.
At the beginning of the experiment, each
plant was given a number for identification and a
control was established to ensure that the plants were
not performing carbon fixation. The control was set
up during the daylight at ambient temperature and
humidity (35%). A leaf from each plant was removed
and placed into the inner chambers of the
photosynthesis tanks. Carbon dioxide levels were
measured.
Appropriate solutions for the various
humidities were predetermined by referencing
previous studies (Greenspan, 1976; Sweetman, 1933;
Winston & Bates, 1960); saturated solutions were
prepared. The 0% humidity environment was
produced by placing 15 mL of drierite in the bottom
center portion of the inner photosynthesis chamber to
absorb all the moisture. A rubber stopper was placed
in the middle of the drierite to prevent direct contact
with the leaf. To produce a 33% humidity
environment, 15mL of saturated MgCl was placed at
the bottom of the inner chamber. The 75% humidity
environment had 15mL of saturated NaCl. The 100%
humidity level was obtained by placing 15mL of
deionized water at the bottom of the center chamber.
Each humidity condition had a rubber stopper in the
chamber so that the leaves could rest without
contamination or damage from the solutions.
Leaves were removed from the plant to
eliminate the effect of soil water potential on
stomatal response during acclimation. Leaves, seven
centimeters (cm) long, were removed by cutting with
scissors. The cut portion of the plant was covered
with parafilm to prevent any potential water loss. In
the absence of light, the leaves with the parafilm
were weighed (grams) and placed in the inner
chamber of the appropriate photosynthesis tank. Two
leaves, from separate plants, were placed in a
photosynthesis chamber to ensure that sufficient
carbon dioxide levels could be detected. The leaves
were paired consistently throughout all trials and
allowed to acclimate for three hours at the respective
environment. Pasco GLX data loggers with carbon
dioxide probes were turned on to record data for
three hours.
Once carbon dioxide data were collected,
leaves were removed from the photosynthesis tanks
and immediately reweighed. Stomatal imprints were
obtained initially by adding a drop of Superglue to a
blank glass slide. The top of the leaf was then firmly
pressed into the wet glue and pressure was applied
for ten seconds. The leaf was carefully removed from
the slide, leaving an imprint behind. The glue was
allowed to dry and the slides were analyzed under a
compound light microscope magnified at 100x. Trials
were repeated with freshly cut leaves for a total of
four trials, to ensure that each plant was rotated
through every humidity level. Stomatal data was
analyzed by photographing a 1 mm 2 area and
determining the percentage of open stomata.
Statistical analyses were conducted using Microsoft
Excel 2003; all data were analyzed by converting
parts per million (ppm) of carbon dioxide to grams of
carbon dioxide produced per gram of plant.
Results
The metabolic rate of the leaves for each
humidity level was averaged and graphed (Figure 1).
There was no significant statistical difference
between plant metabolic rate and humidity level
(p=0.655, ANOVA). At 0% humidity the average
metabolic rate was 7.25x10 -7 ± 4.18x10 -7 ; at 33%
humidity average metabolic rate was 1.20x10 -6 ±
8.62x10 -7 ; at 75% humidity average metabolic rate
was 9.50x10 -7 ± 1.43x10 -6 ; at 100% humidity average
metabolic rate was 1.58x10 -6 ± 2.59x10 -6 . There was
no significant statistical difference between stomatal
response and humidity level (p=0.292, ANOVA). At
0% humidity the average percentage of open stomata
was 16.96% ± 30.1%, at 33% humidity the average
percentage of open stomata was 34.69% ± 8.92%, at
75% humidity the average percentage of open
stomata was 34.20% ± 19.62%, and at 100%
humidity the average percentage of open stomata was
36.65% ± 32.19% (Figure 2).
Average Metabolic Rate
(g CO2 • g plant mass -1 • s -1 )
5.00E-06
4.00E-06
3.00E-06
2.00E-06
1.00E-06
0.00E+00
-1.00E-06
-2.00E-06
0% 33% 75% 100%
Humidity Level
Figure 1. The mean metabolic rates for each
humidity. ANOVA shows no significant difference
between humidities (p=0.655). Error bars indicate
mean ± SEM
2
Saddleback Journal of Biology
Spring 2010