Materials
- a potometer with a rubber tubing, a syringe, capillary tube, and three-way tap
- secateurs
-
leafy shoot of plants (*Rubiaceae, Verbenaceae, Oleaceae, and Rutaceae)
- a 50(w) x 30(d) x 25(h) cm tub (big enough for a potometer to be completely put)
- water
- a clamp stand
- 2 x clamps
- stopwatch
- a 30cm ruler (± 0.1 cm)
- sheets of squared paper (1cm grid)
- a calculator
Method
- Cut four different kinds of branch of plants using secateurs *Make sure the thickness of the branches fits well to the rubbing tube of a potometer
- Put 35 L of water into a tub (much enough for a potometer to be sunk)
- Place a potometer into the water completely to allow all water to enter throughout the whole tube
- Pull plunger to allow water to enter into a syringe
- Turn a three-way tap (the middle one of a three-way tap for closure) to the side of rubbing tube to close the way for water to enter
- Push (not completely) the plunger to have all air bubbles go out
- Then turn the tap again to the side of capillary tube to block the water-passing way
- Push the plunger to make sure there is no air bubble left in the rubbing tube
-
Use secateurs to cut the small end of a leafy shoot of *Rubiaceae (this is done in the water tub) with its cut part still kept in water.
- Insert the branch into the rubber tubing (it is still conducted in the water tub) making sure there is no empty space between branch and the tubing.
- Set up two clamps onto a clamp stand with shorter one to the bottom and longer one to the upper.
- Have a shorter clamp hold the potometer and longer one catch the branch (refer to the Diagram 1 below)
- Open the whole passage except the syringe by turning the three-way tap to the side of it.
- Operate stopwatch for 1minute, measuring from 0mL
-
Measure and record how much distance in cm water has been travelled, or reduced (it can be measured via attached ruler-like markings in cm on capillary tube)
- Measure the radius of the capillary tube by using a ruler
-
Figure out the area of all detached leaves by putting them on 1cm squared grid and counting 1cm2 squares.
- Calculate the volume of water uptake per unit time per unit area
- Repeat step 1-18 with the rest of other species.
- Repeat step 1-19 four times more.
* In this experiment, there cannot be control of no treatment.
Diagram 1: a potometer set up on a clamp stand to measure the transpiration rate
The image of a potometer (Philip Harris Potometer, n.d.)
Result
Table 1: raw data for the distance (± 0.1 cm) that water uptake by four different plants (*Rubiaceae, Verbenaceae, Oleaceae, and Rutaceae) has travelled during one minute
Note: the measurement of potometer is the same as a ruler. Thus uncertainties are based on ± half of the place value of the last measured value, provided there is limit for the precision at both e
nds of the ruler.
Table 2: processed data with the average distance that different plant species’ water uptake travelled and standard deviation (± 0.1 cm)
Note: 1.6 is circled because it is an outlier. However, the calculation for standard deviation requires at least five samples, so the outlier is included in the calculations for both mean and standard deviation.
Sample calculation
-Mean
all the values
Mean = –––––––––––––––––––––
the number of the values
(1.6 + 0.6 + 0.5 + 0.4 + 0.4)
= ––––––––––––––––––––––––
5
= 0.7 cm
-Standard Deviation
√ ∑(x-Mean)2
Standard Deviation = –––––––––––
√ n-1
√ {(1.6−0.7)2 +(0.6−0.7) 2 + (0.5−0.7) 2 + (0.4−0.7) 2 + (0.4−0.7) 2}
= ––––––––––––––––––––––––––––––
––––––––––––––––––––––––
√ 4
= 0.5 cm
Table 3: data required for the calculation of volume of water uptake (cm3) per minute per m2 of four different species (*Rubiaceae, Verbenaceae, Oleaceae, and Rutaceae)
Note: the uncertainty of ± 0.0001m² for total surface area is from 1 cm² grid paper which was used for counting the surface area of plant species. When 1cm² converted to m², it is 0.0001m².
Note: the radius of the capillary tube is 0.5 mm. From this, the uncertainty is ± 0.1mm. Then when 0.1mm³ is converted into cm³, it is 0.0001cm³.
Volume = π r 2 h
= π × (0.05)2 × (0.7)
= 0.0055 cm3
r (radius of capillary tube) = 0.5 mm = 0.05 cm
h = (mean of) distance of water uptake
-
Volume of water uptake per minute per m2
Volume of water uptake per minute per m2 = volume of water uptake ÷ total surface area
= 0.0055 ÷ 0.0072
= 0.7639 cm3 per minute per m2
Observations (qualitative data)
This experiment was done throughout two days. In one day with more humid condition, the rate of water uptake was slower. When there was sometimes wind blown, it affected the speed of water uptake: faster.
What is more, wet branch slowed down transpiration rate. In other words, the first trial with dry branch had the fastest transpiration rate in a minute, compared to the rest of four trials.
The insertion of branch into a rubber tubing was not always perfect. Thus, some water leaked from the crack between two. Though for the solution to that, vaseline was used, it was not that helpful.
*Rubiaceae had spider nest on its surface of leaf. This somehow would have effect on transpiration rate because the process happens at the surface of leaves. The surface hidden under the nest would not have transpired as much.
In fact, as time and trial goes, the skill of removing air bubble out improved. For the *Oleaceae, it was the first time to do experiment. It took much more time and had so various values than the last experiment was conducted. The last experiment was for *Rubiaceae. Except for the outlier 1.6 due to only dry branch, all the trials were precise and close to each other.
Graph 1: the volume of water uptake (cm3) per minute per m2 by four different species of plant (*Rubiaceae, Verbenaceae, Oleaceae, and Rutaceae)
Note: Error bars- data range
Each mean is drawn in bar graphs, and the highest and lowest transpiration rate result for each plant is shown around it.
Conclusion
As mentioned in hypothesis, different species have different transpiration rate from each other. This can be proven by the data above clearly in Graph 1. The plant from Rubiaceae family took 0.7639 mL of water per minute per m² while 0.9512 mL was absorbed for Verbenaceae, 0.2678 mL for Oleaceae, and 0.1297 mL for Rutaceae according to Table 3. This is because all four species have different leaves. Of all features of leaves having effect on transpiration, the thickness of cuticles and stomatal complex are essential.
Cuticles are hydrophobic waxy layers on the surface of plant and thus make water to move not easily through leaves (Transpiration, n.d.). The thicker waxy cuticles can indicate the increasing hydrophobic. Then the transpiration rate increasingly slows down.
About stomatal complex, it involves stomatal density, stomatal size, and stomatal index. All three aspects of it are different from species to species and thus have impact on transpiration rate of each species. For example, Saadu et al. (2009) found that Euphorbiaceae with 16.45a mm−² of stomatal density, 52.30a μm of stomatal size, and 3.03a % of stomatal index has 1.47 x 10−4a mol per m² per sec of transpiration rate (the method for measuring transpiration rate was a cobalt chloride paper method). The family had different transpiration from Cyperaceae with 28.95b mm−² of stomatal density, 104.77b μm of stomatal size, and 17.16b % of stomatal index has 2.68 x 10−4b mol per m² per sec of transpiration rate. To extend further, Saadu et al. (2009) also discovered that especially high stomatal density is effective in water uptake. Convolvulaceae was the family species with the highest stomatal density of 35.75b mm−² and thus had the greatest transpiration rate of 3.07 x 10-4b per m² per sec. Whether the size of stomata and leaf is large or not, it is more important that there are many ways out for water molecule on the surface of leaf for transpiration to happen fast.
Even though different family species were researched, from this, it still can be known that each species has different variations on the features of its leaf. These variations result in different transpiration rate.
Though in Table 3, Oleaceae absorbed the most volume of water, the surface area for transpiring that much amount of water was as large. More accurate data is volume per unit time per unit leaf area, and this is shown apparently in Graph 1. In Graph 1, Verbenaceae absorbed the most amount of water. However, considering the error bar especially of Rubiaceae, it is not reliable whether Verbenaceae with thick waxy cuticles and elliptic pinnate leaves has the fastest transpiration rate. The matter of reliability of this data will be discussed in evaluation.
Evaluation
In this experiment, there are several errors presented. The first source of errors is random including human errors. What was uncontrolled variable was the weather condition. This was also discussed in observations. The weather cannot be controlled to desired or constant condition for two days even for a few hours. Especially when doing experiment for Oleaceae, the condition was so various. The excessively high value of 1.80 cm for the trial 2, as seen in Table 2, was because of wind blown. Blowing wind had the plant transpire faster. As seen in Graph 1, the error bar ranges so widely due to this factor; it was huge impact on the result. Other than wind, the conditions of light, humidity, and temperature would be very different from day 1 to day 2. The experiment on day 1 was in the noon when the temperature was high and humidity was low. On the other hand, the day 2 experiment was in the morning when the temperature was relatively low and the humidity at that time was high. This seems a big issue, affecting enormously the results. Verbenaceae, Oleaceae, and Rutaceae was done in day 1, so they were likely to transpire faster than when they would be in the condition of day 2. With this, the comparison among plants is not reliable much.
There are always human errors. This often involves incorrectly reading the capillary tube on potometer in this case. Thus, the uncertainties of the equipment can reflect this kind of error, not to mention the uncertainties of the measurement itself. There was another random human error. Whenever the experimenter moves, the amount of either sunlight or fluorescent light to which the plants was exposed varied. This human error is random and different each time. Though it is hard to reduce it to zero, it does not seem to have much impact on the result as the variation in the amount of light was actually very short-term.
Other errors contain air bubble, wet leaves and branches, spider nest or dust, and finally the insertion of rubber tubing. Firstly, the removal of air bubble was the most problematic issue. Quite large air bubbles were stuck in the passage, and it was hard and thus time-consuming to take those out despite a number of adjustments of three-way tab and a syringe. Every trial for each of four plants would have different amount of air bubbles. As mentioned in observations, transpiration test for Oleaceae was the very first time trials and therefore, massive air bubbles were stuck in the passage. Seen in the row of Oleaceae in Table 2, the lowest value is 0.5 cm where a large air bubble blocked the passage.
Secondly, all wet leaves and branch were the issues. At the dry state of leaves and branch, i.e. the first trial of each plant, the water uptake was much and thus transpiration rate was very fast. The outlier 1.60 cm in Table 2 was the evidence. Comparing to the rest of four trials, the first trial had much higher value. The wet state of leaves and branch can be considered as the high humidity in the air, so it slows down the diffusion, loss, or transpiration of water molecules into air with already much water. This error could result in much impact on the transpiration rate and thus the result.
The third source of error is the spider nest or dust on the surface of plants. Though they are not that influential enough, they could have slight effect on transpiration of Rubiaceae, blocking some of the stomata. Perhaps the values of distance of water uptake and thus the volume of water uptake per unit time per unit area would be lower than what is expected or supposed. Still this error is small and trivial.
The last error is whether the branch fits into the rubber tubing, as briefly mentioned in observations. When there was room between the tubing and the branch, water leaked there. As soon as the three-way tap was manipulated to open all the passages, the water uptake for few seconds was so great. The problem here is that the water was not absorbed by the plant but moved to the crack. Therefore, sooner or later, the water uptake suddenly stopped or greatly slowed down. The difference is so great that this would have very large effect on the results and their error bars.
There two limitations to consider about. One is whether the removal of root affects the ability of water uptake. The water uptake by roots would not be the same as by branch or stem. The other is whether water uptake can necessarily mean the water loss or evaporation, which is what really transpiration means. Even if two measurements are different, it can be inferred that the large amount of water uptake leads to that much amount of water evaporation.
Time management was also the source of error. Since it was poor, the divided experiment was conducted for two days which had so different condition of weather that the effect on the result was great. All this was discussed in earlier part of evaluation.
To comment on precision and accuracy of this experiment and result, precision only for Verbenaceae and Rutaceae is high and accuracy is high as long as only general difference between species is concerned. In terms of high precision, the standard deviation for those two species was very close to 0 (standard deviation 0.0 in Table 2 was rounded off). This indicates small scatter around the mean and further implies few random errors for them. In fact, most of many random errors took place when conducing the trials of Rutaceae and Oleaceae. Therefore the standard deviation of two species is 0.5, meaning wider scatter around the means and finally the lower precision.
Accuracy is high as long as only the general conclusion is compared. In other words, the general conclusion that different species transpire at different rates also applies for known data or other research paper. However, when it comes to ‘how’ different species result in different transpiration rate, there are so many other aspects of leaves to compare such as waxy cuticles, stomatal complex, succulence, the side of occurrence of transpiration, etc. The result is not that reliable when taking all those account. Whether the family Rutaceae transpire the slowest is not sure. However, the fact of different transpiration rate for different species is quite reliable with the help of high precision of result for Verbenaceae and Rutaceae.
For better experiments, it is the fist thing to correct errors mentioned above. Keeping the weather controlled is nearly impossible. However, the best way can be to complete an experiment as quickly as possible in one day. The weather condition such as the amount of light and temperature is so various even during few hours.
As mentioned briefly, human errors are random so they can not be fully removed. The error of fluctuating amount of light due to moving experimenter can be improved by placing the set up of a potometer in a dark closed system with one source of light, not open space with sunlight and fluorescent light. This further can prevent the change in temperature, wind, humidity, etc.
To solve the air bubble problem, the sufficient amount of actual practice is needed. As experienced during the experiment, as trials went the skill improved. Another practical way is first of all to remove air bubbles in a syringe. The air bubbles in the syringe caused bigger and more bubbles in the passage. Since the syringe can be detach from the potometer, it is possible to pull water up to the end and push the plunger back all the way to the other end to remove all air bubbles. Then fill the syringe up with water again and connect it to the three-way tap. Now water from the syringe would get rid of all air bubbles stuck in the passage when the tap is adjusted properly.
In regards to the error of wet and dry state of branch and leaves, it seems the best way to just wait until the wet branch dries. The paper towel to dry those would be helpful for reducing the waiting time.
Similar to the method right above, it is important to eliminate any water, dust, or even spider nest out of the surface of plant. These blocked stomata and prevented transpiration from occurring at the spot.
The matter of suitable insertion of branch into a rubber tubing could be resolved by having different size of rubber tubings, cutting the branch at the same thickness as the rubber tubing. As temporary expedient, vaseline can be used. This will prevent the leaking of water from the crack between the branch and rubber tubing.
The next step for improved laboratory is to repeat as much as possible. As long as biology is concerned, variations are inevitable. Just to decrease the variations, size of error bars, and standard deviation, many repetitions are required. The more there are repetitions, the greater possibility there is that the results get closer to the expected outcome.
For future research, more than four families of plant species can be data range, which would give more information. Since this laboratory experiment is quite superficial in that it only identifies the differences in transpiration rate, deeper consideration is required on how the differences come from. It would be better not only to check the appearance of leaves but also the quantitative data for example like the thickness of waxy cuticles and stomatal size as in other experiments done by others.
Bibliography
Clegg, C. (2007). Biology for the IB DIPLOMA. London: Hodder Murray.
How to Calculate Leaf Surface Area. (2010). Retrieved on August 31, 2010 from
Philip Harris Potometer [Image] (n.d.). Retrieved on September 5, 2010 from
Roberts, M., King T., & Reiss, M. (1994). Practical Biology For Advanced Level. China: Nelson
Saadu, R. O., Abdulrahaman, A. A. & Oladele, F. A. (2009). Stomatal complex types and transpiration rates in some tropical tuber species. Retrieved on September 22, 2010 from
Transpiration. (2009). Retrieved on September 17, 2010 from
Transpiration. (n.d.) Retrieved on September 19, 2010 from
Transpiration- Factors Affecting Rates of Transpiration. (n.d.). Retrieved on September 22, 2010, from
Appendix
Rubiaceae Verbenaceae
Oleaceae Rutaceae