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The effectiveness of Lemna Minor at removing excess phosphorus from water
Andrew D. Tyson
Biology 220W
Section 012
Michelle Harrison
1
INTRODUCTION
Contaminated soils and waters pose a major threat to both the environment. An effective
way to treat contamination in waters and soils is phytoremediation. Phytoremediation is the use
of green plants to either remove pollutants from the environment or render the pollutants
harmless.1 Rhizofiltration is a specific type of phytoremediation that uses plant roots to absorb
pollutants from water and aqueous waste streams.1 It can be used to uptake harmful metals and
other substances.1
Lemna minor is a plant may be used for effective rhizofiltration since it is known to
uptake excess nutrients and concentrate them in its tissues.2 Lemna minor is a plant that is a
member of the duckweed family.3 It is naturally occurring in almost all freshwater ponds and
slow-moving streams in Asia, Africa, North America, and Europe.3 It is also found in South
America and Australia even though it is not naturally occurring there.3
Each Lemna minor plant floats in the water and has one to four leaves, of thalli, along
with a single root that hangs down into the water.2 Lemna minor typically reproduce asexually by
division.4 Flowers are rarely produced on the Lemna minor.4 It grows in water with high levels of
nutrients and a pH around seven.5 Since Lemna minor grow at an extremely rapid pace, they can
form a complete layer over the top of the body of water they occupy in a very short amount of
time.4
Lemna minor is overpowering compared to its potential competitors. It has adapted its
structure over time in order to grow very quickly.4 This allows it to grow and populate bodies of
water very quickly. Most often, Lemna minor overcomes inter-species competition by growing
thick layers over the top of water bodies.4 These layers take away sunlight from the rest of its
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competition and eliminate them.4 By absorbing as much of the surrounding resources as possible,
Lemna minor overcomes intra-species competition enabling it to grow and reproduce.4
The purpose of this experiment is to determine how changes in nutrient concentration
affect the growth of Lemna minor. It is also to determine if Lemna minor would be effective in
rhizofiltration. If it is effective in rhizofiltration, it would help advance research in different
methods of phytoremediation. For this experiment, I hypothesize that the starting population size
will have no effect on the overall growth rate of Lemna minor as it is expected to grow
exponentially. I also hypothesize that an increase in phosphorus will help Lemna minor grow
more quickly than the control population with no excess phosphorus.
METHODS AND MATERIALS
The overall experiment was divided into two separate experiments. The first experiment
consisted of setting up two 10 oz. containers with pond water and putting 12 and 24 individual
Lemna minor plants in each container respectively. The containers were marked for
identification purposes. After the plants were put in their containers, the number of leaves, also
known as thalli, were counted. For the next five weeks, the number of thalli were counted once a
week to monitor growth rates of each population. Each container was also checked multiple
times throughout the week in order to ensure that the proper amount of water had been added and
that algae growth was kept to a minimum.2
The second part of the experiment was similar to the first with one key difference. Three
10 ounce containers were filled with pond water. Next, 24 Lemna minor plants were put into
each container, and the number of thalli were counted. The key difference in this experiment was
that two milliliters of phosphorus were added to each container. After the experiment was set up
3
and similar to above, the number of Lemna minor thalli was counted for each container each
week for five weeks. Phosphorus must also be added each week. The values counted from the
first experiment and this experiment was compiled into a class data sheet.2
For both experiments, the dependent variable was the number of Lemna thalli counted.
For the first experiment, the independent variable was the number of Lemna plants started with.
For the second experiment, the independent variable was the nutrient added. The control for the
second experiment was the container with 24 Lemna plants initially added.2
Three different equations were used to analyze the population growth. To find the
population growth rate, the number of thalli counted was plotted against time. The maximum
intrinsic rate of growth (rmax) is found by dN/dt = rmaxt where dN/dt is the change in population
size at a specific point in time.2 The geometric growth rate (λ) is the factor by which a population
increases in size in one specific unit of time.2 It is found by λ = Nt+1/Nt where N is the number of
thalli and t is time. When the population reaches its maximum size, the growth rate of the
population will be zero.2 This is known as the carrying capacity (K) of the population. The
carrying capacity is found by N = dN/dt. N = K when dN/dt is zero.2 The carrying capacity can
also be found by (Surface Area of container/Surface area of individual Lemna minor thalli).2
RESULTS
Experiment 1
For experiment one, both the population growth and the maximum intrinsic rate of
increase (Rmax) for both the population of Lemna minor that started with 12 plants and the
population that started with 24 plants needed to be determined.
4
The population growth was found by plotting the number of plants (N) against time. As
Figure 1 illustrates, Lemna minor experienced exponential growth in both the population that
started with 12 plants and the population that started with 24 plants. The population with 12
plants saw its growth rate slow in the last seven days of the experiment while the population with
24 plants saw exponential growth that continued throughout the entirety of the experiment.
Number of Thalli
Population Growth of Lemna Minor
2000
1800
1600
1400
1200
1000
800
600
400
200
0
Started w/ 12 Plants
Started w/ 24 Plants
0
10
20
30
Time (days)
Figure 1. Population growth rates of Lemna minor populations that started with 12 plants and
24 plants respectively.
As the population size approaches its carrying capacity (K), the population growth rate
begins to slow. To determine the maximum intrinsic rate of increase (Rmax), the natural log of N
was plotted against the time in days. The slopes of the linear lines of best fit plotted on Figure 2
show the Rmax values for both populations.
5
Maximum intrinsic rates of increase for
Lemna Minor
y = 0.188x + 2.7141
y = 0.1272x + 3.6845
10
ln(N)
8
Started w/ 12 Plants
6
4
Started w/ 24 Plants
2
Linear (Started w/ 12
Plants)
Linear (Started w/ 24
Plants)
0
0
10
20
30
Time (days)
Figure 2. Maximum intrinsic rates of increase (Rmax) for Lemna minor populations that started
with 12 plants and 24 plants respectively.
The equations for the lines of best fit are shown in Figure 2 for each population. For the
population that began with 12 plants, the Rmax = 0.188. For the population that began with 24
plants, the Rmax = 0.1272.
To find the geometric rate of increase (λ), the equation (N+1)/N was used. The geometric
rates of increase for both populations in Experiment 1 are shown in Table 1.
Time
(Day)
Started w/
12 (λ)
Started w/
24 (λ)
0
6.71153846
5.33018868
14
5.14040115
2.58584071
21
1.79004091
2.56080356
28
N/A
N/A
Table 1. Geometric Growth Rates for Lemna minor populations that started with 12 plants and
24 plants respectively.
Sample Calculation for “Started w/ 24 (λ)”:
λ = (N14/N0)= (282.5/53) = 5.33018868
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As Table 3 shows, both populations had the greatest increase in size during the first time
interval from Day 0 to Day 14. However, since the population that started with 12 plants had a λ
value of 5.140 during the second time interval compared to a λ value of 6.712 during the first
time interval, it can be argued that the population that started with 12 plants had a higher growth
rate during the second time interval as it was half as long.
The carrying capacity (K) can be estimated by using the formula (Surface area of
container/ Surface area of Lemna minor thalli). Using this formula yields a carrying capacity of
approximately 1453 Lemna minor thalli. The calculations can be seen below.
Calculations for Carrying Capacity:
(Surface area of container/ Surface area of Lemna minor thalli)
(5809 mm2/4 mm2)
= 1452.24 thalli
The carrying capacity (K) of the population can be determined by plotting λ against N.
Figures 3 and 4 show λ against N for both populations in Experiment 1.
λ
Lambda vs N for Lemna minor
8
7
6
5
4
3
2
1
0
-1 0
-2
-3
y = -0.0053x + 6.4964
Started w/ 12
Linear (Started w/ 12)
500
1000
1500
2000
Number of Thalli (N)
Figure 3. Carrying Capacity for Lemna minor population that started with 12 plants.
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Lambda vs N for Lemna minor
6
5
4
3
y = -0.0035x + 4.7479
2
λ
Started w/ 24
1
Linear (Started w/ 24)
0
-1 0
500
1000
1500
2000
-2
-3
Number of Thalli (N)
Figure 4. Carrying Capacity for Lemna minor population that started with 24 plants.
To determine the carrying capacity (K), λ is plotted against N. Using the equation of the
linear line of best fit, K can be determined by inputting a value of 1 for y and solving for x. For
the population that started with 12 plants, K = 1037.06. For the population that started with 24
plants, K = 1070.83. The calculations for both are shown below.
Calculations for Carrying Capacities:
1 = -0.0053x + 6.4964
-0.0053x = -5.4964
x = 1037.06
1 = -0.0035x + 4.7479
-0.0035x = -3.7479
x = 1070.83
Experiment 2
In all cases, the control is a population of Lemna minor that was not treated with any
excess nutrients unless stated otherwise. Both populations started with 24 plants each.
Again, both populations experienced exponential growth. However, as Figure 5 shows,
the population treated with excess phosphorus grew at a faster rate than the control.
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Population Growth of Lemna minor
Number of Thalli (N)
2500
2000
1500
Phosphorus
1000
Control
500
0
0
5
10
15
20
25
30
Time (days)
Figure 5. Population growth of Lemna minor treated with excess phosphorus and no excess
phosphorus respectively. Both populations started with 24 plants.
To determine the carrying capacity (K) for the population of Lemna minor treated with
phosphorus, λ was plotted against N. Table 2 below shows the values of λ that were used in
Figure 6. These values were found the same way as described above.
Time
(days)
Started w/
24 (λ)
Phosphorus
(λ)
0
5.33018868
8.2058824
14
2.58584071
3.2461183
21
2.56080356
2.5027598
28
N/A
N/A
Table 2. Geometric Growth Rates for Lemna minor control and population treated with
phosphorus. Population started with 24 plants.
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The Lemna minor population treated with phosphorus had a higher geometric growth rate
in both of the first two time intervals. Both populations had almost equal geometric growth rates
in the last time interval.
Lambda vs N for Lemna minor treated
w/ Phosphorus
10
8
6
4
Treated w/ Phosphorus
λ
2
0
-2 0
500
1000
1500
2000
2500
-4
y = -0.0055x + 6.8802
-6
-8
Linear (Treated w/
Phosphorus)
Number of Thalli (N)
Figure 6. Carrying capacity for Lemna minor treated with phosphorus. Population started with
24 plants.
Comparing Figure 5 to Figure 6, there is almost no difference between the carrying
capacities of the control and the population treated with phosphorus. For the control, K =
1070.83, and for the population treated with phosphorus, K = 1069.13. The calculations for
determining the K value of the population treated with phosphorus are shown below.
Calculations for Carrying Capacity:
1 = -0.0055x + 6.8802
-0.0055x = -5.8802
x = 1069.13
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DISCUSSION
Experiment 1
The population growth rates for both the Lemna minor population that started with 12
plants and the population that started with 24 plants were very similar. This is not surprising as
the 12 plant population should theoretically start growing at the same exponential rate as the 24
plant population as soon as it reaches that amount of plants. Over a long period of time, such as
the one this experiment was performed under, this difference in starting population size becomes
negligible.
As expected, the population that started with 12 plants also had a higher maximum
intrinsic rate of growth than the population that started with 24 plants. This is due to the fact that
the population with 12 plants starts with a smaller population and, therefore, can grow at a faster
rate than the population that started with 24 plants since they have more room for growth. The
smaller population also will grow at a faster rate since they have fewer plants, while having the
same amount of nutrients to use as the larger population.
The population with 12 plants also had higher geometric rates of increase during the first
two time intervals. This is not surprising for the same reasons stated above. The smaller
population had more nutrients to use per plant and could therefore grow at a quicker rate.
Two methods were used to find the carrying capacity. The first method found the amount
of Lemna minor thalli that could fit on the water surface of the container using their surface
areas. The second method plotted geometric growth rates against the number of Lemna minor
thalli. The second method is more accurate as the first method does not account for the fact that
the Lemna minor thalli will not fit perfectly together to take up every available square millimeter
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of space. It also does not account for the fact that Lemna minor can grow in layers, and can grow
in more than one layer.
The carrying capacities for both population sizes were expected to be the same since the
populations were in equal size containers under equal conditions. This was not the case as the
population that started with 12 plants had a carrying capacity of approximately 1040 while the
population that started with 24 plants had a carrying capacity of approximately 1070. While this
is a small difference, it is enough to warrant questioning. This could be due to some source of
error such as the starting amount of thalli in each population or the effect that water and algae
amount had on the population.
Experiment 2
As was hypothesized, the population treated with phosphorus grew at a slightly faster rate
than the control. This is attributed to the extra nutrients, in this case phosphorus, that allowed the
plants to grow. Since phosphorus is usually a limiting nutrient, the extra nutrients have a large
effect on the population.
As was also expected, the phosphorus had a higher geometric growth rate over the course
of the experiment. This is easily explained by the excess nutrients the phosphorus population
possesses, allowing it to grow at a much faster rate as phosphorus is no longer a limiting nutrient.
The phosphorus had a carrying capacity of approximately 1069 while the control
population had a carrying capacity of approximately 1070. This similarity is expected as the
populations were kept in the same size containers under the same conditions.
How does this relate to Lemna minor use in rhizofiltration? Since the Lemna minor was
shown to suffer no ill effects from absorbing excess phosphorus, it could be considered a viable
solution for treating bodies of water with an excess of phosphorus. New problems could arise
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from using Lemna minor as a rhizofiltration method in bodies of water. Lemna minor is very
invasive and faster growing by nature.4 If allowed to grow under conditions that expedited its
growth, such as with excess phosphorus, Lemna minor would eventually cover the top of a lake
or pond entirely, taking sunlight and nutrients from other organisms in the population.4 This
could lead to just as much damage, and possibly even more, than the harmful substances in the
water originally were. To counter this, the Lemna minor population treating any body of water
would need to be carefully monitored and kept at a number where it could successfully remove
excess phosphorus from the water without overpopulating and killing off other species.
Sources of Error
Many different factors could have affected the accuracy at some points of the experiment.
At the beginning of the experiment, the started populations were set by number of plants, but the
number of thalli was recorded for each population at each time interval. This could have affected
experiment one, as the thalli were not in a two-to-one ratio to start the experiment, only the
plants were.
The Lemna minor populations could have been affected by the amount of algae in the
containers. Since the algae were only cleared out every other day, the algae that were present on
days when the containers were not cleaned could have negatively affected the plant growth.
These algae could have stolen significant amounts of sunlight and/or nutrients.
Future Experimentation
Many changes could be made to these experiments to make the results more accurate or
more widespread. Counts of the thalli of the Lemna minor could be taken more than once a week
or even daily to ensure more accurate growth rates, maximum intrinsic rates of growth, and
carrying capacities. Different nutrients such as sulfur, fluorine, and chlorine could be tested to
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see if the plants could successfully survive while removing these elements from the water. The
experiment could also be done in larger containers to see how the growth rate and carrying
capacity change when allowed to grow for a longer time over a larger area.
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REFERENCES
1) Salt, D. E., Smith, R. D., & Raskin, I. (1998). Phytoremediation. Annual review of plant
biology, 49(1), 643-668.
2) Hass, C.A., D. Burpee, R. Meisel, and A. Ward. 2013. A Preliminary Study of the Effects of
Excess Nutrients and Interspecies Competition on Population Growth of Lemna minor
and Salvinia minima in a Laboratory Manual for Biology 220W: Populations and
Communities. (Burpee, D. and C. Hass, eds.) Department of Biology, The Pennsylvania
State University, University Park, PA.
Adapted from Beiswenger, J. M. 1993. Experiments to Teach Ecology. A Project of the
Education Committee of the Ecological Society of America. Ecological Society of
America, Tempe, AZ. pp. 83-105.
3) Lemna Minor L. (2002, December 13). Retrieved April 7, 2013, from Germplasm Resources
Information Network: http://www.ars-grin.gov/cgi-bin/npgs/html/taxon.pl?400078
4) Lemon, G. D., Posluszny, U., & Husband, B. C. (2001). Potential and realized rates of
vegetative reproduction in Spirodela polyrhiza, Lemna minor, and Wolffia borealis.
Aquatic Botany, 70(1), 79-87.
5) Leng, R., Stambolie, J., & Bell, R. (1995, October). Duckweed - a potential high-protein feed
resource for domestic animals and fish. Livestock Research for Rural Development, 7(1).
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