1. Name: Aditya Parmar Date: 15th November 2010
Supervisor: Dr. Olga Calvo di Alegre
Institute: Institute of Landscape and Plant Ecology.
Experiment: Lemna spp. (Duckweed) Growth Inhibition Test with 3,5-DCP as toxin (Inhibitor)
Introduction
Lemna plant was used to characterize the effect of different concentrations of 3,5-DCP on its growth. This aquatic plant was used due to
its high representation in the ecosystem and its unique characteristics of being small in size, simple structure, asexual reproduction,
genetic uniformity and short generation time. None the less, plant reduces the use of animal (for e.g. Fish) for ecotoxicological tests.
Principle
Lemna shows a significant reduction in growth when exposed to different organic and inorganic toxins. The inhibited growth is
depicted in terms of lower frond number, lower frond area and lower quantity of pigments such as chlorophyll.
Methodology
The experiment was carried out in a time period of 1 week. Day 1, five concentrations (0, 1.25, 2.5, 5, 10,20mg/L) were prepared to
expose Lemna plant to the toxin (3, 5-DCP). Equal numbers of frond were added to all six samples. Day 3, frond area, frond number
and appearance was noted down in the data sheet. The nutrient solution was changed, samples were rotated and ph levels were
measured again to compare them with Day 1 readings. Day 7, again frond number, frond area and appearance was noted down in the
datasheet. The ph was also checked, on Day 7 chlorophyll pigment levels were calculated with SPAD test for different concentrations
of toxin.
Results
Frond Numbers and Frond areas were plotted against different doses of the 3,5-DCP concentrations in point graphs as Figure 1 and
Figure 2 respectively, showing variation for duration and concentration relationship to frond area and frond number. On the last day
chlorophyll pigment was also measured and represented in a bar graph in Figure 3.
Mean Frond Numbers vs Concentration- Figure 1 Area vs Concentration - Figure 2
30 4.00
3.50
25
3.00
Frond Ares ( cm2)
Frond Numbers
20 2.50
15 20, 15 Fronds Day 3 2.00
20, 1.78
1.74
10
20, 12 Fronds Day 7
1.50 Day 3
1.00 Day 7
5 Fronds Day 1
0.50
0 0.00
0 5 10 15 20 25 0 5 10 15 20 25
(mg/L) 3,5-DCP (mg/L) 3,5-DCP
SPAD Vs Concentration - Figure 3 NOTE:
From the above results we can describe the reduced growth of
25
Lemna plant at high concentration of toxin. Figure 1 and 2 shows
20
15
that at more than10ml/L 3, 5-DCP the plant has shown almost no
10
growth in terms of increase in frond numbers and frond area after
5 the 3rd day of the experiment.
0
1 2 3 4 5 6
The plant has shown a steep decline in chlorophyll levels with
increasing concentration of toxin. Figure 3 show how pigment
Concentration Mean SPAD level decreased with the increasing concentrations of 3,5-DCP.
Conclusion and Discussion
The experiment showed the reduction in growth as an influence of toxin on plant as expected. Lemna plant responded to higher
concentration in a larger influence on its frond number and area. The increasing reduction in pigment levels were also depicted with
increasing concentrations of toxin. The test clearly gave an indication that Chlorinated Hydrocarbons (CHC) significantly reduces the
growth and pigment levels at higher concentrations (more than 10ml/L). This test might not give accurate indication for very low
concentrations of toxins, as at lower concentrations the significance was much less.
Calculation of frond number and area could be a source of possible mistake if not done in a proper disciplined manner. During the
experiment some of the samples had difficulties in estimation of area as the pictures took were not of very good quality. However, test
gave appropriate results in the end and users were satisfied with the estimates.