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Sammy Quiroz, Savannah Stowe, LeighAnn Halik, Sana Navaid

Mrs. Norris

AP Environmental Science

02 March 2017

LD50 Lab Report

Introduction

The LD50 of a substance, “the individual dose required to kill 50% of a population of

organisms” (Environmental Protection Agency), is a necessary calculation that must be made by

researchers in many fields of science. In particular, the calculation of LD50 is most necessary in

pharmacology in order to accurately predict the mortality risk for patients when prescribed a

certain drug. The data from LD50 experiments is used as a precaution when prescribing certain

doses of drugs and/or medications in order to ensure a low risk and no chance of death. The

calculations found through the collection of LD50 are represented by a dose-response

relationship and are recorded on a dose-response curve. The curve also demonstrates the

threshold dose for the given substance. The threshold is the point of the graph where the

response is first observed (Barron's, Thorpe). In addition to using LD50 in order to measure the

risks in pharmacology, it can also be used for finding the toxicity of common pollutants. In the

past, salt has been used as a preventative measure for keeping roads from getting icy in the

winter time. What had not been considered is the point at which the salt causes harm to the

environment. Salt from the roads has eventually gotten into runoff, and infiltrated the ground

water. The polluted soils have had harmful effects on vegetation. This information led to the

question, “at what point is the salt concentration in water harmful to surrounding vegetation?” In
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order to find a solution to this question, an experiment was conducted in which radish seeds were

grown in petri dishes in water diluted to different salt concentrations. Through the calculation of

LD50 and observation of the threshold of negative effects of salt in the radish seeds, the question

is resolved, and an answer is found that can be used to accurately define the risk of using salt to

ice roads in a real-life situation.

Purpose

To determine the effect of differing salt concentrations on the germination and radicle

root growth of radish plants.

Hypothesis

If 60 radish seeds are placed into 6 different petri dishes with varying degrees of salinity

in the water, then the data will show the LD-50 of these radish seeds occurs at a 25% salt

concentration. In addition, as salt concentration in the water increases, the radicle root length will

decrease by 10% for each increasing concentration.

Materials

1. 10 mL graduated cylinder

2. Concentrated salt solution

3. 24 circular pieces of paper towel

4. 60 radish seeds (10 per petri dish)

5. Calculator

6. 6 petri dishes with lids

7. Masking tape to label everything

8. Water
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9. Metric ruler

10. Pencil

Procedure

1. Using 20 ml of the original concentration of salt perform a serial dilution to create five

different salt concentrations, plus a sixth with distilled water for the control.

2. Count out 60 seeds, 10 for each water sample

3. Put 10 seeds into a folded paper towel, space the seeds out evenly on the paper towel so

that they do not touch each other

4. Wet the paper towel/seeds and seal in a baggie, be sure to label the bags with the salt

concentration for that sample

5. After 4-5 days examine the seeds, counting the number of each that germinated. You

will measure the response of the radish seeds at various salt concentrations.

6. Remove the lid of the control dish. Count the number of seeds that germinated.

Calculate the percentage of seeds that germinated and

record in Table 2. ​Note: if fewer than 80% of the seeds in

this ​control sample​ germinate, this indicates a problem with

the experiment.

7. Measure the length of the radical for each of the germinating radish seeds to the nearest

millimeter (mm). Look carefully at each sprout to make sure you are measuring just the

root, not the shoot as well. In the picture below, you would measure just the part between

the two arrows, not the shoot and cotyledons to the left.

8. Repeat steps 6-7 for each petri dish.

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9. For each treatment, calculate the mean radicle length for each salt solution. Add the total

radical lengths for each salt solution and divide by the total number of seeds that

germinated. ​Do not include​ data from seeds that did not germinate. Record data in

column labeled, “Mean Radicle Length (mm).”

Data and Analysis

a. Data

Table 1: Root Length of Radishes Grown

Concentrati # of Radicle Root Length (mm)

on of Salt seeds
germina 1 2 3 4 5 6 7 8 9 10 T= mean Difference
ted (out Radicle in Radicle
of 10) Length Length C-T
(mm)

Control 10 80 75 110 95 90 121 172 143 114 142 114.2 0

0%

6.25% 9 96 65 60 85 50 120 83 70 115 N/A 82.67 76.42

12.5% 4 50 10 65 60 N/A N/A N/A N/A N/A N/A 46.25 33.75

25% 4 31 42 15 16 N/A N/A N/A N/A N/A N/A 20.8 4.2

50% 0 N/A N/A N/A N/A N/A N/A N/A N/A N/A N/A 0 N/A

100% 0 N/A N/A N/A N/A N/A N/A N/A N/A N/A N/A 0 N/A
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Fig 1: Radical Root Length for Various Salt Concentrations (Comparison with Class)

Fig 2: LD-50 of Salt Concentration upon Radishes

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b. Analysis

The table above contains data collected from the petri dishes on the amount of radishes

that germinated depending on the amount of salt concentration in the water that was used. Each

petri dish contained a different amount of salt concentration within the water used; 0%, 6.25%,

12.5%, 25%, 50%, and 100%.

After collecting data from the class averages and comparing

them to the group averages of radish seed germination, basic trends

were discovered. Overall, a minimum of 50% salt concentration

caused no radish seeds to germinate. According to the class

averages, at an average of around 12-13%, half of the radish seed

populations were ungerminated. As shown (Table 1), 100% of the

radish seeds germinated in the control group due to the purity of the

water. There is a strong correlation between salt concentration and the radical root lengths of the

radishes. It was discovered that using higher amounts of salt-concentrated water impeded the

germination and growth of radish roots. This explains why 90% of the radish seeds germinated

with a 6.25% salt concentration, while 40% of the radish seed population germinated with a 25%

salt concentration… the salt reduced germination.

After transferring the group data of radish germination onto a dose-response curve, the

LD-50 of salt concentration on radish seeds was 12%. This indicates that if a population of

radish seeds were exposed to 12% salt-concentrated water, 50% of the population would not

germinate.
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Conclusion

The results from the lab differed from the initial hypothesis, in that half of the radish seed

population exposed to 12% of salt-concentrated water would die. This contradicts the original

hypothesis, as it was first predicted that 25% salt concentration would cause 50% of the radish

population to die. From this lab, it was discovered that the control group would have higher

radical root lengths than seeds that germinated with a certain amount of salt concentration. The

results from the lab also disproved the original hypothesis, as it was initially predicted that as the

salt concentration increased, the radicle length would decrease by 10%. However, after

collecting data of radical roots, it was found that the length decreased by 28%, then 44%, and

then 55%. Instead of decreasing by a constant 10% each time, the radical lengths decreased

unsteadily, as the results varied by an even larger amount between each different level of salt

concentration. The reason for these unexpected results is that salt concentration had a much

larger impact on germination and root length than anticipated.

Possible sources of error include the process of taking the germinated radishes out of the

petri dishes. A few of the radish roots were broken during the process of disentanglement from

the paper towel, after being taken out of the petri dishes. This may have lead to inaccurate root

length measurements. The issue could have been avoided if the radish roots were not so tangled

and intertwined, or if more caution had been taken while collecting data. If the radish roots were

unraveled more smoothly with minimal tangles, the measurements of the roots would have been

more accurate. Also, instead of using fingers to place the radish seeds into the petri dish, tongs

could have been used in order for there to be fewer possible sources of contamination. Possible
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contamination of the seeds by germs on fingers used may have altered the growth of the radish

roots.

In this experiment, the initial purpose was achieved by finding the effect of differing salt

concentrations upon the germination and radical root growth of radish seeds. In comparison to

the rest of the class, measurements from ‘Group 7’ (Halik, Navaid, Quiroz, Stowe) were most

similar to those of “Group 3”. Compared to other groups, Group 7’s radish roots were much

longer, excluding roots grown under 50% and 100% salt concentration. This may have been

attributed to the overall health of the radish seeds prior to the distribution of seeds to different

groups for experimentation.

This lab may be related to current environmental issues, such as the amount of

application of salt on icy roads. Not only does the amount of salt effect snow and/or ice being

melted, but also the organisms and habitats surrounding the roads, as runoff is common. In

addition, road salt effects the potential toxicity of groundwater. The Environmental Protection

Agency recommends that levels be kept below 230 mg/L, which is measured over a course of

four days (Environmental Protection Agency, 2017). Also, soil salinity affects agricultural land

during the process of crop cultivation. If soil salinity is too high, crop growth is negatively

impacted, as it may be slowed or halted.

Overall, this lab has exemplified how various salt concentrations impact organisms, as

well as crops; such as radishes. In both the group and class data, a strong correlation between the

salt concentrations and germination/radical root length was found. Both data sets found that the

root lengths of radishes in waters of lower salt concentrations were greater than the root lengths

of radish seeds in water with a higher salt concentration, or did not germinate at all. “Deicing salt
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is usually refined rock salt, consisting of about 98.5 percent sodium chloride, 1.2 percent calcium

sulfate, 0.1 percent magnesium chloride, and 0.2 percent rock.” (Perry, Leonard). That being

said, road salt consists of 98.5% indicating a clear potential risk in the use of road salt upon the

environment. At 50%, no germination was found in the radish seeds, therefore, with a salt

concentration of 98.5%, the potential environmental consequences could be unprecedented.

Works Cited

"Food Agricultural Salt." ​Salt Institute​. Web. 01 Mar. 2017.

Gould, Ann. "Impact of Road Salt on Adjacent Vegetation." ​Plant & Pest Advisory​. 10 May

2016. Web. 16 Feb. 2017.

Miller, Lonnie. Salinization Lab, Teacher Lab Template. College Board, 2003. PDF.

Perry, Leonard. "Salt Damage to Plants." University of Vermont. Web. 13 March. 2017

"Snow and Ice." ​EPA​. Environmental Protection Agency, 06 Jan. 2017. Web. 27 Feb. 2017.

Stromberg, Joseph. "What Happens to All the Salt We Dump On the Roads?" ​Smithsonian.com​.

Smithsonian Institution. Web. 09 Feb. 2017.

Thorpe, Gary S. ​Barron's AP Environmental Science​. Hauppauge, NY: Barron's, 2013. Print.

Zielinski, Sarah. "Earth's Soil Is Getting Too Salty for Crops to Grow." ​Smithsonian.com​.

Smithsonian Institution. Web. 09 Feb. 2017.