The objective of the Pharmacology Lab is to find the lethal chemical(s) or the LD50 relevant to a certain genus. In order for a chemical to be considered the LD50, 50% of the population of the organism present in a container must die. A total of three trials and four tests are completed to determine whether or not the chemical has the ability to kill the brine shrimp. The three trials contain 5mL of the chemical in each container, while the four tests have different amounts of the chemical in each container. The amount of water used for both the trials and tests remain constant at 40mL. Once the correct amount of the required substances and brine shrimp are placed into their respective and labeled containers for both trials and tests, they are then moved under a desk light. For the next duration of time, the death rate of the brine shrimp and the physical or behavioral changes visible are observed and recorded. After performing the experiments, the data shows that a great number of these selected regular household items have deadly chemicals in them that can kill other living organisms if given the right amount of dosage, including humans. Out of all five products, the LD50 was only found in soda and hydrogen peroxide. Lemon juice, nail polish remover, and hand sanitizer were not considered the LD50 because the death rate percentage was higher than 50%. This research provides a better understanding of how toxic household chemicals are. Products that were created for consuming such as the Lemon Juice and Soda were found to instantly and over a certain amount of time, kill the brine shrimp once in contact.
II. Introduction
In this experiment, one of the main purposes is to find the LD50 of several household chemicals on Brine Shrimp. LD means lethal dose of a certain product. The LD50 formula gives the estimate of deaths of half a population when receiving the molarity of each chemical. A possible solution to this problem is to perform and experiment where an organism is tested on with different types of chemicals for the death rate. Brine shrimp, the genus used, are aquatic crustaceans that can be grown by adding water and brine shrimp eggs into a tank. This process takes approximately two days. Brine shrimp lives in salt water, but can also survive in regular water for 24 hours. Completing this experiment determines whether they can survive a certain about of chemicals in different time intervals. This lab contains five different chemicals including: nail polish remover, hydrogen peroxide, lemon juice, hand sanitizer, and lemon soda. These household chemicals are tested as the positive controls, while the negative control is water. With five positive controls, there would be different results from each chemical. As the experiment takes place through four tests; each containing 3 trials; the observation of the deaths of brine shrimp demonstrates the amount of time they can survive in new obscure habitats.
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The test of each chemical shows whether there are traces of deadly substances, which could help find the measures of toxicity in each chemical. Testing the toxicity by molarity and time shows the maximum amount of toxicity that humans can handle to be able to survive. The experiment is an example of how living organisms react to chemicals. The brine shrimp will play the role of the organism. However, different organisms will have different reactions depending on their adaptations and physical traits. As the brine shrimp take in the chemicals, the inability to survive increases. The death of the aquatic crustaceans shows how toxic harmless household chemicals can truly be.This lab will explore the ingredients from the five chemicals used. The observations of the LD50 will determine which chemical is deadlier than the others, causing the public to be more aware of these fatal factors.
III. Methods:
The brine shrimp are grown by adding water and brine shrimp eggs into a tank. Brine shrimp takes two days to mature enough to be experimented on. The lethal chemical(s) or the LD50 are found by placing different substances mixed with water into containers with nine brine shrimp. Through a series of tests and trials, the lethal chemicals are distinguished. A total of three trials and four tests are completed to determine whether or not the chemical has the ability to kill the brine shrimp. The three trials contain 5ml of the chemical in each container, while the four tests have different amounts of the chemical in each container; (5ml, 10ml, 15ml, and 20ml). The amount of water used for both the trials and tests remain constant at 40ml. Once the correct amount of the required substances the brine shrimp are placed into their labeled containers for both trials and tests. Next, the Brine shrimp are moved under a desk light because they are attracted to any form of light. The death rate and physical or behavioral changes of the brine shrimp are observed and recorded in tests with intervals of 10-minute trials and 5-30 minutes.
This lab consists of four tests and three trials for five different chemicals to find the LD50. The positive controls include: Super Chill Lemon Lime soda, hydrogen peroxide, House Recipe lemon juice, 2.2 oz. of Assured Aloe Vera scented hand sanitizer and 2.2 oz. of ONYX Professional lavender scented nail polish remover, while the negative control is water throughout the entire experiment. For each test, there are three trials performed beforehand. Twenty seven brine shrimp are needed for the trials. For each of the three trials, nine brine shrimp are obtained from the twenty seven. Then, 40mL of distilled water is added to the brine shrimp container through pipettes and graduated cylinders; the total volume of water for each trial is 40mL in order to maintain consistency. From each chemical, 5mL are added into the labeled containers of the 40 mL distilled water. Once all substances are added, the containers are placed under direct light from a desk lamp because brine shrimp are attracted to light. Through five tests after the three trials, various amounts of nail polish remover, hydrogen peroxide, lemon juice, hand sanitizer, and Sprite soda are added to its assigned containers. For each test, there will be four containers for the different amounts used of each chemical. The first container contains 5mL of the chemical while the other three containers will have an increase of 5mL from the prior container. This process will be done five times for the five chemicals used. The counts of deceased shrimp after each time interval are recorded over a course of 5 minutes to 30 minutes in percentages.
Different reactions from the brine shrimp were recorded in the data table as well as pictures being taken. This procedure was used according to the outline given in order to achieve the objective of this lab. Given a certain the amount of time, three trials and four tests were used. In the procedure, several changes were included, instead of adding 40ml of water with the nine brine shrimp, 40ml of water was added to the desired chemical first to have enough time to collect the brine shrimp for the specific trial and test.
IV. Data
Data table for Trial: Peroxide (3 trials total) @ (5ml of chemical) PH: 7
Time interval
Observations
Deaths (%)
Deaths (#)
10 mins
move erratically; attracted to light; hitting against the cup
0%
0
10 mins
move erratically, attracted to light
11%
1
10 mins
move erratically, attracted to light
11%
1
Data table for Test: Peroxide – 5mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
has bubbles
0%
0
10 min
has bubbles
0%
0
15 min
has bubbles
22%
2
20 min
has bubbles
22%
2
25 min
has bubbles
33%
3
30 min
has bubbles
33%
3
Data table for Test: Peroxide – 10mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
has bubbles
44%
4
10 min
has bubbles
44%
4
15 min
has bubbles
44%
4
20 min
has bubbles
44%
4
25 min
has bubbles
44%
4
30 min
has bubbles
44%
4
Data table for Test: Peroxide – 15mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
has bubbles
33%
3
10 min
has bubbles
44%
4
15 min
has bubbles
44%
4
20 min
has bubbles
55%
5
25 min
has bubbles
55%
5
30 min
has bubbles
55%
5
Data table for Test: Peroxide – 20mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
has bubbles
33%
3
10 min
has bubbles
33%
3
15 min
shrimp turn orange
44%
4
20 min
has bubbles
55%
5
25 min
has bubbles
55%
5
30 min
has bubbles
55%
5
Data table for Trial: Nail Polish Remover (3 trials total) @ (5ml of chemical) PH: 7
Time interval
Observations
Deaths (%)
Deaths (#)
10 mins
hitting against the cup under the light
22%
2
10 mins
hitting against the cup under the light
22%
2
10 mins
hitting against the cup under the light
22%
2
Data table for Test: Nail Polish – 5mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
has bubbles
11%
1
10 min
has bubbles
22%
2
15 min
has bubbles
22%
2
20 min
has bubbles
55%
5
25 min
has bubbles
66%
6
30 min
has bubbles
66%
6
Data table for Test: Nail Polish – 10mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
has bubbles
100%
9
10 min
has bubbles
100%
9
15 min
has bubbles
100%
9
20 min
has bubbles
100%
9
25 min
has bubbles
100%
9
30 min
has bubbles
100%
9
Data table for Test: Nail Polish – 15mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
has bubbles
100%
9
10 min
has bubbles
100%
9
15 min
has bubbles
100%
9
20 min
has bubbles
100%
9
25 min
has bubbles
100%
9
30 min
has bubbles
100%
9
Data table for Test: Nail Polish – 20mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
has bubbles
100%
9
10 min
has bubbles
100%
9
-15 min
has bubbles
100%
9
20 min
has bubbles
100%
9
25 min
has bubbles
100%
9
30 min
has bubbles
100%
9
Data table for Trial: Lemon (3 trials total) @ (5ml of chemical) PH: 2
Time interval
Observations
Deaths (%)
Deaths (#)
10 mins
some floats
100%
9
10 mins
transparent with some orange
100%
9
10 mins
completely transparent
100%
9
Data table for Test: Lemon – 5 mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
completely transparent
100%
9
10 min
completely transparent
100%
9
15 min
completely transparent
100%
9
20 min
completely transparent
100%
9
25 min
completely transparent
100%
9
30 min
completely transparent
100%
9
Data table for Test: Lemon – 10mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
completely transparent
100%
9
10 min
completely transparent
100%
9
15 min
completely transparent
100%
9
20 min
completely transparent
100%
9
25 min
completely transparent
100%
9
30 min
completely transparent
100%
9
Data table for Test: Lemon – 15mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
completely transparent
100%
9
10 min
completely transparent
100%
9
15 min
completely transparent
100%
9
20 min
completely transparent
100%
9
25 min
completely transparent
100%
9
30 min
completely transparent
100%
9
Data table for Test: Lemon – 20mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
completely transparent
100%
9
10 min
completely transparent
100%
9
15 min
completely transparent
100%
9
20 min
completely transparent
100%
9
25 min
completely transparent
100%
9
30 min
completely transparent
100%
9
Data table for Trial: Hand Sanitizer (3 trials total) @ (5ml of chemical) PH: 7
Time interval
Observations
Deaths (%)
Deaths (#)
10 mins
shrimp turns orange
33%
3
10 mins
all on the bottom
33%
3
10 mins
against the wall
22%
2
Data table for Test: Hand Sanitizer – 5 mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
turns orange
66%
6
10 min
turns orange
66%
6
15 min
turns orange
77%
7
20 min
turns orange
77%
7
25 min
turns orange
100%
9
30 min
turn orange
100%
9
Data table for Test: Hand Sanitizer- 10mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
turns orange
22%
2
10 min
turns orange
33%
3
15 min
turns orange
55%
5
20 min
turns orange
55%
5
25 min
turns orange
88%
8
30 min
turns orange
100%
9
Data table for Test:Hand Sanitizer- 15mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
turns orange
44%
4
10 min
turns orange
66%
6
15 min
turns orange
66%
6
20 min
turns orange
66%
6
25 min
turns orange
77%
7
30 min
turns orange
100%
9
Data table for Test: Hand Sanitizer- 20mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
turns orange
88%
8
10 min
turns orange
88%
8
15 min
turns orange
100%
9
20 min
turns orange
100%
9
25 min
turns orange
100%
9
30 min
turns orange
100%
9
Data table for Trial: soda (3 trials total) @ (5ml of chemical) PH: 7
Time interval
Observations
Deaths (%)
Deaths (#)
10 mins
bubbles
22%
2
10 mins
some are orange
22%
2
10 mins
some are orange
22%
2
Data table for Test: soda – 5 mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
bubbles
11%
1
10 min
bubbles
11%
1
15 min
bubbles
11%
1
20 min
bubbles
11%
1
25 min
bubbles
22%
2
30 min
bubbles
22%
2
Data table for Test: soda – 10mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
bubbles
55%
5
10 min
bubbles
55%
5
15 min
bubbles
55%
5
20 min
bubbles
55%
5
25 min
bubbles
55%
5
30 min
bubbles
55%
5
Data table for Test: soda- 15mL
Time Interval
Observations
Deaths (%)
Deaths (#)
5 min
bubbles
44%
4
10 min
bubbles
44%
4
15 min
bubbles
55%
5
20 min
bubbles
66%
6
25 min
bubbles
66%
6
30 min
bubbles
77%
7
Data table for Test: soda – 20mL:
Time Interval
Observations
Death (%)
Deaths (#)
5 min
bubbles
88%
8
10 min
bubbles
88%
8
15 min
bubbles
88%
8
20 min
bubbles
88%
8
25 min
bubbles
88%
8
30 min
bubbles
88%
8
V. Analysis
The LD50 for hydrogen peroxide is approximately 0.24 molars. Hydrogen peroxide has 3% Hydrogen Peroxide (Stabilized), Purpose First Aid Antiseptic/Oral Debriding Agent. Its Inactive Ingredient is Purified Water. Hydrogen peroxide is a deteriorizing agent that is commonly used as a hair bleach when diluted. An Oral debriding agent describes what the hydrogen peroxide can do. This means that the hydrogen peroxide when diulted in water can help heal minor mouth and gum irritation. Since all living organisms have an enzyme called peroxidase, it helps decompose low concentrations hydrogen peroxide into water and oxygen.
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There is no LD50 for nail polish remover. Nail Polish remover has the ingredients: Acetone, aqua, denatonium benzoate, tocopheryl acetate, aloe barbadensis leaf juice, parfum(means perfume in french), Cl 45410(dye), Cl 60730(dye). Denatonium benzoate is a chemical compound known to prevent and discourage people from drinking harmful products. Tocopheryl acetate, also known as vitamin E acetate provides beneficial antioxidant effects. Aloe barbadensis leaf juice, also know as Aloe Vera is a plant. When consumed, it can help relieve irritable bowel syndrome and heartburn. Acetone is an organic compound that is a colorless and flammable liquid. Concluding that acetone is the lethal ingredient and causes the death of the brine shrimp when ingested, it will cause liver problems, which causes the liver to have a hard time breaking them down. Large intake of acetone can lead to unconsciousness and death.
There was no LD50 for Lemon; the ingredients in the lemon juice packets include water, lemon juice concentrate, sodium benzoate, sodium bisulfite, and lemon oil. Sodium Benozoate is a type of salt often used a preservative because it works well at killing bacteria, yeast, and fungi. Sodium bisulfite is a food addictive. When in contact with chlorine bleach, it will release harmful fumes.The brine shrimp died within 20 minutes because of the sodium benzoate in the lemon juice packet, which is a type of salt mixed together with vitamin C causes a carcinogen substance called benzene. Benzene is a carcinogen because when exposed into the air, it can cause leukemia. Eating or drinking substances with high levels of benzene can cause vomiting, dizziness, sleepiness, and or death. It may not be lethal enough to kill humans yet, but having a high amount of the lemon juice packets can potentially kill a human or any living organism.
There is no LD50 for Hand Sanitizer but the graph decreases and increases because there was a slight change in the number of brine shrimp needed in each container. The ingredients contained in this chemical are: water, triethanolamine, glycerin, propylene glycol, aloe barbadensis gel(another form of aloe vera), carbomer, fragrance, blue 1(dye), yellow 5(dye), and 62% ethyl alcohol. Triethanolamine is a chemical compound that occasionally causes contact allergy. Glycerin is a simple alcohol compound that is used as a sweetener and sometimes a preservative in food and beverages. Propylene glycol is an organic compound used in many applications such as a food addictive called E number 1520, moisturizer in medicine, and a solvent for food coloring. Carbomer is a synthetic polymer that forms an adhesive eye gel. 62% ethyl alcohol is a flammable liquid used as solvents and fuel. The brine shrimp died because of the 62% ethyl alcohol in the hand sanitizer, when consumed can cause death. Although used in wine and any alcoholic drinks, the drinks only contain small concentrations of alcohol, legal enough to drink.
The LD50 for Super chill lemon lime soda is approximately 0.19 molars. The ingredients in the soda are carbonated water, high fructose corn syrup, citric acid, natural flavors, potassium citrate and potassium benzoate. High frutose corn syrup is comprised of many corn syrups that has undergone enzymatic(enzymes that catalyze) process that converts glucose into frutose for desired sweetness. There have been health concerns about raising the risks of diabetes and cardiovascular disease. Citric acid is a weak organic acid that is a natural preservative and used to add a sour taste to food and soft drinks. Contact with dry citric acid can result in skin or eye irritation. potassium citrate is a food addictive and used to regulate acidity. It is also a potential health hazard when administered incorrectly. Elevated blood levels of potassium citrate can result in heart attacks and death. Potassium benzoate is a food preservative that inhibits the growth of yeast, mold, and some bacteria. The reason why the brine shrimp died because of potassium citrate in the soft drink. Too much potassium citrate can cause irregular heartbeat and a risk of having a heart attack. Although there are potassium citrate pills sold in stores, they are only recommended if the person had been having a case of fatigue or muscle weakness. The kidney is responsible for eliminating excess potassium in the body.
The molarity formula: iM1V1=1M2V2 was used to find the LD50 and true concentration of each chemical after all the substance has been added to container and totaled. The LD50 for Hydrogen Peroxide is 0.24 molars. The LD50 for Super Chill lemon lime soda is 0.19 molars. There are no LD50 for lemon, nail polish remover, and hand sanitizer because the deaths were higher than fifty percent. The standard deviation at 20 ml for Hydrogen Peroxide is approximately 0.833, for Nail Polish remover, lemon, hand sanitizer, and soda is 0. The chemicals included in this experiment are normally used in everyday use in the household. Chemicals such as peroxide are used to clean injuries or cuts. Hand sanitizer is used to clean 99.9% of bacteria on hands. Nail polish remover is used for getting the nail polish off of the fingernails. Lemon juice packets are used in making the dressing for a salad. The LD50 for Nail Polish remover is 1.159 g/kg for humans when digested. Unfortunately, the LD50 for the rest of chemicals are unavailable because they have only been conducted on rats and mice.
Although the results of the data were both consistent and gradual in terms of death percentages, a few of the brine shrimp died before 5 minutes of data was recorded. This indicated that the brine shrimp could not live in the high toxicity levels on some of the chemicals used in this experimentation even with the addition of water.
VI. Conclusion
This lab is significant because the participants are able to discover the true dangers of certain household products such as hydrogen peroxide, soda, hand sanitizer, lemon juice, and nail polish remover. With everyday household chemicals, a person’s health can be affected, thus finding the LD50 will tell us how toxic a certain item is. The use of the LD50 is to study the amount of a chemical that will kill 50% of the tested animal, which will show us how dangerous a product can truly be. Some insights include the participants testing the dangers of “harmless†products. The lab offered similarities and an opportunity for a hands on experience of the procedures a toxicologist, like finding the cause and effects of poison. By using mathematical formulas like the molarity formula and the standard deviation formula, accurate and reliable data was collected to find the true concentration of the LD50 chemicals. For example, the data and calculations from these formulas show that one of the chemicals, Hydrogen Peroxide is the LD50 with a molarity of 0.24 molars. The data tables show that chemicals like the lemon juice, nail polish remover, and hand sanitizer are not the LD50. This is most likely due to the fact that these products contain amounts of citric acid and alcohol which results in death rates that are over 50%. In the trials and tests, many occurrences of errors and mistakes which could have affected the outcome of the results. In one of the hand sanitizer tests, 18mL was added into the mixture of substance instead of 20mL. Instead of counting the number of death at a certain time, some of the intervals were missed by a minute or two. This lab was not as accurate as expected because of the difficulties experienced during the beginning of the lab. For example, lotion was one of the chemicals to be tested on the brine shrimp prior to the lab. However, because the lotion was dense, it was difficult to mix with the water. Also, the color made it nearly impossible to see and count the brine shrimp. These errors could be easily avoided in the future by double checking the amount of a substance being added, keeping track of time, and using transparent products. For a clearer understanding several alterations of a procedure outline, being careful with the amount of measurements and constant data collection could be applied in the future.
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