Plant Defensive Compounds Lab Report PDF

Preview

Summary

Download Plant Defensive Compounds Lab Report PDF

Description

Running Head: EFFECTS OF PLANT DEFENSIVE COMPOUNDS ON HEART RATE

1

Effects of Plant Defensive Compounds on Heart Rate Alyssa Alonso BIOL 1510 DO1 Professor Eldakar Yvanna Strait October 18, 2019

Abstract The objective of this experiment was to examine the effects of various plant defensive compounds, milkweed and Cuban tilo extract, on the leg flapping rate, to be used as a measure of heart rate, of Artemia Salina, a small invertebrate that thrives in salt water. The mean heart rate for the control, sea water, was 150 fpm, whereas the mean heart rate for milkweed was 194.67 fpm and 140.17 fpm for the Cuban tilo extract. There was a significant heart rate increase in A. Salina exposed to milkweed versus control (T milkweed,control = 3.54, T crit 11,0.05 = 2.20). In comparison to milkweed and control, tilo had no significant effect on heart rate (T milkweed,tilo = 4.86, T crit 11,0.05 = 2.20); (T tilo,control = 0.72, T crit 11,0.05 = 2.20). These results suggest that milkweed, but not tilo, significantly increases heart rate in exposed A. Salina.

EFFECTS OF PLANT DEFENSIVE COMPOUNDS ON HEART RATE

2

Introduction A. Salina is a small invertebrate, also known as a brine shrimp, that thrives in salt water and is characterized by a body with a discrete head and compound eyes, a number of leaflike limbs, and a slender abdomen. They use their leaflike limbs to swim, rhythmically beating the water to create thrust (The Editors of Encyclopedia Britannica, 2019). These small crustaceans also feed upon green algae and can regulate the salt balance within their bodies (Urry et. al., 2016). The organism’s heart rate, which is being tested in this experiment, can be affected with exposure to various plant defensive compounds. These compounds can be found in all plants to protect themselves from herbivory. However, plants protect themselves in two different ways, through structural defenses and/or chemical secretions. The structural defenses may be anything from thorns to spines, that allow the plant to ward off larger herbivores, who cannot maneuver around such structures (Bio II Lab Manual, 2019, Pg. 1). Chemical defenses, also known as plant defensive compounds, are primarily involved in preventing smaller organisms, such as insects, from destroying the plants’ leaves or branches. Different plants have diverse chemical compounds, which have varying effects on the organisms they target. Conducting research into these different compounds’ effects is vital to the understanding of how they function and how they affect the surrounding population. The chemicals that are secreted by plants for defense are known as allelochemicals. These serve as toxins to either disrupt the behavior, growth/development, or overall survival of whatever herbivore comes in contact with it. The most common category of plant toxins, however, are the alkaloids. These compounds can vary in effects, acting as stimulants, analgesics, depressants, or even psychedelics (Bio II Lab Manual, 2019, Pg. 1). The effects of such substances on humans is well documented, but the effects they have on small organisms is key to the understanding of how plants are able to mount a defense against predators despite lacking the ability to move. In this experiment, we used two varieties of allelochemicals: cardenolide and coumarin, also known as milkweed and Cuban tilo extract respectively. Milkweed is part of the flower family Asclepiadoideae, characterized by milky juice, flowers with five united petals, pod-like fruits, and tufted seeds (The Editors of Encyclopedia Britannica, 2013). Cuban tilo extract, on the

EFFECTS OF PLANT DEFENSIVE COMPOUNDS ON HEART RATE other hand, is a commonly used medicinal plant in Central America and the Caribbean, known for its anti-inflammatory, analgesic, and sedative effects (Roersch, 2018). The objective of this experiment was to examine the effects of various plant defensive compounds on the leg flapping rate of Artemia Salina, the leg flapping being used as a measurement of the organism’s heart rate. Using the information we know about allelochemicals and the properties of milkweed and tilo extract, it can be hypothesized that there will be a significant change in leg flaps per minute as a result of the cardenolide (increase) and coumarin (decrease), compared to control.

Methods In the first part of the experiment, a pipette was used to place a single A. Salina onto a petri dish, making sure to leave enough water for the A. Salina to breathe, but not enough to allow it to swim around the dish. A drop of salt water was then added to the dish and placed under the microscope with the light off. This allowed for the A. Salina and the solution to acclimate to the new chemical addition. After one minute, the light was turned on and the number of leg flaps were counted for 5 seconds. This was used to estimate the heart rate, using the number of leg flaps in 5 seconds, multiplied by twelve, to get an estimate of the beats per minute, in this case, referred to as the fpm. The leg flaps were recorded for a total of three 5second intervals, in order to record a more accurate mean fpm. This was done two more times with two additional A. Salina, for a total of three trials in the control scenario (Bio II Lab Manual, 2019, pg. 4). Following the collection of control data, the effects of milkweed and Cuban tilo extract were tested in the same manner as above (Bio II Lab Manual, 2019, pg. 4). Once all of the trials were conducted, the significance of the data needed to be calculated. This was done using a Ttest, to determine whether or not the mean of the leg flapping differed significantly among the chemical treatments. A statistical significance level of 5% was used.

Results First, the mean heart rate for each treatment needed to be calculated. As can be seen in Figure 1, the control (seawater) had a mean heart rate of 150 fpm, the milkweed had a mean of 194.67 fpm, and the Cuban tilo extract had a mean of 140.17 fpm. The control had a SD of

3

EFFECTS OF PLANT DEFENSIVE COMPOUNDS ON HEART RATE 36.19, milkweed had a SD of 24.56, and finally Cuban tilo extract had a SD of 30.11. Assuming a significance level of 5% and 11 degrees of freedom, the critical value for t was found to be 2.201. A series of T-tests were then conducted to compare heart rates in Artemia Salina exposed to milkweed and Cuban tilo extract, and in an unexposed control, seawater. The mean heart rate for the control, sea water, was 150 fpm, whereas the mean heart rate for milkweed was 194.67 fpm and 140.17 fpm for the Cuban tilo extract. There was a significant heart rate increase in A. Salina exposed to milkweed versus control (T milkweed,control = 3.54, T crit 11,0.05 = 2.20). In comparison to milkweed and control, tilo had no significant effect on heart rate (T milkweed,tilo = 4.86, T crit 11,0.05 = 2.20); (T tilo,control = 0.72, T crit 11,0.05 = 2.20). These results suggest that milkweed, but not tilo, significantly increases heart rate in exposed A. Salina.

The Effect of Plant Defensive Compounds on Leg Flapping in Artemia Salina

Mean Heart Rate (fpm)

250 200 150 100 50 0

Control

Milkweed Extract Treatments

Cuban Tilo Extract

4

EFFECTS OF PLANT DEFENSIVE COMPOUNDS ON HEART RATE

5

Figure 1. Mean heart rate (+/- SD) in A. Salina exposed to milkweed or tilo compared to control.

Discussion The objective of this experiment was to examine the effects of various plant defensive compounds, milkweed and Cuban tilo extract, on the leg flapping rate (heart rate) of Artemia Salina. In doing this, we found that milkweed, but not tilo, significantly increases heart rate in exposed A. Salina. In terms of milkweed extract, the mean heart rate was 194.67 fpm, suggesting that the compound had a largely positive effect on the A. Salina’s heart rate. The t-stat for milkweed was also above the critical value for t (T milkweed,control = 3.54, T crit 11,0.05 = 2.20), further suggesting that it had a significant affect against control. In regard to the Cuban tilo extract, the results clearly showed a much lower mean of 140.17 fpm, suggesting that it had a negative effect on the heart rate of the A. Salina. However, compared to the mean of the control, 150 fpm, the difference was not of enough significance. Furthermore, the t-stat for the tilo extract was below the critical value for t (T tilo,control = 0.72, T crit 11,0.05 = 2.20), further asserting the notion that tilo extract was not as successful in affecting the A. Salina’s heart rate. Similar studies have explored the effects of temperature changes, light intensity, and oxygen consumption on the heart rates of comparable small crustaceans. In a study conducted in 1990, researchers found that heart rate was significantly lower in crayfish at 18oC and higher at 22oC, suggesting that temperature plays a major role in the survival of such organisms (Villareal, 1990). Another study done in 1998, found a positive correlation between heart rate and locomotor activity. Temperature and light intensity also had a positive and negative influence on heart rate, respectively (Bojsen et. al., 1998). Both of these studies show the importance of researching and understanding these organisms, if just to understand how best to use them in experiments such as the one conducted in this report (Rice, 2004). Without knowledge of how

EFFECTS OF PLANT DEFENSIVE COMPOUNDS ON HEART RATE

6

these organisms work, one would not know to use such crustaceans in these examinations. Including further research into the possible medicinal effects that can come from some of these plant defensive compounds in organisms other than small invertebrates. The results found in this experiment partially supported the hypothesis. The milkweed, but not tilo, affected the heart rate of A. Salina compared to control. These results seem to be supported by known scientific facts about milkweed and Cuban tilo extract, however, there may have been some errors that occurred during lab. The amount of each compound added could have been inconsistent. This could be a result of the pipette being used; it could have formed bubbles of the compound rather than actual drops, making it seem as if a drop was added when, in reality, there wasn’t. To correct this, one should tap the pipette to remove any air bubbles before pipetting the compound onto the dish. Another error could have occurred in counting the leg flaps. Such a short counting period contributes to the high chance of error when the leg flapping increases in speed. In order to avoid this issue, future researchers should consider recording the A. Salina and slowing down the footage to accurately count the number of leg flaps. Any further research into this topic should investigate the effects of a wider variety of plant defense compounds on various small organisms. This would allow for a better understanding of these compounds and a more thorough examination into how plants defend themselves. Testing compounds from different parts of the world, in particular, would provide an interesting view into how geographical location also has an impact on production of these compounds and the plants’ adaptive toxins for different locations and organisms.

EFFECTS OF PLANT DEFENSIVE COMPOUNDS ON HEART RATE

References Bio II Lab Manual. (2019) Testing Plant Defensive Compounds. Nova Southeastern University Biology Department Bojsen, B. H., Witthøfft, H., Styrishave, B., & Andersen, O. (1998). In situ studies on heart rate and locomotor activity in the freshwater crayfish, Astacus astacus (L.) in relation to natural fluctuations in temperature and light intensity. Freshwater Biology, 39(3), 455– 465. doi: 10.1046/j.1365-2427.1998.00297.x Rice, S. A. (2004). Brine Shrimp Bioassays: A Useful Technique in Biological Investigations. The American Biology Teacher, 66(3), 208–215. doi: 10.2307/4451655 Roersch C. (2018) Justicia pectoralis Jacq.. In: Albuquerque U., Patil U., Máthé Á. (eds) Medicinal and Aromatic Plants of South America. Medicinal and Aromatic Plants of the World, vol 5. Springer, Dordrecht The Editors of Encyclopedia Britannica. (2013, September 17). Asclepiadoideae. Retrieved from https://www.britannica.com/plant/Asclepiadoideae. The Editors of Encyclopedia Britannica. (2019, June 21). Brine shrimp. Retrieved from https://www.britannica.com/animal/brine-shrimp#ref3495. Urry, L. A., Cain, M. L., Wasserman, S. A., Minorsky, P. V., Orr, R. B., & Campbell, N. A. (2016). Campbell biology (11th ed.). New York, NY: Pearson.

7

EFFECTS OF PLANT DEFENSIVE COMPOUNDS ON HEART RATE Villarreal, H. (1990). Effect of temperature on oxygen consumption and heart rate of the australian crayfish Cherax tenuimanus (Smith). Comparative Biochemistry and Physiology Part A: Physiology, 95(1), 189–193. doi: 10.1016/0300-9629(90)90031-m

8...