How do microrespirometers measure cellular respiration in germinating seeds?

Microrespirometers measure cellular respiration by tracking oxygen consumption. As germinating seeds consume oxygen for energy, carbon dioxide is released and absorbed by potassium carbonate. This reduces the overall gaseous pressure inside the sealed chamber, causing manometer fluid to move up the capillary tube. The distance traveled indicates the rate of oxygen consumption and cellular respiration.

Why is potassium carbonate used in the microrespirometer setup?

Potassium carbonate absorbs the carbon dioxide released by germinating seeds during cellular respiration. By removing CO2 from the chamber, only oxygen consumption affects the gas pressure inside the microrespirometer. This allows the manometer fluid movement to accurately reflect the rate of oxygen uptake and respiration activity.

What is the purpose of using glass beads as a control in this experiment?

Glass beads serve as a control because they do not undergo cellular respiration. By comparing manometer fluid movement in beads versus germinating seeds, researchers can isolate the respiration signal from other physical or chemical changes. Any pressure changes in the bead control indicate experimental error rather than biological activity.

How does temperature affect the respiration rate of germinating seeds?

The experiment tests whether higher temperatures increase respiration rates in germinating seeds. Seeds placed in heated water baths are compared to those at room temperature. If respiration rate increases with temperature, the manometer fluid in heated microrespirometers will travel farther in the same time period than in room temperature controls.

What does the slope calculation represent in the respiration data analysis?

The slope represents the rate of cellular respiration over time. It is calculated by dividing the distance the manometer fluid traveled by the time elapsed. A steeper slope indicates faster oxygen consumption and higher respiration rates, allowing comparison between experimental conditions and controls.

Why must the microrespirometer chambers remain undisturbed after sealing?

Once sealed with manometer fluid, the microrespirometer creates a closed system where pressure changes directly reflect oxygen consumption. Disturbing the chamber—bumping the table or inserting objects—introduces physical pressure changes unrelated to respiration. This would produce inaccurate measurements and invalidate the experimental results.

How is the bar graph used to compare respiration rates across experimental groups?

The bar graph plots respiration rates calculated from slope values for each microrespirometer group: control room temperature, control heated, experimental room temperature, and experimental heated. Visual comparison of bar heights reveals differences in respiration between seeds and beads, and shows the effect of temperature on respiration rates.

Cellular Respiration

Concepts Instructor Prep Student Protocol

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Everything that is alive requires an energy source to fuel its activities. Ultimately, that source of energy is the sun. How then did the organisms on Earth harness this energy? It all starts with the photosynthesizers. These organisms are able to take carbon dioxide and water and then use energy captured from the sun as photons to force these molecules together, producing glucose and oxygen. The glucose is the key to the next critical step, one that nearly all organisms use in one form or another - cellular respiration. Cellular respiration is carried out by enzymes, either in the cell membrane in prokaryotes or in the mitochondria in eukaryotes.

The chemical reaction begins by breaking down the glucose using oxygen to create carbon dioxide and water again, but in the process, the energy that originally went into making the glucose molecule goes to two new places. One is into synthesizing ATP, or adenosine triphosphate molecules, an energy source that cells can readily use. The rest is lost as heat. So far, we have been talking about a kind of cellular respiration that uses oxygen, and this is referred to as aerobic respiration, but some organisms and cells are capable of doing respiration in the absence of oxygen. This is called anaerobic respiration. And instead of producing CO2 and water, this process makes ethanol as a byproduct. Fermentation is an example of this type of respiration. This is how yeast are able to produce alcohol in sealed containers. We can use a tool called a respirometer to measure aerobic respiration. Quite simply, this device measures the amount of oxygen used by an organism, germinating plant seeds in this case. Germinating seeds are respiring, which means we can expect them to be using up oxygen and releasing carbon dioxide. They don't have their green parts yet, so they aren't photosynthesizing.

Measuring respiration in a respirometer uses a clever method based on the ideal gas law, P times V equals n times R times T. P is the pressure of the system. V is the volume of the gas. n is the number of moles of gas present. R is the ideal gas constant…and T is the absolute temperature. Simply summarized, this means that you can figure out how many molecules of gas are present in a sample by measuring its volume, assuming you know the pressure and temperature, because R is a constant.

Respirometers contain potassium hydroxide, which traps carbon dioxide in solid form as potassium carbonate. So, due to cellular respiration, the oxygen in the sealed tube gets used up by the seeds, and they release carbon dioxide, which in turn gets trapped as potassium carbonate. Thus, as cellular respiration progresses, the total volume of gas inside the system decreases. We can quantify this by attaching the respirometer to a device called a manometer. Here, as the oxygen molecules are consumed, the pressure drops inside the respirometer chamber, and a colored liquid inside a tiny capillary tube is pulled in the direction of the decreased pressure. We can then estimate the quantity of gas remaining in the respirometer tube by reading off the value on the manometer level. This versatile system can be set up with many different variables, like different temperatures for example, and can be used to test the rate of respiration in many different types of living organisms.

In this lab, you will use a respirometer and manometer to measure the respiration rate of germinating seeds.

Print Procedure Steps

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Quantifying Respiration using Microrespirometers
- NOTE: In this experiment, you will measure the rate of cellular respiration for germinating seeds by measuring the rate of exchange for oxygen. As oxygen is consumed to provide energy, germinating seeds release carbon dioxide. This carbon dioxide is absorbed by potassium carbonate and thus the overall gaseous pressure of the respirometer will be reduced. HYPOTHESES: The experimental hypothesis is that germinating seeds will show a greater rate of respiration than control glass beads. Additionally, that at higher temperatures, the rate of cellular respiration in the seeds will increase. The null hypotheses are that both glass beads and germinating seeds will display a similar rate of cellular respiration and that temperature has no effect on respiration.
- To set up the control, first remove the plunger from a prepared microrespirometer and label it with a “CR” or “CH” respectively, depending on whether it is the control room temperature or heated treatment.
- Add the glass microbeads to each of the microrespirometers up to the 0.5 mL mark, and push the plunger in the syringe to the 1 mL mark.
- Place 3 - 4 washers onto the flared end of each microrespirometer to weigh them down while in the water bath. NOTE: The microrespirometer will sit capillary tube end up.
- Set the control devices to the side.
- To set up the experimental seated respirometers, remove the plunger from a prepared microrespirometer and label it with “ER” or “EH” respectively, depending on whether it is the experimental room temperature or heated treatment.
- Measure 0.5 mL of germinating seeds into the tuberculin syringe of the microrespirometer.
- Then, push the syringe to the 1 mL mark.
- Place 3 - 4 washers onto the flared end of each microrespirometer to weigh them down while in the water bath.
- To use the microrespirometers, carefully press the “CR” and “ER” microrespirometers into the room temperature water bath. The entire syringe should be submerged, but the capillary tubes should be entirely exposed. Remove water from the bath until this position is achieved. Be sure the top of the capillary tube is open.
- Perform the same procedure (step 9) for the “CH” and “EH” microrespirometers to set up the higher temperature experimental group.
- Wait 5 minutes before moving on to allow the temperature in the microrespirometers to equalize with the water bath.
- Add a single drop of manometer fluid to the top of each of the four capillary tubes which seals the microrespirometer chambers. NOTE: If the chambers are working properly, the manometer fluid will be sucked into the capillary chamber.
- Use the plunger set in the bath to suck the manometer fluid into the capillary. Stop when the fluid is about halfway down the capillary tube. IMPORTANT: The microrespirometers should not be disturbed once they are added to the bath. Do not bump the table or insert or remove anything to or from the water bath once the chambers have been sealed and equalized.
- Mark the bottom edge of the manometer fluid that is closer to the chamber. This represents time point zero.
- Set a timer to sound in 5 minutes.
- After 5 minutes, create a new mark where the manometer fluid is on each microrespirometer. Repeat every 5 minutes until 25 minutes has passed or until the manometer fluid has traveled the entire length of the capillary tube.
- Then, remove the respirometers from the water baths.
- Measure the distance between each of the marks and record them in Table 1 next to the correct time point. Click Here to download Table 1
- During clean up, manometers should either be deconstructed to be reused or disposed of in the trash.
Results
- To analyze the data you collected, first plot the data points with the x-axis as time and the y-axis as the distance the manometer moved, representing oxygen consumption.
- Calculate the slope of each line using this equation.
- Record these values for each microrespirometer in Table 2. NOTE: This value represents the amount of cellular respiration that took place over the course of the experiment. Click Here to download Table 2
- Plot the respiration rates as a bar graph.
- Examine both of the graphs. Consider how your results fit with the hypotheses. Note any observed differences in respiration rate between the control beads and the germinating seeds. Determine if increasing the temperature had any effect on the respiration rate.