C. Metabolic Processes · Metabolic Processes

Investigation: Detecting the Products of Cellular Respiration

C2.2 C2.1 C3.1 A1.1 A1.4 A1.5 A1.6 A1.8 A1.10 A1.11About 75 minutes

Learning goals

I can plan a controlled investigation to identify the products of cellular respiration in germinating seeds or yeast.
I can use chemical indicators and temperature measurements to detect CO₂, heat and O₂ use.
I can record qualitative observations clearly and display them in an appropriate format.
I can interpret observations, identify sources of error, and connect results to the respiration equation.

Words to know

germinating seed
A seed that has absorbed water and started to grow; it respires rapidly, using stored starch or oil as fuel.

bromothymol blue (BTB)
An acid-base indicator that is blue in basic solution, green near neutral and yellow in acidic solution; it detects CO₂ dissolved in water.

limewater
A solution of calcium hydroxide that turns milky (cloudy) when CO₂ bubbles through it, because insoluble calcium carbonate forms.

respirometer
A device that measures the rate of respiration, usually by measuring how much O₂ is used in a sealed chamber.

qualitative observation
An observation described in words (colour, cloudiness, smell) rather than measured with numbers.

quantitative observation
An observation recorded as a measurement with a number and a unit (e.g. 26.0 °C).

negative control
A set-up in which no effect is expected (e.g. boiled seeds or glass beads), used to show that any change is caused by the variable being tested.

Seeds that breathe

A dry pea in a jar seems lifeless. Soak it overnight, though, and it wakes up: enzymes digest its stored starch, and its cells begin to respire rapidly to power growth. If cellular respiration really is C₆H₁₂O₆ + 6O₂ → 6CO₂ + 6H₂O + energy, then germinating seeds should use O₂, release CO₂ and release heat. C2.2 asks you to test this in a laboratory investigation, record qualitative observations, and display them clearly. In this lesson you'll learn how such an experiment is designed and analysed, then carry out a safe version at home.

Remember from Lessons 4 to 8

From Lessons 4 to 8: aerobic respiration releases CO₂ in pyruvate oxidation and the Krebs cycle, makes water at complex IV, uses O₂ as the final electron acceptor, and releases about two-thirds of glucose's energy as heat. Yeast without O₂ ferments, producing ethanol and CO₂.

Designing the investigation

Question: Do germinating seeds carry out cellular respiration, and what are its products?

Hypothesis: If germinating peas carry out cellular respiration, then a sealed tube containing them will show a rise in CO₂ (indicator changes colour), a rise in temperature, and a decrease in O₂, compared with tubes of boiled peas or glass beads, because respiration oxidizes glucose to CO₂ and releases energy as heat.

Variables:

Independent: whether the tube contains living (germinating) peas, dead (boiled) peas or glass beads.
Dependent: indicator colour (CO₂); temperature (heat); O₂ use (in a respirometer).
Controlled: number and volume of peas/beads, volume of indicator, tube size, temperature of the room, time.

Why two controls? Glass beads show what happens with no living material (a check on the indicator and set-up). Boiled peas show what happens with the same material but dead cells: their enzymes are denatured, so any change must be caused by microbes or by non-biological effects, not by the peas' respiration.

A typical school set-up. A wire gauze keeps the peas out of the indicator; each tube is stoppered. Tube A holds germinating peas; tubes B (boiled peas) and C (glass beads) are negative controls.

Detecting the products

CO₂. CO₂ dissolves in water to form carbonic acid (H₂CO₃), lowering the pH. Two common tests:

Bromothymol blue (BTB): blue → green → yellow as CO₂ increases.
Limewater: clear → milky as CO₂ reacts to form solid calcium carbonate: Ca(OH)₂ + CO₂ → CaCO₃ + H₂O.

Heat. Place germinating and boiled seeds in two vacuum flasks (thermoses) with thermometers, plugged with cotton wool (which lets some air in). Read the temperature every few hours.

O₂ use. In a respirometer, a chemical such as potassium hydroxide absorbs the CO₂ produced, so the gas volume in the sealed tube falls only because O₂ is being used. A drop of coloured liquid in an attached capillary tube moves toward the seeds. (Potassium hydroxide is corrosive; it is used only in a school lab with goggles and gloves.)

Water. Water is a product too, but it is hard to detect because the seeds are already moist.

Bromothymol blue changes from blue to green to yellow as dissolved CO₂ makes the solution more acidic. (Here the colours are the point of the diagram.)

Lab safety (A1.4)

Wear safety goggles; BTB, limewater and potassium hydroxide can irritate or burn eyes and skin. Know the WHMIS symbols on each bottle.
Boil the control seeds with care (hot plate, tongs) and let them cool to room temperature before use, so they don't add heat or warm the indicator.
Rinse seeds with a dilute disinfectant solution provided by the teacher (to reduce mould and bacteria), then with water.
Wash your hands after handling seeds and indicators. Dispose of solutions as your teacher directs.

Recording and displaying observations

C2.2 asks you to interpret qualitative observations and display them in an appropriate format. A good qualitative data table has:

a clear title,
one row per set-up (A, B, C) and columns for each time,
precise descriptive words ("yellow-green", "slightly cloudy") rather than "changed",
a column for notes (mould, condensation, smell).

For quantitative data (temperature, distance moved by a respirometer drop), use a line graph with time on the x-axis.

Sample temperature data from two thermoses (germinating vs boiled peas). The germinating peas warm up as respiration releases heat; the boiled peas stay at room temperature.

Worked example

Let's interpret a set of results

A student's BTB results after 30 minutes: Tube A (germinating peas): yellow. Tube B (boiled peas): blue. Tube C (glass beads): blue. Temperature after 48 h: germinating 28.0 °C, boiled 21.0 °C, room 21.0 °C. Interpret the results and write a conclusion.

Tube A turned yellow: the indicator became acidic, so CO₂ was released by the germinating peas.
Tubes B and C stayed blue: no CO₂ from dead peas or beads. This shows the change in A was caused by living pea cells, not by the indicator or the tube.
Temperature: germinating peas warmed by 7.0 °C; boiled peas didn't. Living peas released heat.
Link to theory: respiration oxidizes glucose (from stored starch), releasing CO₂ (pyruvate oxidation and Krebs) and energy, about two-thirds of it as heat.
Conclusion: the evidence supports the hypothesis that germinating peas carry out cellular respiration, producing CO₂ and heat.

Germinating peas released CO₂ (BTB turned yellow) and heat (+7.0 °C), while the controls showed no change, so the changes were caused by cellular respiration in living cells.

Worked example

Let's find sources of error

In a repeat, Tube B (boiled peas) turned green after 24 hours, and the boiled-pea thermos warmed by 2 °C after 3 days. Suggest an explanation and how to improve the procedure.

The boiled peas are dead, but bacteria and moulds growing on them are alive and respire, producing CO₂ and heat.
This is more likely over long times (24 h, 3 days) and if the seeds weren't disinfected.
Improvements: rinse all seeds in a dilute disinfectant before the experiment, shorten the time, use fresh solutions, and repeat each set-up several times to check reliability.

Microbial respiration on the dead seeds is the likely cause. Disinfecting the seeds and shortening the time would reduce this error.

Just for fun

Germinating seeds can produce enough heat in a large, damp pile to raise its temperature noticeably. That's one reason grain must be dried well before it goes into a storage bin: a soggy bin of seeds is basically a slow-cooker that runs on cellular respiration.

Hands-on activity

Home investigation: Do germinating seeds respire?

You need: about 2 cups of dried peas, chickpeas or mung beans, 2 clean wide-mouth jars with lids, 2 small cups that fit inside the jars, red cabbage indicator (from Lesson 5: cabbage soaked in hot tap water, strained), 2 small insulated mugs or thermoses with lids, a kitchen thermometer, cotton balls, a pot (for boiling).

Safety note: boil the control seeds carefully (oven mitts, pot handle turned inward, never leave the stove unattended) and let them cool completely before use. Wash hands after handling seeds.

Day 0: soak half the seeds in water overnight (these will germinate). Boil the other half for 10 minutes, then cool them completely (dead control).

Part 1: CO₂ (2 to 3 days).

Rinse the soaked seeds. Put a 2 cm layer of moist seeds in jar A; the cooled boiled seeds in jar B.
Put a small cup with 2 cm of cabbage indicator in each jar, resting on top of the seeds, then close the lids.
Compare the indicator colours (against white paper) at the start, after 24 h and after 48 h. A shift toward pink or red-violet means CO₂ has dissolved.

Part 2: heat (2 days).

Fill one insulated mug with damp germinating seeds and another with damp boiled seeds. Push the thermometer into the middle of each (or read each in turn) and loosely plug the top with cotton.
Record the temperature twice a day for 2 days.

Display: make a qualitative observation table for Part 1 and a line graph for Part 2. Write a conclusion and two sources of error.

What to look for: the indicator over the germinating seeds should shift slightly toward pink or violet compared with the control (colour changes can be subtle; photos help), and the germinating seeds should become a few degrees warmer than the boiled seeds. Possible errors: mould on the boiled seeds, lids not sealed, temperature of the room changing, different amounts of seeds.

Your turn

State a testable hypothesis for an investigation of respiration in germinating seeds.

Identify the independent and dependent variables in the three-tube BTB experiment.

Why are boiled peas used as a control?

How does limewater show the presence of CO₂? Write the equation.

Why is potassium hydroxide placed in a respirometer?

Give two features of a good qualitative data table.

Why is water, a product of respiration, not usually tested for in this experiment?

More practice: warm-up

What colour is BTB in the presence of a lot of CO₂?

What gas makes limewater go milky?

Name the three products of aerobic respiration you could test for.

What is a qualitative observation?

Why soak seeds before the experiment?

More practice: on your own

Predict how the BTB in Tube A would change if the tube were kept at 5 °C instead of 25 °C. Explain.

A student uses yeast and sugar water instead of peas, sealed with no air. Which products would you expect, and how would you show that one of them is ethanol without tasting it?

In a respirometer, the coloured drop moves 18 mm toward germinating peas in 10 min. With glass beads it moves 2 mm. What is the corrected rate? Suggest why the bead tube moved at all.

Explain why the germinating-pea thermos is plugged with cotton wool, not sealed airtight.

A classmate concludes, "Germinating seeds produce CO₂, so they are not photosynthesizing." Evaluate this conclusion.

Challenge

Design a respirometer experiment to test whether germinating peas respire faster at 30 °C than at 20 °C. Include the controls, variables and how you'd process the data.

A tube contains germinating peas and KOH. Another contains germinating peas without KOH. The drop moves in the first but hardly moves in the second. Use the respiration equation to explain both results.

Peas store mostly starch; sunflower seeds store mostly oil (fat). Predict, with reasons, which would use more O₂ per gram of stored fuel respired, and how this would show in the respirometer (with KOH present).

Check your understanding

Quiz

In a respiration experiment, why is a tube of boiled peas included?

Bromothymol blue in a tube with germinating peas turns yellow. This shows that the peas:

In a respirometer, potassium hydroxide is used to:

Which is a qualitative observation?

A thermos of boiled peas warms slightly after three days. The most likely explanation is:

Why should a thermos of germinating seeds be plugged with cotton rather than sealed?

For parents and teachers

Big idea: This lesson covers the C2.2 investigation: detecting CO₂ (BTB, limewater), heat and O₂ use from germinating seeds, with proper controls, and recording and displaying qualitative observations (A1.1, A1.4 to A1.6, A1.8, A1.10, A1.11).

How to help: The home investigation takes 2 to 3 days. Check that your teen boils the control seeds safely (oven mitts, stove never left unattended). Ask your teen to show you the data table and graph and explain what each control tells them. Indicator chemicals such as BTB, limewater and potassium hydroxide are for the school lab only; the cabbage indicator is the safe home alternative.

Common difficulties:

Forgetting the purpose of each control (beads: the set-up; boiled seeds: living vs dead).
Vague observations ("it changed") instead of precise descriptions.
Not realizing that microbes on dead seeds can respire too.
Thinking that a respirometer without a CO₂ absorber measures O₂ use.