Investigating Anaerobic Respiration in Yeast

Yeast is a tiny single-celled fungus. When there is no oxygen available, yeast can still release energy from glucose by anaerobic respiration. In yeast this is also called fermentation.

The word equation you must know is:

glucose → ethanol + carbon dioxide (+ a little energy)

Notice there are two products: ethanol (an alcohol) and carbon dioxide gas. The carbon dioxide is the key to the experiment — because it is a gas, we can see it and measure it. The faster the yeast respires, the faster carbon dioxide is produced.

So the whole investigation comes down to one idea: measure how fast carbon dioxide is given off, and that tells you how fast the yeast is respiring anaerobically. We then change one condition (for example the temperature or the sugar concentration) and see how the rate of CO₂ production changes.

Exam tip. Learn the word equation exactly. A very common slip is to write "carbon dioxide + water" (that is aerobic respiration). Anaerobic respiration in yeast makes ethanol + carbon dioxide, with no water and no oxygen used.

A reliable basic method looks like this:

1. Add a measured volume of glucose solution to a measured mass/volume of yeast (suspension) in a boiling tube or flask.
2. Add a thin layer of vegetable oil on top of the mixture. This floats and seals the surface so oxygen cannot get in — this keeps the conditions anaerobic.
3. Connect the tube with a delivery tube either to a tube of limewater, or to a gas syringe / inverted measuring cylinder of water to collect the gas.
4. Leave the apparatus at a set temperature (for example in a water bath) so the temperature is controlled.
5. Measure the carbon dioxide produced — count bubbles per minute, read the volume of gas collected, or time how long the limewater takes to turn cloudy/milky.
6. Repeat at each temperature (or each sugar concentration) and calculate a mean for reliability.

You can picture the apparatus like this:

The layer of oil is the part examiners love to ask about. It floats on top, stops air (oxygen) reaching the yeast, and so makes sure the yeast is respiring anaerobically rather than aerobically. Without it, oxygen could dissolve in and the yeast might respire aerobically instead.

Exam tip. If asked "why is oil added?" the answer is: to keep oxygen out / keep the conditions anaerobic. Do not say it is to keep the yeast warm — that is the job of the water bath.

Every good practical answer sorts the factors into three groups. You should be able to do this for this experiment.

Independent variable — the one thing you change. Usually either:

• the temperature of the water bath (e.g. 10 °C, 20 °C, 30 °C, 40 °C, 50 °C), or
• the concentration of sugar (glucose) solution (e.g. 1%, 2%, 5%, 10%).

Dependent variable — the thing you measure as a result. Here it is the rate of carbon dioxide production, for example bubbles per minute, cm³ of gas per minute, or the time for the limewater to turn cloudy (a longer time means a slower rate).

Control variables — everything you keep the same to make it a fair test:

• volume of yeast suspension and of glucose solution,
• the same type of sugar (e.g. glucose) and the same mass of yeast,
• the same pH,
• the same time for each reading,
• and the factor you are not investigating (if you change temperature, keep the sugar concentration the same; if you change sugar concentration, keep the temperature the same).

If you do not control these, you cannot tell whether a change in the CO₂ rate is due to your independent variable or due to something else — the experiment would not be a fair test.

Exam tip. A favourite question gives you a method and asks you to name the independent, dependent and two control variables. Practise writing these out quickly — they are easy marks if you have the structure ready.

You record your readings in a results table, then calculate the rate (for example, volume of gas ÷ time, or bubbles ÷ time) and plot a graph.

Drawing the graph correctly (easy marks):

• Put the independent variable (temperature or sugar concentration) on the x-axis.
• Put the rate of CO₂ production on the y-axis.
• Choose sensible scales, label both axes with units, plot points accurately, and draw a smooth line/curve of best fit.

What the graph usually shows:

If you changed temperature: the rate rises as temperature increases up to an optimum (around 30–40 °C for yeast), because the molecules and enzymes in the yeast work faster. Above the optimum the rate falls sharply because the high temperature denatures the enzymes (changes their shape) so they no longer work. This gives a graph that goes up to a peak and then down.

If you changed sugar concentration: the rate generally increases as more sugar is available, then levels off once there is plenty of sugar for the yeast to use (other factors become limiting).

Exam tip. If your temperature graph peaks and falls, explain it with enzymes: faster up to the optimum, then denatured above it. Do not say the yeast is "killed by the cold" — at low temperatures the yeast is just slow/inactive, not dead.

Top marks in a practical come from being able to evaluate — spot what could go wrong and suggest improvements.

Reliability (getting consistent results):

• Repeat each reading (e.g. three times) and calculate a mean — this reduces the effect of random error and identifies anomalous results.
• Use the same equipment and method each time.

Accuracy (getting close to the true value):

• Use a gas syringe to measure the exact volume of CO₂ rather than only counting bubbles (bubbles can be different sizes, so bubble-counting is less accurate).
• Use a water bath / thermostat to keep the temperature steady, and a thermometer to check it.
• Allow the apparatus to reach the set temperature before starting timing (an equilibration / warm-up time).

Common problems to mention:

• If the bung leaks, some CO₂ escapes and the rate looks too low.
• Bubbles of different sizes make bubble-counting unreliable.
• At the start the apparatus may not be at the right temperature, so early readings are off.

Improvements: measure gas volume with a syringe, use a water bath with a thermometer, repeat and average, and make sure all joints are airtight.

Exam tip. "Suggest one improvement" questions are common. Strong answers say why the change helps — e.g. "use a gas syringe so the exact volume of CO₂ is measured, which is more accurate than counting bubbles of different sizes."