Required Practical 1: Enzyme Rates

0:24A reaction we can measure

First, a reaction we can measure. Catalase breaks down hydrogen peroxide into water and oxygen. Oxygen is a gas, so we can collect it in a gas syringe and read off the volume. The enzyme comes from an extract of something living, such as liver or potato.

More oxygen in less time means a faster rate. That is how we turn an invisible reaction into numbers.

0:49Independent, dependent, control

Now the three kinds of variable. The independent variable is the one you change. Here, that is temperature. The dependent variable is the one you measure. Here, that is the volume of oxygen collected. Everything else is a control variable, and it must stay the same.

If two things change at once, you can never say which one caused the result.

1:12Water baths and temperature range

To change temperature, use water baths. Set up a range of at least five temperatures, say ten, twenty, thirty, forty and fifty degrees, and check each one with a thermometer. Put the enzyme and the hydrogen peroxide in separate tubes in the same water bath, and leave them there for a few minutes. That way both reach the right temperature before you mix them.

1:37Controls and the buffer

Now the controls, where students lose easy marks. Use the same volume and concentration of enzyme, and the same volume and concentration of hydrogen peroxide, every time. Control the pH with a buffer solution, because a change in pH changes the shape of the active site. Don't just name a control variable. Say how you control it.

1:59Measuring oxygen over time

Mix the two, start the stopwatch, and record the volume of oxygen at regular time intervals. When you plot volume against time, the curve starts steep, then flattens. It flattens because the substrate is being used up, so the rate falls. Eventually the line goes flat, because all the hydrogen peroxide has been broken down.

2:21Why initial rate (tangent)

So we do not use the average rate over the whole reaction. We use the initial rate. At the start, substrate concentration is highest and nothing else has changed, so it is a fair comparison between temperatures. To find it, draw a tangent to the curve at time zero. Then work out the gradient: the change in volume, divided by the change in time. The unit is cubic centimetres per second.

A steeper tangent means a faster initial rate. Do this for every temperature, and you can compare them fairly.

2:55Repeats and the mean

Repeat at each temperature, and calculate a mean. That makes your results more reliable. Finally, plot initial rate against temperature. You should see the rate rise to a peak, then fall as the enzyme denatures. You can also state the uncertainty in each measurement, for example from the smallest division on the syringe.

3:51Examiner trap and recap

The trap. Writing, keep everything else the same, earns nothing. Name the variable and say how. Recap. Change one thing, measure oxygen over time, control the rest, find the initial rate with a tangent, and repeat.

Next lesson, we move on to nucleic acids, and what a nucleotide is made of.