Compare the time for amylase to digest starch at different pH values. Use prepared enzyme, starch, buffers, iodine, a spotting tile, pipettes, stopwatch and thermometer.Labelled apparatus schematic; not to scale. Follow the measurements and connections, not the drawn dimensions.
The independent variable is pH, maintained by buffers. The dependent measurement is the time until starch is no longer detected; for equal amounts of starch, use 1/time to compare the relative rates (higher 1/time means faster reaction).
Control temperature, reagent concentrations and volumes, total volume, sampling interval and mixing procedure. Changing acid volume alone can also change dilution.
Pearson includes a beaker water bath heated with a Bunsen burner to maintain temperature. Monitor it; a stable initial temperature is insufficient if the bath cools later.
Use eye protection, avoid enzyme/iodine contact and handle hot water and glass safely. Prepared amylase avoids saliva hygiene problems.
Continuous sampling method
Put iodine drops in separate tile wells. Measure equal reagent volumes using clean pipettes and label each tube with buffer pH.
Allow the starch, enzyme and buffer to reach the water-bath temperature before mixing. Keep enzyme and starch separate during this waiting time so digestion does not begin before timing starts.
Mix the reactants and start timing promptly, keeping the mixture at the controlled temperature.
At fixed intervals, such as 10 seconds, place a small sample into a fresh iodine well. Iodine belongs on the tile, not in the main reacting mixture.
Blue-black indicates starch remains. The first sample that stays orange-brown indicates no detectable starch; record the elapsed time.
Repeat with independent fresh mixtures at each pH. A no-enzyme control should retain starch under otherwise matching conditions.
Results, precision and evaluation
Put pH on the horizontal axis and mean time or calculated relative rate on the vertical axis, with units. The fastest reaction has the shortest time and highest 1/time. Many amylase preparations work fastest near neutral pH, but use your results to decide.
If starch is present at 50 s and absent at 60 s, the endpoint lies within that interval; it is not known to 0.001 s. Shorter sampling intervals improve endpoint resolution.
Use a preliminary (pilot) test to avoid reactions too fast to time. Keep iodine/sample volumes and colour judgement consistent; a validated colorimeter method can reduce subjective judgement.
Extreme pH can disrupt bonding and active-site shape, causing denaturation. Substrate binding becomes less effective away from the optimum.
If no endpoint is reached, record a time limit such as greater than 180 s rather than zero. The iodine test detects starch, not the amount or identity of sugar formed.
Cross-contamination of starch stock with amylase starts digestion early. Use separate clean pipettes. A consistently late clock start underestimates time and overestimates 1/time.
Calculate a mean and compare repeat spread. Repeated readings from one well do not substitute for independently mixed reactions.
Exam skills: planning, precision and evaluation
State what you change (the independent variable), what you measure (the dependent variable) and what you keep the same (control variables). Explain how you keep each control variable constant, rather than just saying “make it fair”.
Accuracy means how close a result is to the true value. Precision means how close repeated measurements are to each other. Resolution is the smallest change an instrument can show. More digits on a display do not automatically mean a more accurate result.
Repeat measurements for each condition, calculate a mean and describe how spread out the results are. This helps assess and reduce the effect of random errors. Repeating cannot fix an error that pushes results consistently in one direction (a systematic error), such as a stopwatch consistently started after mixing.
Repeatability means getting similar results when the same person repeats the same method with the same equipment. Reproducibility means getting similar results when someone else, or different suitable equipment, repeats the experiment. Results can be consistent but still inaccurate.
Check that instruments read zero correctly and are calibrated where needed. Read scales at eye level: looking from an angle can give a wrong reading (parallax error). Choose suitable ranges, measurement intervals and scale divisions (resolution).
Write down the original readings straight away in a table, with units in the headings. Use decimal places that match the instrument’s resolution. Keep the original data and round only when needed. Do not discard a result just because it differs from your prediction.
An anomalous result does not fit the pattern of the other results. Repeat that measurement and check the method. Only leave it out of a mean if you have a clear reason; state which result you excluded and why.
For continuous variables, plot the independent variable on the horizontal axis and the dependent variable vertically. Use sensible scales, units and a best-fit line or curve; do not automatically join every point or force the graph through zero.
Find the gradient of a straight best-fit line using a large triangle: vertical change ÷ horizontal change. For a curve, draw a tangent to estimate the gradient at one point. Explain what the gradient shows in this experiment, include its units and use measured values to support your conclusion.
Uncertainty describes the possible range around a measurement. For one reading on a scale, half the smallest division is a useful classroom estimate unless the question says otherwise. If you subtract two readings, both have uncertainty. Percentage uncertainty = absolute uncertainty ÷ measured value × 100. Follow the method specified in the question.
Use results as evidence and then explain what they mean. A pattern linking variables (a correlation) does not prove that one causes the other. If the ranges of repeat results overlap, a claimed difference may be less convincing. Keep conclusions within the range tested and suggest an improvement that tackles a specific error.
Mr Kidman's Classroom