Question 1 — Estimating hydrogen peroxide concentration by iodine-clock timing
Serially dilute 2.0% hydrogen peroxide by half four times, then time how long each dilution takes to turn blue-black with the iodine–starch–thiosulfate mixture. Use the same timing method on an unknown 'patient sample' U to estimate its hydrogen peroxide concentration, then interpret Cambridge-supplied data on bacterial hydrogen peroxide production at two temperatures.
RUN IT ON THE BENCH
Your readings feed straight into the answer table below as you record them.
ON YOUR BENCH — FROM THE CONFIDENTIAL INSTRUCTIONS
Question 1 — solutions
- solution R1(100 cm³)
- solution R2(10 cm³)
- solution R3(10 cm³)
- solution R4(10 cm³)
- solution H(25 cm³)
- solution U(10 cm³)
- solution W(100 cm³)
Question 1 — apparatus
- 10 cm³ syringes(2)
- 1 cm³ syringes(5)
- beakers(5)
- test-tubes, large(8)
- test-tube rack(1)
- glass rod(1)
- container labelled 'For washing'(1)
- container labelled 'For waste'(1)
- paper towels(8)
- glass marker pen, permanent(1)
- stop-clock or timer showing seconds(1)
- suitable eye protection(1)
STEP 1 — SERIALLY DILUTE STOCK H (2.0% H₂O₂) BY HALF, FOUR TIMES
Each step transfers 10 cm³ of the previous beaker's solution into the next and tops it up with 10 cm³ of distilled water W — exactly halving the concentration every time.
STEP 2 — MIX WITH R1–R4 AND TIME THE BLUE-BLACK END-POINT
1 cm³ of 2.0% H₂O₂ + R1 (acid) + R2 (starch) + R3 (iodide) + R4 (thiosulfate). Start the clock the moment they mix.
STEP 3 — UNKNOWN SAMPLE U — SAME METHOD, THREE REPEATS
1 cm³ of the patient sample U + R1 + R2 + R3 + R4, timed the same way. Repeat three times.
Why does the tube suddenly go blue-black instead of gradually darkening?
H₂O₂ slowly oxidises iodide to iodine, but the fixed, small amount of thiosulfate mops up that iodine as fast as it forms — reducing it straight back to iodide. Nothing is visible while thiosulfate remains. The instant it runs out, iodine starts to accumulate for the first time, and starch turns blue-black almost immediately. That abrupt switch is what makes it a "clock" reaction — you're timing when a fixed resource runs out, not watching a gradual colour change.
Why does halving the concentration roughly double the time?
The same fixed amount of thiosulfate has to be used up either way. At half the H₂O₂ concentration, iodide is oxidised at roughly half the rate, so it takes roughly twice as long to consume the same amount of thiosulfate. That inverse relationship is what lets you use time as a stand-in for concentration — and why the calibration curve is built from time vs. concentration, not time vs. time.
Why time sample U three times instead of once?
A single timing could be an anomaly — a late reagent addition, a missed start, a slightly dirty tube. Three repeats let you check for consistency, and averaging them reduces the effect of random error on the final estimate. It's the same logic as concordant titres in a titration.
YOUR TASKS · 0/4
- ✓Serially dilute H down to 0.125%
- ✓Time the blue-black end-point at all five concentrations
- ✓Time sample U three times (U1, U2, U3)
- ✓Calculate the mean time for sample U
TABLE 1(A)(II) — CALIBRATION SERIES
| 2.0% | 1.0% | 0.5% | 0.25% | 0.125% |
|---|---|---|---|---|
| — | — | — | — | — |
Time for blue-black colour to appear / s.
TABLE 1(A)(III) — SAMPLE U REPEATS
| U1 | U2 | U3 |
|---|---|---|
| — | — | — |
Time for blue-black colour to appear / s.
YOUR ANSWERS
Complete Fig. 1.1 to show how you will prepare your serial dilution of the 2.0% hydrogen peroxide solution, H, by half at each step. Each beaker needs a labelled arrow for the volume of H2O2 transferred, a labelled arrow for the volume of distilled water W added, and a label underneath for the resulting concentration.
Record, in an appropriate table, the time taken for a blue-black colour to appear for each of the five hydrogen peroxide concentrations (2.0%, 1%, 0.5%, 0.25%, 0.125%). If the colour hasn't appeared after 180 seconds, stop timing and record 'more than 180'.
| 2.0% | 1.0% | 0.5% | 0.25% | 0.125% | |
|---|---|---|---|---|---|
| time for blue-black colour to appear / s |
Record the time taken for a blue-black colour to appear for U1, U2 and U3 (three repeats of sample U).
| U1 | U2 | U3 | |
|---|---|---|---|
| time for blue-black colour to appear / s |
Calculate the mean time taken for the blue-black colour to appear for sample U. Show your working.
Use your results in (a)(ii) and (a)(iv) to estimate the concentration of hydrogen peroxide in sample U.
Explain why repeating the measurement for sample U allows you to have more confidence in your estimate.
With reference to your estimate for sample U, describe one other modification to the procedure that would allow a more accurate estimate of the concentration of hydrogen peroxide in sample U.
Scientists investigated the effect of temperature on hydrogen peroxide production by Streptococcus pyogenes at 20°C and 37°C over 168 hours (Table 1.2). Plot a graph of this data on the grid in Fig. 1.2.
State two conclusions from the results of the investigation at the two temperatures.
A sample taken at 20°C showed 14.5% of the bacteria were able to produce hydrogen peroxide. Use your graph in Fig. 1.2 to estimate when the sample was taken. Show on your graph how you obtained your estimate, and give your answer to the nearest hour.
State one variable that the scientists would need to keep constant so that the results at the two temperatures could be compared.