What you need to know
- 1.14Core Practical 2: investigate vitamin C content of food and drink.
Practical study guide for a supervised school laboratory. The teacher must set reagent strengths, operating ranges and local risk controls before use. This is original preparation and evaluation material, not a replacement for the school’s approved laboratory protocol.
By the end, you should be able to…
- Explain the purpose of a vitamin C standard.
- Calculate concentration from endpoint volumes.
- Evaluate colour, extraction, oxidation and endpoint uncertainty.
Build your understanding
The measurement principle
Vitamin C reduces DCPIP, causing its blue colour to disappear under the chosen test conditions. Add standard or sample to a fixed volume of DCPIP and measure the volume needed to reach a consistent endpoint. More sample needed means a lower vitamin C concentration, if other reducing substances and matrix effects are negligible.
Use a known vitamin C solution to establish how much vitamin C reacts with that DCPIP aliquot. The standard and samples must use the same DCPIP batch, volume and concentration. Fresh preparation and consistent storage matter because vitamin C can be oxidised during handling.
Prepare a fair comparison
Use labelled tubes, measured syringes or pipettes, fresh standard, DCPIP and samples prepared consistently. Filter cloudy juice if necessary, recording any treatment. For a solid food, weigh a known mass, homogenise with measured extraction liquid, filter and make the extract up to a known final volume. Record dilution and extraction volumes.
If comparing heating or storage, divide one starting sample into matched portions and vary only the chosen factor. Keep sample mass or starting volume, extraction procedure, delay before testing and storage conditions consistent. Use independently prepared samples as well as repeat endpoint measurements where possible.
Find and repeat the endpoint
Place a measured DCPIP aliquot in a tube. Add the known standard in small measured amounts, mixing after each addition, and add more slowly near the endpoint. Record the total volume from the instrument readings. Repeat with fresh aliquots at least three times, then test prepared samples in the same way.
Use a white background and a consistent endpoint definition. Acidic samples can give pink colour and naturally coloured juices can obscure the endpoint; do not assume every mixture becomes visibly colourless. Check whether a sample blank, dilution or an alternative validated assay is needed rather than forcing an ambiguous endpoint.
Calculate concentration and content
Under the assay assumptions, Csample = Cstandard × Vstandard / Vsample. For a standard of 0.80 mg cm⁻³ requiring 1.5 cm³ and a sample requiring 4.0 cm³, Csample = 0.30 mg cm⁻³. If the sample was diluted threefold, the original concentration is 0.90 mg cm⁻³.
For an extract, estimated vitamin C mass = extract concentration × total extract volume. Divide by original food mass to obtain mg g⁻¹, or multiply that value by 100 for mg per 100 g. This assumes representative sampling and effective extraction. Do not confuse the volume used in one titration with the whole extract volume.
Safety and evaluation
Wear eye protection, avoid contact with reagents and never taste samples once brought into the laboratory. Use safe food-cutting and homogenising procedures under supervision, and follow local disposal instructions.
A large final drop can overshoot the endpoint. Repeat concordant measurements and report spread; investigate unusual values rather than deleting them automatically. Other reducing substances may react with DCPIP, and extraction or oxidation can bias the result. Describe the estimate as assay-dependent rather than exact nutrient composition.
NOT: a larger titre does not mean the drink is richer in vitamin C.
- Use a fresh standard because vitamin C oxidises
- The endpoint must be consistent
- Other reducing substances can bleach DCPIP
- Correct for any dilution of the food extract
Two traps this diagram blocks
- Do not say a bigger titre means more vitamin C.
- Do not ignore oxidation of the standard between repeats.
From the diagram alone
Question
Explain why a vitamin C standard is needed in a DCPIP titration of a fruit juice. [3]
Show a model answer
The standard of known concentration shows how much vitamin C decolourises the fixed volume of DCPIP. The juice titre can then be compared with that volume. Without the standard, the titre alone does not give a concentration.
The vitamin C titration lab
Calibrate a fixed DCPIP aliquot, compare fictional samples and defend an estimate. The numbers and illustrations are invented.
Simulation only. This is preparation and analysis, not a protocol for unsupervised work. A teacher sets real reagent strengths, apparatus settings and risk controls. Never taste laboratory samples.
Method 7 steps
- Make a fresh vitamin C standard of known concentration.
- Pipette a fixed volume of DCPIP solution into a conical flask standing on a white tile.
- Fill a burette with the standard and record the initial reading at the bottom of the meniscus.
- Add the standard to the DCPIP, swirling, until the blue colour just disappears. Record the final reading: this is the rough titre.
- Repeat with fresh DCPIP, adding drop by drop near the endpoint, until you have concordant titres.
- Repeat the titration with each juice (diluted if needed), using the same volume and concentration of DCPIP.
- Use the mean concordant titres to calculate the vitamin C concentration of each juice, correcting for any dilution.
Aligned with the order of the specification practical. Simulation only: a teacher sets the real volumes, concentrations and risk controls; this is not a laboratory protocol.
Variables
- Independent
- The juice or sample tested, and so its vitamin C concentration.
- Dependent
- Volume of juice needed to decolourise the DCPIP (the titre).
- Controlled
- Volume and concentration of DCPIP
- The same endpoint definition: first permanent loss of blue
- Storage time, light and temperature of the juices (vitamin C oxidises)
- Swirling and rate of addition near the endpoint
Safety
- Wear eye protection. DCPIP and acidic juices can irritate eyes and skin; wash off splashes.
- Clamp the burette securely. Fill it below eye level with a funnel, then remove the funnel before titrating.
- Never taste juices or food in a laboratory. Clear up broken glass as your teacher directs.
Follow your school’s risk assessment; these notes are not a substitute for it.
Interpreting
- A smaller titre means more vitamin C: less juice was needed to decolourise the same DCPIP.
- Calculate C(sample) = C(standard) × V(standard) ÷ V(sample) from mean concordant titres, then multiply by any dilution factor.
- Coloured juices can hide the endpoint, and other reducing substances also decolourise DCPIP (overestimate). Oxidation during storage lowers the result (underestimate).
- Report the spread of repeats and the assumptions. One assay does not show a health benefit.
Guided method · step 1 of 10
- In 1 · Set up a fair comparison, just below these steps, press Prepare fresh fictional vitamin C standard.
- The standard has a known concentration of 0.80 mg cm⁻³.
Vitamin C (ascorbic acid) is oxidised by air, light and warmth, so only a freshly made standard has a reliable known concentration. Titrating it first calibrates how much vitamin C your DCPIP aliquot needs.
1 · Set up a fair comparison
Vitamin C reduces blue DCPIP. Keep the DCPIP aliquot and concentration matched for the standard and every sample. The first run for each is rough; later runs are repeats.
Known standard: 0.80 mg cm⁻³. The reference uses a fictional 2 cm³ DCPIP aliquot. A sample titre alone cannot give its concentration.
A naturally coloured or acidic sample may leave a pink background when blue DCPIP disappears. Use the same endpoint definition and consider a sample blank or another validated assay.
Burette tap
Play opens the tap. Normal flow is about 1 cm³ s⁻¹; slow is about 0.2 cm³ s⁻¹. Step adds 0.1 cm³.
Toolbox
Prepare the fresh standard, choose a solution, then start a run.
3 · Select repeats and calculate
Keep every result visible. Select at least two concordant repeat titres for the standard and your chosen sample. Investigate omitted readings rather than silently deleting them.
Record a standard run to begin the results table.
Evaluate the claim
Repeat titrations test endpoint variability; independently prepared extracts test preparation variability. Coloured samples may obscure the endpoint, vitamin C can oxidise in storage, and a sample blank or alternative validated assay may be needed.
Make your reasoning visible
Check the direction of the ratio
With fixed DCPIP, a smaller sample volume indicates more concentrated vitamin C. The standard volume goes above the sample volume in the ratio.
Carry units through
mg cm⁻³ × cm³ gives mg. Dividing by grams of food gives mg g⁻¹.
Separate repeats from fresh preparations
Repeating an endpoint on one extract checks assay variability; repeating extraction checks additional preparation variability.
Key explanations
- The volume added to reach the endpoint is the titre.
- Extracts may need a dilution correction before the concentration in the original food is stated.
Avoid this misconception
The claim: A darker juice must contain more vitamin C.
The correction: Natural pigment intensity does not measure vitamin C and can obscure the DCPIP endpoint.
Check your understanding
Try each question before opening the suggested answer.
Practice question
Retrieval and explanation · suggested answer
A 1.0 mg cm⁻³ standard takes 1.2 cm³ and an unknown takes 3.0 cm³ to reach the same endpoint. Calculate concentration before any dilution correction.
Show suggested answer
1.0 × 1.2/3.0 = 0.40 mg cm⁻³.
Practice question
Retrieval and explanation · suggested answer
An extract contains 0.20 mg cm⁻³ in 50 cm³ made from 5.0 g food. Estimate mg per 100 g.
Show suggested answer
Total = 0.20 × 50 = 10 mg. Per gram = 10/5.0 = 2 mg g⁻¹. Per 100 g = 200 mg, assuming complete representative extraction.
Practice question
Retrieval and explanation · suggested answer
Why test heated and unheated samples after the same storage delay?
Show suggested answer
Vitamin C can change during storage, so unequal delays could confound the effect attributed to heating.
Take the idea further
Apply the same specification knowledge to an unfamiliar problem. Any extra assumptions are supplied in the question.
Stretch question
A heated sample needs more volume to decolourise DCPIP, but it also lost water by evaporation. Why should the original volume be restored before comparing concentration?
Show a hint
Heating may affect both vitamin C amount and solvent volume.
Explore the reasoning
Evaporation concentrates the remaining solutes and can mask a fall in vitamin C amount. Restoring to a known matched volume helps separate the change in vitamin C from concentration caused by water loss.
Recall this lesson
Lesson recall questions
Question 1 of 5: Why can vitamin C decolourise DCPIP?
Vitamin C in food and drink
Practice question
Retrieval and explanation · suggested answer
Why can vitamin C decolourise DCPIP?
Lesson: CP2: Vitamin C Content of Food and Drink
Try this from memory first.
Beyond the classroomTesting a nutrient claimCareers · food analysis · Enrichment, not required recall
Practical Biology describes a DCPIP assay using a known vitamin C standard. It illustrates how a chemical response can be used to compare foods, while sample colour and other reducers complicate interpretation.
Back to the spec: Specification 1.14: calibrate a response and evaluate the assumptions behind a food measurement.
Practical Biology: vitamin C assayQuestion
Why should a lab avoid reporting an uncertain visual endpoint to six decimal places?
Explore a possible response
The displayed precision would exceed the measurement’s resolution and uncertainty. Report a justified number of significant figures with relevant limitations.