Vitamin C comparisons and endpoint calibration

What you’ll learn

• Use a reference solution to interpret a vitamin C endpoint.
• Explain dilution, oxidation and sample-colour limitations.

Understand the idea

Core practical 2 compares vitamin C in food or drink. In an appropriate DCPIP assay, vitamin C reduces the coloured indicator. When a fixed amount of indicator is used with a consistently defined endpoint, the vitamin C amount needed is the same for the reference and unknown. A more concentrated sample therefore needs a smaller sample volume to reach that endpoint. The inverse relationship is easy to reverse by mistake.

A known reference concentration links sample volume to vitamin C amount. Apply any extraction or dilution factor before comparing the original food or drink. Fair comparisons require a clearly stated basis, such as milligrams per cubic centimetre of drink or milligrams per gram of starting food, rather than comparing unmatched volumes of extracts. Repeated endpoints reveal variation, but do not correct an incorrect reference concentration.

Vitamin C can be oxidised during storage and handling, so elapsed time and exposure conditions can alter the value being measured. An extract's own colour may obscure the endpoint, and other reducing substances may affect assay specificity. Use appropriate blanks, matched handling and calibration to evaluate these limitations. An assay result alone does not establish the health benefit of a particular drink.

Key formulae

$$c_{\mathrm{reference}}V_{\mathrm{reference}}=c_{\mathrm{unknown}}V_{\mathrm{unknown}}$$

$$c_{\mathrm{original}}=c_{\mathrm{assayed}}\times\mathrm{dilution\ factor}$$

Worked example

A reference containing $0.20\ \mathrm{mg\,cm^{-3}}$ of vitamin C needs $1.50\ \mathrm{cm^3}$ to reach the defined endpoint. A fivefold-diluted drink needs $3.00\ \mathrm{cm^3}$ for the same indicator amount. Assume the response is due to vitamin C alone. Estimate the original drink's concentration.

1. The reference supplies $0.20\times1.50=0.30\ \mathrm{mg}$ of vitamin C at the endpoint.
2. The diluted drink supplies the same amount in $3.00\ \mathrm{cm^3}$, so its concentration is $0.30/3.00=0.10\ \mathrm{mg\,cm^{-3}}$.
3. Reverse the fivefold dilution: $0.10\times5=0.50\ \mathrm{mg\,cm^{-3}}$.

Answer: Approximately $0.50\ \mathrm{mg\,cm^{-3}}$.