12.2.13—Redox indicator investigation

DCPIP and methylene blue are redox indicators that can accept hydrogen and change from blue to colourless when reduced. In a yeast suspension, the speed of this colour change is a proxy for respiration rate: faster discolouration indicates faster hydrogen release, not a direct measurement of ATP production.

• Respiratory dehydrogenation removes hydrogen from substrate molecules.
• The indicator accepts hydrogen and becomes reduced, producing the visible blue-to-colourless change.
• A faster respiration rate gives faster hydrogen release and faster indicator reduction.
• For a fixed colour endpoint, rate is inferred from the inverse relationship: a shorter time means a faster rate. Use the same endpoint and time basis for every condition.
• The SME investigation uses living yeast and can compare one independent variable at a time, such as temperature or substrate concentration.

1. Prepare matched yeast suspensions and choose either a controlled temperature range or a range of concentrations of the same substrate.
2. Keep yeast volume and concentration, substrate type, dye identity and volume, dye concentration, temperature of the added dye and other relevant conditions constant.
3. Add the indicator consistently, start timing at the same point, and record the time for the same defined colour endpoint.
4. Repeat each condition, use a control containing the yeast suspension without dye when the suspension has its own colour, and calculate a representative rate from the repeated readings.
5. Compare or plot rate against the independent variable. Interpret the colour change as an indirect proxy and avoid treating endpoint time as a direct ATP or oxygen measurement.

A fair test changes one independent variable at a time. Do not compare different dye volumes, yeast amounts or colour endpoints as if they were equivalent, and do not assign an unsupported optimum temperature or numerical rate. This indicator method is distinct from a respirometer, which measures oxygen consumption during aerobic respiration.