What are the three steps in the 3-step precipitation method?

• A. Precipitation, Centrifugation, Trinder Coupled Enzymatic Reaction
• B. Precipitation, Filtration, Colorimetric Reading
• C. Extraction, Centrifugation, Enzymatic Reaction
• D. Precipitation, Vacuum Drying, Enzymatic Reaction

Answer: (A)

The question tests understanding of the three-step precipitation method, which uses an order designed to remove interfering substances and then generate a measurable color signal. The first step is precipitation, where a reagent pulls proteins and other potential interferents out of solution. This cleanup helps prevent false or biased color readings. Next is centrifugation, which pellets the precipitated solids so the analyte remains in the clear liquid and a clean supernatant can be analyzed. The final step is a Trinder coupled enzymatic reaction, which on the clear extract produces a colored product whose intensity can be measured and is proportional to the analyte concentration. This sequence—precipitation to remove interferences, centrifugation to separate solids, followed by enzymatic color development using a Trinder reaction—best fits the method described.

The other options don’t fit because they replace essential components of the method: skipping the enzymatic color development would leave you without a measurable color signal; using filtration or extraction in place of centrifugation changes the cleanup approach; and including vacuum drying isn’t part of this protocol.