HPLC: Reversed Phase, Gradients and Method Development · seed 1 · A4, ink-friendly. The answer key prints on its own page for grown-ups.

Tuning a liquid separation

Science · Chemistry · ages 19-21
Name ______________________   Date ____________

  1. On a reversed phase column, which compounds elute first?

    • Nonpolar ones
    • Polar ones
    • Charged metals only

  2. Why is HPLC run at high pressure?

    • To heat the sample
    • To force liquid mobile phase through tightly packed powder
    • To make peaks taller directly

  3. Each HPLC peak is one compound leaving the column, and its time hints at its identity.

    Circle one:   True   False

  4. An acidic drug washes straight through a reversed phase column. What should you try?

    • Raise the pH far above its pKa
    • Lower the pH below its pKa so it stays neutral
    • Switch off the detector

  5. Early peaks crowd and late peaks drag broad in a fixed blend. Which fix fits?

    • Inject a larger sample
    • Remove the column
    • Run a gradient of rising solvent strength

  6. You change the solvent blend and all peaks move. What does that show?

    • Retention times shift with the sticking balance
    • The column has broken
    • Peaks always stay fixed

  7. On a reversed phase column, neutral molecules stick while ions wash through.

    Circle one:   True   False

  8. Early peaks look fine but late peaks are broad and dragging. What do you change first?

    • Steepen the gradient to sweep late peaks out
    • Lower the pressure to slow everything
    • Raise the pH at random
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Answer key

For grown-ups. Fold this page away before handing over the rest.

Tuning a liquid separation W1-mt_hlJl6nXQqT-s1

  1. Polar ones · Polar compounds ride the polar mobile phase and hurry through.
  2. To force liquid mobile phase through tightly packed powder · Fine packing resists flow, so pressure keeps the liquid moving.
  3. True · Retention time under fixed conditions is characteristic of each compound.
  4. Lower the pH below its pKa so it stays neutral · Neutral molecules grip the nonpolar phase while ions wash away.
  5. Run a gradient of rising solvent strength · Rising strength sweeps late peaks out sharp while early spacing holds.
  6. Retention times shift with the sticking balance · Each compound rebalances between sticking and dissolving.
  7. True · Ions prefer the polar liquid and skip the nonpolar powder.
  8. Steepen the gradient to sweep late peaks out · Only the late end suffers, so strengthen the solvent later in the run.
Worksheet · LightMySky