Paper Chromatography (2)

Preparative Chromatography

Preparative = “More” • More mixture – Isolate and characterize separated compds

• More silica gel – Larger container (column v. thin-layer plate)

• More solvent • More time

Agenda • Building (packing) column – Equipment, silica, compression + test

• Applying mixture (loading) • Collecting (eluting) fractions • Testing fractions

Pack Column • Sintered glass funnel + conical adaptors + filter flask (+ aspirator) • Load silica LOOSELY to brim • Compress 1) Rubber or glass stopper 2) Apply vacuum

• Test – Head space – Solvent flow (filter paper protects) – Cavities + channels

Apply Mixture • Make mixture – Weigh mixture (~ 400 mg) – Add few drops EtOAc & mix – Dilute w/ hexane

• No vacuum • Apply entire mixture to paper (spread) • Apply vacuum

Collect Fractions • Insert clean test tube – Funnel tip INSIDE tube

• Apply vacuum – Squeeze (& listen)

• Pour solvent mixture (~25 mL) – Flows into tube (don’t overfill)

• Release vacuum • Remove tube

Solvent Mixtures • 0% EA: 100% H Æ 40% EA: 60% H – Increment %EA between fractions

• Method – Make mixture in 100 mL grad cylinder – Pour (#1) – Add little EA, mix & pour (#2) – Add little EA, mix & pour (#3) – Add little EA, mix & pour (#4)

• Copy recipes into lab notebook (LEFT)

Test Fractions • Collect 20 fractions • Find HOT fractions (absorb UV) 1

2

3

20 19 18

4

5

6

7

8

9

10

17 16 15 14 13 12 11

• Elute HOT (only) vs. 5% unknown • Must get both compds off column 7

U 8

Lucky People Only • Combine fractions that contain same pure compound • Evaporate – Once using rotary evaporator • Switch from large RBF to small RBF (rinse large) • Weigh small RBF

– Again using vacuum pump • Weigh small RBF