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Chromatography using acetone!

  1. Oct 2, 2004 #1
    chromatography....using acetone!

    the protocol:

    Vg = volume not accessable to solvent (volume of the gel matrix)

    Vg cannot be measured directly. However, it should apparant that:
    Vg = Vt - volume accessable to solvent

    We can measure the volume accessable to the solvent, using a small molecule such as acetone which can be easily visualized by a UV detector.

    example:
    Pharmacia Superose 6 column - volume accessable to solvent = 19.5 mL (measured with acetone)
    Vg = Vt - 19.5 mL = 24.4 - 19.5 mL = 4.9 mL

    ---------------------------------------------------------------------------------------

    hi guys!
    i just find this calculation very unreasonable, since as we know Vt=Vi+Vg+Vo,
    where Vt =total column volum, Vo= void volum due to EXTREMELY huge substances, Vg= volum of matrix and Vi=volum inside the beads due to EXTREMELY small substances.

    my question is, how come we use acetone (small molecule) to determine both Vo and Vi, since Vg= Vt- (Vo+Vi)?


    hopes for replies!

    thanks!
     
  2. jcsd
  3. Oct 2, 2004 #2
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