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column chromatography - (Mar/03/2013 )

What calculations do we use for packing any column.i.e. how much volume of beads should be taken for how much compound? For different types of adsorbents, the calculation vary. Like when we run silica gel column we take 1gm slurry for 10mg compound (???)...what about c18, g25 etc.?..I have tried to find out on google for some basic calculation..but did not succeed.. ..so experts please help me, I am confused about column chromatography

-neuron-

the mode of separation is important in determining how much to use.

for modes which bind the sample you should find the binding capacity on the bottle (or a specification sheet) in which the matrix comes. personally, if i'm packing the column fresh for each sample, i would use ~10% more than required to bind the sample.

with stationary phases like g25 (gel filtration) the important factors are width and length of the column. for classical lc you want a separation length of at least 50 cm (we prefer 100 cm for packings more stable than sephadex). for hplc/fplc 30 cm is the acceptable length (when longer is needed 2 or more columns can be run in tandem).

you can download some useful handbooks about the various chromatographic methods (and more) from this page at the ge healthcare website.

-mdfenko-

Thanks mdfenko, the link is useful..:)

-neuron-