How to partition free amino acids from proteins in plant extract

Michael Witty mw132 at mole.bio.cam.ac.uk
Tue Jul 31 16:02:19 EST 2001


Dear Tony,
         how about conventional Sephadex chromatography?  Mike.

> Hi,
>     I am trying to quantitatively recover free amino acids away from
> total soluble proteins in a plant leaf extract.  The extract is made of
> 5-100 mg frozen leaf material homogenized in 1 mL of 80% ethanol and
> then centrifuged to remove particulates.  The amino acids in the
> extracts will ultimately be quantified by either HPLC of PITC
> derivatives, or GCMS of MTBSTFA derivatives.
>     I have heard of a method where such extracts are passed through a
> C18 SPE column to adsorb the proteins and recover the free amino
> acids in the eluate.  If this works, it would be an attractive
> alternative to using expensive and time-consuming low-molecular-weight
> cut-off centrifugation or protein precipitation methods.  However, I
> can't find a protocol for such a method using C18 SPE columns (i.e.
> rinse, load, elution conditions).  Can anyone help?
>
> thanks
> Tony
>




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