I always look forward at your replies.And,just for this,I did some
major editing to my original reply.I was having some vision problems,
but this is not a good excuse for the errors.It should be more
understandable.You correct that in most cases it does not matter if there
is any of the D-form,except if you want to use the compound in enzymatic
assays.Enzymes are very specific for L-amino acids.
Vladimir Titov <vmtitov@aha.ru>@aecom.yu.edu> on 08/01/2000 08:08:03 AM
Please respond to Vladimir Titov <vmtitov@aha.ru>
Sent by: Association of Biomolecular Resource Facilities
<abrf-request@aecom.yu.edu>
To: Recipients of ABRF List <abrf@aecom.yu.edu>
cc: Recipients of ABRF List <abrf@aecom.yu.edu>
Subject: Re[2]: Racemization/polarimeters
High resolution NMR detects diastereomers, but you need an expert to
interprete the spectra. HPLC often works, but not always. Polarimetry
is not the method for peptides, because optical rotation depends on
concentration of peptide in the sample which is subject to another
analysis. Threrefore, the errors from the two analyses would be
multiplied. I have heard about AAA on chiral phases but have never
seen anyone who used it. In my cinical opinion, for most research
there is no difference which substance is used at all, so it's better
to forget about racemization and present HPLC and mass spec. If you
are producing a pharmaceutical compound, it makes a difference but you
would have resources to study the compound and develop analytical
methods.
On 31/07/2000, fperini@unmc.edu wrote:
> I have been spending a good deal of time trying to get an universal
> procedure,and I
> found out that no matter which method I tried,it does not work all the
> time.
> If the peptide is relatively small,and I will not be pinned down on
size,a
> reverse phase HPLC separation will do the job.If you can get the same
> column,conditions,etc.as the initial
> run,it should work when combined with amino acid analysis.Once,I got a
> peptide which gave
> baseline resolution of two peaks of identical size.I found out that the
> researcher
> synthesized the derivatives,using the D,L-amino acid,but it was only
five residues in length..
> To get the percentage of D-amino acid is not so easy.I have seen
> publications,which took advantage of the chiral properties,and I tried
> the procedures.They work for only a few amino acids.Probably,the best
> method is that of
> N.Nimura and T.Kinoshita,J.Chromat.,vol.352,169-177(1986).It relies on
the
> reaction of
> primary amino acids with o-phtaldehyde(OPA),in presence of a sulfur
compound
> which is usually in
> the L-form.A ternary complex is formed,and it is possible to separate the
> one with the
> L-amino acid from D-amino acid,by the fact that there is a L-L and D-L
> form.The authors and
> I have been using N-acetyl-L-cysteine,although other sulfur compounds
have
> been used.
> It is a precolumn derivatization and detection is fluorometric.I do not
> think that any
> other physico-chemical method will work.
Vladimir Titov
Bokiron Ltd., Moscow, Russia
This archive was generated by hypermail 2b29 : Fri Aug 18 2000 - 07:56:59 EDT