Back to the Top
The following message was posted to: PharmPK
Dear All,
I would like to know some informations about chromatography analisys of
chemical strutures with fenol groups. Have any derivation assay to
stabilizy
the compound ? We have been some problems with HPLC assays for this
substances.
Thanks,
Daniel
Back to the Top
The following message was posted to: PharmPK
Dear Daniel,
I'm not an expert on HPLC, but I think your problem with phenols may be
caused by transition metals such as iron. The problem can often be
solved
by using a protected stationary phase such as Zorbax RX or SB (there are
many other column packings which will work). Traditional column
packings
have enough residual metal ions to cause severe tailing of some
analytes,
particularly if they chelate iron or nickel.
Derivatisation is possible, but you will also derivatise much of the
matrix,
and change all the retention times.
Goood luck.
Ted
Back to the Top
Hello Daniel: you may want to consider including a small percentage of
antioxidant in the mobile phase and the soutions of the analyte. Such
a strategy should stabilize the phenol.
Sodium metabisulphite may be suitable.
Angus McLean Ph.D.
BioPharm Global Inc.
8125 Langport Terrace,
Suite 100
Gaithersburg,
MD 20877
301-869-1009
301-869-5737
BioPharm Global Inc.
Back to the Top
The following message was posted to: PharmPK
what are the problems?
Back to the Top
The following message was posted to: PharmPK
Dr Emary,
We are using GC-MS to analise urine samples of rat in a research about
phenols. The standard samples of phenol when injected in chromatography
system do not provide any response. We tried to derivate with PFP/PFPA
but
nothing have been improved.
Is there another derivative agent that may be used ?
Any informations will be welcome.
Daniel
Back to the Top
The following message was posted to: PharmPK
A more detailed question would have been lot helpful to get a precise
answer
a general approach (with reasonable recoveries of > 75% and good
chromatography)can be found in the following link.
http://www.jtbaker.com/techlib/documents/en-502.html
Without much information given in the question, I am assuming three
problems
with phenols 1) Stability, 2) poor recovery and 3) poor peak shape.
Stability of the phenols can be addressed by adding anti-oxidant in the
sample and extract Dr. McLean already addressed this issue, if you are
concerned about using a harsh anti-oxidant, try to use ascorbic acid or
you
can use BHA and BHT if you are using non polar solvents and
chromatographic
interference from BHA and BHT is not an issue.
Recovery: can be corrected by using the strategy outlined in the above
link.
Peak tailing: OMG you got lot of work to do, adding different end cap
modifiers in the mobile phase such as tetra butyl ammonium hydroxide,
tetra
methyl ammonium hydroxide. Often times adding acetic acid to the mobile
phase do the trick (don't ask me for the mechanism).
Derivitization!!!!! There were couple of articles published in J.
Chromatography, that might help you. Usually you have to pick
derivitizing
agent taking your matrix into consideration. When you have nothing
tosylation is the general approach.
Have you tried DRYLAB simulations? For optimizing the chromatographic
work.
Hopefully this helps your lab work.
Regards,
Prasad Tata
St. Louis, MO 63134
Back to the Top
The following message was posted to: PharmPK
Hi Daniel,
I would have thought your approach should work. I assume you used
negative
chemical ionization and did a full scan analysis of your PFP derivative
and
compared against a control sample (don't jump right to MRM or SIM). HF
pops
off easily for PFP and HFB derivatives and sometimes rather than
starting
with a M- radical ion, you possibly may have an abundant (M-HF)-
radical.
You probably already know this but the sample needs to be very dry
before
starting the reaction and sometimes a little mild base like trimethyl
amine
helps (I think deprotonates the phenol so it can attack the acyl group).
Pentafluorobenzoyl chloride is an acylating reagent for negative MS
work.
BSTFA or BSA bis(trimethylsilyl)acetamide are popular silyating
reagents,
BSTFA part # 15243 from Fluka. It will hit all your active hydrogen
atoms
of other moieties also and works in positive ion mode.
Dan Knapp/ Handbook of analytical derivation reactions has a textbook
covering conditions for most things you would want to do.
Before all the commercial stuff became available, people used to make
the
ether derivatives of alcohols with diazomethane or CH3I/base but those
are
difficult things to work with (diazomethane can explode) and potentially
carcinogenic and would only be used carefully after other approaches are
exhausted.
PharmPK Discussion List Archive Index page
Copyright 1995-2010 David W. A. Bourne (david@boomer.org)