By karen on Tuesday, May 28, 2002 - 06:42 am:
-I am at a dead end with troubleshooting a system. We are using an established liq/liq extraction procedure with DPBEA to analyse for noradrenaline/adrenaline/DHBA. Chromatograms are consistently giving extra peaks between NA and A and a late one which carries over.It only appears when samples are extracted and only on this system and cell. Standards are clean.
I have changed all reagents and mobile phase, cleaned the rheodyne and cleaned the electrochemical cell (Antec flexcell with in situ Ag/AgCl ref)to no avail.
Column- Hypersil c18 2.00x100mm
mobile phase - sodium acetate,EDTA,sodium lauryl sulfate, 3mM NaCl(for cell), 20% MeOH.
Potential- 0.65V
any suggestions gratefully received!
By banquet on Wednesday, May 29, 2002 - 02:28 am:
Karen,
Try an alternative extraction method. Are there any other products that you may be inadvertently extracting that are giving you your extra peaks?
By karen on Thursday, May 30, 2002 - 08:33 am:
Thanks banquet- not aware of other methods for plasma cats. I don't think that we are inadvertently extracting extra components as I can run my extracts on one of our other systems without any problems, although this does have a slightly different cell with a salt bridge ref. I suspect that something strange is happening with the chemistry in the cell.
By Anonymous on Saturday, June 29, 2002 - 05:48 am:
Call ESA. They really know cats and and EC detectors. Or, better yet, call the distributor who sold you the Antec detector.
When you purchase a piece of LC equipment you are also buying a piece of the company. Now you'll see what you bought.
Plasma cats by HPLC-ECD is easy if you have the right procedure.
By Marcelo on Friday, July 5, 2002 - 01:17 pm:
I don't agree. Plasma catecholamine assay is trace analysis in a very complex matrix, I couldn't call it easy.
Karen, if you extract diluted standard solutions
Does appear the interference?
Have you tried different potential (+0.55 V)
Have you passivates your stainless steel surfaces, including sample loop?
Is the cell using a soli state reference electrode + KCl in the mobile phase? In my experience, KCl 3M/salt bridge ref has worked better.
Regards
Marcelo
By Anonymous on Wednesday, July 10, 2002 - 06:46 am:
Hi Karen,
There are many published papers on solid phase extraction of plasma catecholamines (and urinary catecholamines) using acid treated Alumina. Perhaps you can look into it. The recovery is about 60%-70%, but reproducibility is good, less extraneous peaks too.
Good Luck!
By newtonchristopher on Thursday, July 11, 2002 - 03:56 am:
1. As you said standards are OK. Then the problem is fixed to your sample preparation.
2.Simulate a blank preparation with out sample and analyse as per the same method. If you observe the extra peaks in that try out the possibilities of discarding one component at a time in the sample extraction and run the chromatograph. That is it. You will get it done.
3. In my opinion Acetonitrile is better for ECD than using Methanol. The extraneous peaks may be due to ammonium salts or amines contamination in the sample matrix.