By paolino on Tuesday, June 22, 2004 - 04:44 am:

I had a strange shaped peak in an RP C18 analysis monitored by ELSD. One peak eluting at 7 mins shows extra peaks while the peak intensity is decreasing. In its right side the peak looks like an NMR multiplet.
The column is not overloaded or badly-packed. There are no dead volumes.
Has anyone had a similar problem? If so, what is the solution?

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By Kostas Petritis on Tuesday, June 22, 2004 - 10:49 am:

Paolino,

Have you checked with another detection mode the peak shape? This will answer if it is the column/tubing/valves or detector to be blamed.

What is the flow rate you operate and which ELSD do you use?

Kostas

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By Anonymous on Wednesday, June 23, 2004 - 09:31 pm:

Sometime, the ELSD detector can give the peak shape like that, depending on the flow rate and droplets size distribution. If the nebulizer doesn't function well, some large droplets travel later than most fine droplets and it gives additional peak due to higher scattering response. It could be fixed through increasing the gas flow

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By paolino on Thursday, June 24, 2004 - 07:55 am:

The flow comes out of the column at 1.0 mL/min. Unfortunately my compound is not UV-visible at any waveleght and RID is not sensitive enough to detect it. So I can't check the peak shape by another kind of detector.
The ELSD is a brand new SEDERE Sedex 85 and the gas pressure is set at 3.5 atmosphere. Increasing the temperature of the evaporative chamber yielded only a slightly better peak shape, with a loss of sensitivity of course.
I haven't increased the gas flow yet. The upper limit is 4.5 atm. I'm going to check as soon as possible.

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By Kostas Petritis on Thursday, June 24, 2004 - 04:23 pm:

Paolino,

The flow rates that you are using are normal so it can not be what the Anonymous indicated (if you were using more than 4 mL/min then it could be a concern but then again not with the Sedex 85).

Have you seen this with other compounds of about the same retention? What happen when you decrease the injected amount of your coumpound? Is your compound kind of semi-volatile?

Kostas

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By paolino on Friday, June 25, 2004 - 12:48 am:

Kostas,
the compound is not volatile.
There must be something wrong with the chromatography itself rather than with the detector, despite the fact the method has always worked well so far.
I systematically checked any parameter of the method: the aqueous phase composition, molarity and pH, the % of acetonitrile, the flow, the column itself.
Every seemed to be OK but the peak has a terrible shape and I haven't a clue about where I'm wrong.
In my tests peak shape gets better at shorter retention times - as it should be - this is why I guess there something wrong in the chromatography itself. Anyway I'm more than curious about why this method decided not to work anylonger.

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By Kostas Petritis on Friday, June 25, 2004 - 08:41 am:

Paolino,

You seem to imply that you have a generic LC method for all your analytical needs. Maybe it is that your method does not work very well for that particular compound and another column/mobile phase composition must be developed for it...

Kostas

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By paolino on Tuesday, June 29, 2004 - 04:15 am:

Kostas,
The method indeed has worked well so far in other R&D labs but as soon as it was started on my HPLC station it began giving me troubles.
Now it seems to work much better as I lowered the molarity of the acqueous phase from 10 to 4 mM.
The peak shape is nearly acceptable although I still have problems of the retention times which change from day to day.
I'd rather stop worrying with this method and create a brand new one...