By Anonymous on Tuesday, April 13, 2004 - 02:11 pm:

Hi,

I am devloping new method for some quaternary ammonium compounds using ion-pair mechanism (sodium salt of heptane sulfonic acid and diethyl amine at PH=4). I got not reproducible results based on peak areas while reproducible results comes from peak heights. By contrast i got vice versa with nitrate analysis. so I am wondering when will chemist use peak area or height in HPLC chromatography.

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By A.Mouse on Tuesday, April 13, 2004 - 03:16 pm:

Both should work the same way, typically area gives better results. If you have problems, I would check the pulsation of the pump.

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By M. Stone on Tuesday, April 13, 2004 - 04:44 pm:

There are a few cases where peak height integration can be a better approach than peak area integration. For one, if you are at low levels or there is a lot of baseline noise. Second, if the peaks are tailing (with a consistent profile) such that it is hard to know where the peak ends. Third, if you have a concentration sensitive detector and the flow rate is not constant.

Peak area is better in most other cases. Especially if your peak profile is not exactly the same from run to run (for example if the column is not completely equilibrating between runs) or if you have a mass-sensitive detector and the flow rate is not constant.

Good luck.
Mark

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By HW Mueller on Tuesday, April 13, 2004 - 11:25 pm:

A recurrent question, it seems, though, that the prime reason for using Hight was badly overlapping peaks.

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By bill tindall on Thursday, April 15, 2004 - 08:05 am:

Agreed with HW. Back when I was doing complex environmental samples pk ht gave better precision for unresolved peaks, especially when they are not all the same width.

There is more area at the bottom of a peak. hence, if there is uncertainty in locating a reliable baseline, or figuring out how much tails overlap, then the baseline error on the height is less than on the area.

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By lmull@jhsph.edu on Tuesday, June 8, 2004 - 02:15 pm:

I actually have a question and am at my wits end. I am trying to
quantitatively detect AdoHcy and S-AdoMet in a plant extract
done in o.25M HClO4. I am using a C-8 with 50mM NaHPO4 and
10mM Heptanesulfonic acid, pH3.2 monitoring absorbance at
254nM. My flow rate is at 0.6mL/min. and my retention times
are consistent between runs.
My problem is reproducibility among sample groups for each
compound. I have run standard curves with control reagents and
use a quadratic equation to reverse integrate the peak areas in
order to determine the uMs of each compound. I am maxed at
the amount (95uL) of acid extract I put on the column. My
STDEVs are larger than some of my sample values even if I
normalize to another peak area. HELP or I shall never
graduate!!!!!!!!!!!!!!!!