Effect of Peak RT on Area

[shamitakohli]

What is the effect of Peak retention time on peak area?

[Mattias]

In the ideal case: none.

However if the change in retention time is due to different flow, the peak area will be affected. Double the flow and you will get 50% of the area.

I am not sure if the area will change if you increase or decrease the amount of organic. It is probably negligible in the normal case, but I can imagine that the UV-response of a compound can be different in e.g. 5 vs 50% acetonitrile.

[nschwartz]

Peaks with longer retention times usually have decreased peak height due to broadening, although peak area should be about the same. Keep in mind that as the peak broadens, there is less certainty, i.e., more error in measurement of the area.

[shamitakohli]

Thanx Mattiaz & nschwartz

[unmgvar]

it depends what causes your shift in retention time

for example if you only change the composition ration between buffer and organics then it does not matter the shape the peak area stays about the same

if you change the flow then the area will change in accordance
less flow more area and opposite

if you change the organic or mostly the buffer pH or type of buffer then with the same RT or other RT, the area will change.

i do not remember right now the effect of temperature

[HW Mueller]

unmgvar, how does

"for example if you only change the composition ration between buffer and organics then it does not matter the shape the peak area stays about the same"

jive with

"if you change the organic or mostly the buffer pH or type of buffer then with the same RT or other RT, the area will change."
?

[unmgvar]

HW

softwares like chromsword help you develop separation methods
one of the trick they use is to follow the peaks as they change RT,
but the area stay virtually unchanged if you only change the ration between the A and B phases and they are the same. the software then knows to follow the compounds and learn their separation behavior and from there it helps you find the better suited method for your application

but have you taken a buffer and changed the pH from 2-3 to 6-7 and then seen the compound behavior change, like for an amine and this effects the spectra as well and then the area change using the same WL?

[HW Mueller]

unmgvar,
I am not sure about what you are trying to say, but it is known that solvent polarity changes and pH changes can influence absorbance, thus Chrom. areas (UV-VIS detectors).

[unmgvar]

Dear HW Mueller
maybe i should take extra care to be more explanatory in the future for you so that you do understand what i mean.

I do think however that i was detailed enough,
but if you like, we can make an issue of it. like several times in the past.
i do not think it would be our, but mostly your first time.

[Alexandre]

The longer compounds are in the column, the more interactions, sometime irreversible with the column materials they have. For a certain retention range, say if Rt is 5 min, the Rts 4-6 min effect is normally negligible, however if you observe strange effects it has to be investigated. In UPLC of complex samples I observed coelution resolved.
For most environmental, biological methods effect of Rt on area due to column material interaction is minute compare to other factors contributing to method’s uncertainty.

[Gilson_PLC2020]

A later RT will result in a shorter fatter peak, this means that both the start and the end of the peak become less defined. As such the peak may be seen as smaller due to small amounts of the front and tail 'blending in' with the baseline. For an earlier peak the reverse is true.

Assuming suitable resolution is obtained earlier RTs are preferable.