Peak width decreasing as ret.time increases!Logical or not?

[project44]

Hi there,
I want an opinion and any possible advise about some results i have extracted a few days ago.
I run a gradient elution program for the separation of two compounds(tetracyclines) and i observed the following situation: The late eluting compound(12,3min) and its main metabolite(12min) had narrower peaks than the earlier eluting peak(5min).From theory i know that as retention time increases, the width of a peak also increases and the plate number decreases.
Is there any mistake for this elution?I mean, is everything ok with the peak width of the late eluting peaks? I shall note that the first peak has As=1,59(not good enough)
(I use a mobile phase ACN: KH2PO4 50mM pH8 30:70 v/v as a starting point).
The gradient has as follow:
1-5min 30% ACN-70% buffer
5-8min 65% ACN-35% buffer
8-15min 65% ACN-35% buffer
15-17min 30% ACN-70% buffer
17-30 min 30% ACN-70% buffer
The column's dimension are: 250mmx4,6mm 5μm and the flow rate is 1mL/min.
Thanks for any suggestion and advise!

[danko]

The peak at 5 min is eluting under isocratic conditions whilst the peak/peaks at 12 min under gradient conditions.

Best Regards

[project44]

Hi Danko, thanks for the help...but is it acceptable anyway?
Thank again...

[danko]

You bet it is. No lawyer will dream of suing you for that

Best Regards

[project44]

Thanks, I TRUST U!

[tom jupille]

The only other comment that I would add is that the rule of thumb that later peaks are wider applies only to isocratic separations. In gradients (to a first approximation) all peaks have about the same width.

As danko pointed out, your 5 minute peak is eluting isocratically before the gradient part of your run.

[project44]

Thanks a lot for your help!
Regards