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effect of flow rate??

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Hi

What is effect of flow rate on sensitivity and resolution ???
(P.S:- plz let me know if it effects other parameters also – the part w.r.t to pressure is little bit clear to me)

Thanks

Peaks are getting a little bit wider (in volume units) with increasing flow rate (above the minimum of the van Deemter curve). Thus the sensitivity drops and the resolution decreases. The change in sensitivity is small enough to be ignored, the change in resolution may or may not be relevant, depending how close the elution of a peak pair is.

The pressure increases in direct proportion to the flow rate.

hi

recently i ran one of my sample under two different flow rate ( 1 ml/min and 0.8 ml/min)..and found that by running method at 0.8 ml/min..post-main peak- impurities have merged completely main peak , while pre peak impurities were resolved little more in comparison to the chromatogram obtained with 1 ml/min…
one of my colleague suggested this may be due to diffusion under low flow rate….but if so then why only this affects post peak impurties???..plz help me clarify this doubt..

thanks

Hi Rick

I assume the retention times were prolonged at 0.8 mL/min compared to the 1 mL/min setting.
If that is the case then diffusion is the explanation – which is proportional to the time a compound spends on the column. And because the latest peak is retained for longest time it’s also the most affected.

However, if you had adjusted the retention time by increasing the strong elution component in your mobile phase so that the retention time for e.g. the main peak was constant at both flow rates, then the explanation would be of a chemical nature (selectivity, solubility etc.)

Best Regards
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Dancho Dikov

Rick,

The event that you are describing is rather unusual for an isocratic method. Are you running a gradient? If that is the case, the story is more complicated due to the fact that you are changing the gradient slope. If this is the case, I will explain it further in a later post.

hi Uwe Neue,

yes its a gradient elution method, and in both case i have kept all other parameters same expect the change in flow rate...

When you change the flow rate in a gradient while keeping everything else constant, you are really changing a lot more than it appears on first glance.

When you decrease the flow rate in this case, you are creating a smaller gradient volume, and you are making the gradient steeper. This may or may not change the selectivity of the separation, but it definitely decreases the true retention factor at the column outlet. This makes all your peaks narrower (in volume units), which in turn will give you a better response factor (higher peaks). However, the resolution may get worse, since the distance between the peaks may decrease more than the peak width (both in volume units).

Overall, the expectation is that you will loose resolution, but this may depend on the details of the elution pattern of your analytes. You have seen this: you lost resolution at the beginning of the chromatogram and gained some at the end of the chromatogram.

In principle, you should be able to observe the same pattern as you have observed at 0.8 mL/min by running the chromatogram at 1 mL/min with a gradient that has only 80% of the run time of the original gradient. (There are additional complications due to gradient delay volume that can create additional effects, but let us not worry about this at the moment.)

hi

does this means their is no effect of diffusion and it is more related to "creating a smaller gradient volume"???

also does'nt narrower peak means more separation??(plz forgive me if i am wrong) ...

is there a relationship between resolution and flow rate???(the resolution equation doesnt have any term related to flow rate..plz correct me if i am wrong..)

To 1: The effect of plate count and diffusion is smaller than the effect of the change in the gradient slope.

To 2: Narrower peaks do not always mean better resolution. In the case of gradients, you will get the narrowest peak by running a step gradient. But if you do that, all peaks will overlap.

To 3: The peak width is part of the resolution equation, and it depends on the plate count. The plate count in turn depends on the flow rate. This is straightforward for isocratic chromatography. When you do gradient chromatography and conduct an experiment as the one described by you, you are also changing the position of different peaks towards each other, and this can complicate things.

If you want more info, I will need to try to write equations...
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