Sensitivity of 1mm over 4.6mm column

[project44]

Hi all, i have a small question and i would like your opinion.

I have a column of 150 x 4.6 mm, 5μm and another one of 150 x 1 mm, 5 μm.
If i inject the same concentration of an analyte in both columns, how much sensitivity do i gain from the 1mm column over the 4.6mm column? (i use a uv detector).
I tried to answer this by thinking the dilution factor between the two columns, that is the void volumes (Vm) of the columns. The 4.6mm column has a Vm=1.5mL and the 1mm column Vm=0.080mL. So, the question is: am i correct if i imply that the sensitivity of the 1mm column will be increased by a factor of x20 over the 4.6m column?

Thank you all, in advance!

[jiang295]

i think you are right.

Hi all, i have a small question and i would like your opinion.

I have a column of 150 x 4.6 mm, 5μm and another one of 150 x 1 mm, 5 μm.
If i inject the same concentration of an analyte in both columns, how much sensitivity do i gain from the 1mm column over the 4.6mm column? (i use a uv detector).
I tried to answer this by thinking the dilution factor between the two columns, that is the void volumes (Vm) of the columns. The 4.6mm column has a Vm=1.5mL and the 1mm column Vm=0.080mL. So, the question is: am i correct if i imply that the sensitivity of the 1mm column will be increased by a factor of x20 over the 4.6m column?

Thank you all, in advance!

[project44]

Yes, i agree with the first state. Increase in peak height. But how do you explain the 5 fold increase? (u got the ratio 4.6mm/1mm?)

it differs in peak height and peak area.
peak height wont change but peak area will increase 5 fold.
need for other expert answer.

Hi all, i have a small question and i would like your opinion.

I have a column of 150 x 4.6 mm, 5μm and another one of 150 x 1 mm, 5 μm.
If i inject the same concentration of an analyte in both columns, how much sensitivity do i gain from the 1mm column over the 4.6mm column? (i use a uv detector).
I tried to answer this by thinking the dilution factor between the two columns, that is the void volumes (Vm) of the columns. The 4.6mm column has a Vm=1.5mL and the 1mm column Vm=0.080mL. So, the question is: am i correct if i imply that the sensitivity of the 1mm column will be increased by a factor of x20 over the 4.6m column?

Thank you all, in advance!

[jiang295]

I think peak height definitely gain as you said. but peak area will vary with your flow rate your set for each column.

Hi all, i have a small question and i would like your opinion.

I have a column of 150 x 4.6 mm, 5μm and another one of 150 x 1 mm, 5 μm.
If i inject the same concentration of an analyte in both columns, how much sensitivity do i gain from the 1mm column over the 4.6mm column? (i use a uv detector).
I tried to answer this by thinking the dilution factor between the two columns, that is the void volumes (Vm) of the columns. The 4.6mm column has a Vm=1.5mL and the 1mm column Vm=0.080mL. So, the question is: am i correct if i imply that the sensitivity of the 1mm column will be increased by a factor of x20 over the 4.6m column?

Thank you all, in advance!

[project44]

Yes, but the question is a little more specific. I ask how much will the sensitivity be increased and how this property can be described by e.g. an equation which explains this increase. I proposed my explanation, but i don't know for sure that this is true. I would appreciate some more help.
Thanks again.

[jiang295]

roughly as you said.

Yes, but the question is a little more specific. I ask how much will the sensitivity be increased and how this property can be described by e.g. an equation which explains this increase. I proposed my explanation, but i don't know for sure that this is true. I would appreciate some more help.
Thanks again.

[Consumer Products Guy]

I think that's assuming that you inject the same volume of sample/standard. Oftentimes you'll need to inject less volume with the smaller-diameter columns, especially if your injection solvent is straight or mostly organic.

[Jumpshooter]

1.5/0.08 = 18.75; therefore, sensitivity will be up-ticked by ~20-fold.

In practical terms, this would manifest as an injection of 10 uL of a 10 ug/mL would give a peak area nearly equivalent to an injection of 10 uL of 0.5 ug/mL sample, when the injects on the same sample were compared in Relative terms. This in turn would suggest an LOD that could be stated as 'lower" i.e.,yielding a "more sensitive" assay.

[project44]

Thanks for the responses, i feel much better now!!

[thohry]

the small column has its own drawback: extra column volume dispersion, column load etc.. so you may not have as much gain in sensitivity.

[mbicking]

The correct way to look at this is the peak volume for each column.

The general equation for the 4 sigma peak volume is:

Vp = 4*Vm*(1+k)/sqrt(n),

Where Vm is the void volume.

If you divide the equations for each column, the ratio becomes Vm2/Vm1, which can be simplified to dc2^2/dc1^2.

In other words, it is the ratio of the square of the column diameters. In this case = 21.16.

If you inject the same mass (in the same volume), it elutes from the smaller column in a volume that is 21 times smaller. Therefore, the concentration is 21 time larger, and a concentration sensitive detector like absorbance would show an increase. A mass sensitive detector however (MS) would not.

As noted, the same injection volume would probably overload the smaller column, and the peak shape would be poor. It can be even split in some cases.

Typically, you will reduce the injection volume in similar proportions, so that this is not an issue. This means that a 5 uL injection on a 4.6 mm column should be adjusted to a 0.25 uL injection on a 1.0 mm column.

[project44]

Thank you very much!

So, if i understood well, there are 2 distinct ways that contribute to the increase in sensitivity: 1) ''Ab initio'' due to the diameters of the columns (the peak volume for every peak in the 1.0 mm column will always be 21,16 times lower, and 2) from the necessity of reducing the injection volume (keeping the mass constant) in the 1.0 mm column.

Thanks again!

[mbicking]

Let me clarify, because your point 2) is not exactly what I meant.

If you inject the same solution (same concentration) at the same volume, you will get a larger peak on a more narrow column, because of the peak volume (your point 1)).

To avoid problems with overloading and extra-column effects, most methods will simply reduce the injection volume of the same solution (same concentration). The result of this is that you in fact lose the advantage in response. People use the narrow bore columns to save on mobile phase, not increase sensitivity.

In practice, you don't have to follow this rule exactly. For example, rather than decreasing your injection volume by 20X, you could decrease by 10X, and if you still get good peak shape, you should see an increase in sensitivity.

Most LC systems would not allow you to see the benefits of a 1 mm column, which is why I prefer to switch from 4.6 to 3.0 mm columns. The flow rate is reduced by 60%, but often I can keep the same injection volume, or only decrease it slightly, not by 60%. And many LC systems will not degrade the performance.

[project44]

yes you are right, my comments above were not exactly what you meant before. However, i think that comment 2) is still valid, when the final purpose is to increase the sensitivity.

The problem in my situation, is that i don't have the luxury of sample availability. So, by using a 1.0 mm column i get the advantage of both the comments ( 1) and 2) ). If i reduced my injection volume (keeping the mass constant) in the 4.6 mm column, i would gain sensitivity only from the ''dilution factor''.

Anyway, this is not an offence to your comments. You helped me very much, and i appreciate the advice from the experts.

[mbicking]

Yes, you can proceed as you describe, but be aware that simple concentration overload can still occur, even with a small volume injected. So, if you have peak shape and retention time differences, you may have to reduce your concentration somewhat until the problem disappears, or at least becomes acceptable.

Good luck.