Could anyone tell me an approximate value of dead volume of Merck LiChrocart 250-4 RP-8 LiChrospher 5mm

thank you

Dear anonymous;

There is a rule of thumb that gives dead volumes, of 4.6mm ID columns, as L (cm)/10 expressed in mL. In other words a 250x4.6mm ID column of most packings has a dead volume of 2.5 mL. Is yours 4mm ID?, then you have to apply a correction proportional to the square of the ratio of diameters, or 2.5mlX (4/4.6)**2 or 2.5 mlx 0.756, that equals 1.89 mL.

Good Luck;

Benjamin

Thank you Benjamin

I will remember this rule throughout my entire analytical career (which by the way I have just started).

Benjamin,
Your rule of thumb seems to be a good approximation, but I suppose the particle size (smaller particles gives denser packing and hence smaller dead volume) and also porosity of the particles may have an influence on the void volume.

No, smaller particle sizes do not give a denser packing.
The particle porosity plays a role, but what are you going to do - it's a rule of thumb.

Guys!
Just measure it!!! :)
Inject water in AcN-water system))) the minimum of first negative peak (200-220 nm) is zero volume (minor disturbance method). A great number of other methods are possible.
Constantine.

You could get a lot of people arguing about this. you are absolutely correct - a great number of other methods are possible, and they all give different results.

Yes, for gel permentation mechanism, for example is always involved. If you need supercorrect physical measurements, you are to consider all these factors, but if you are an analyst -- it`s no use.

What is a supercorrect physical measurement in this case? The retention volume of water? The retention volume of acetonitrile? The retention volume of salt? The retention volume of your compound under conditions when it is not retained? The extrapolated retention volume of a homologous series?
The rule of thumb by Benjamin is as good as any of these things for the average chromatographer. Actually, it is better, because it is simpler!!!

Using an RI detector equilibrate w MeOH/water -say
50/50 then inject a 50 /50 soln of MeOH/D2O. The
inflection will essentially be the void volume.

instead of water u can use uracil to check
dead volume of column in water acetonitrile or methanol water system

If the volume isn't dead, you should kill it. After it is dead, you need to bury it.
Honestly, here is a beautifully simple estimate of the column dead volume that works like a charm:

VD = 0.5 * L * D^2

L is the length, and D the diameter of the beast

I need to add something to you 'killer' anon,
L and D are expressed in cm.
-Mic.Raj

I need to add something to you 'killer' anon(Just kidding),
L and D are to be expressed in cm.right?
-Mic.Raj

There are many ways to measure the dead volume of a column. Unfortunately, there is no general accepted method. Even prominent compendia, like the European Pharmacopoeia or United States Pharmacopeia do not prescribe an appropriate method, although the value is needed in a number of formulas. Many ways to calculate dead volume are described: measuring the retention of uracil, thiourea, nicotinic acid or nitrate. Therefore a lot of confusion remains. There is really need to a general and harmonized method!

The different ways of measuring the dead volume give different results. However, for the practitioner, the only thing that counts is to have a reliable and reprodubible number. For this purpose, any of these methods is fine. What is not OK is to use "the first departure of the base line", as some people do.

I agree with constantine you might inject methanol or ACN to your system.

What detector do you use when you inject acetonitrile and methanol?

UV-VIS dedector

Then how do you see acetonitrile and methanol?

To use acetonitrile or methanol with a UV detector, briefly, you inject the sample and wait for the injection disturbance -- it looks like a sharp negative peak followed by a sharp positive peak -- caused by the flow disturbance due to the injection loop turning. If you system has a pulse dampener, or your column is a wide bore prep column, you may not see anything.

If your mobile phase is an aqueous buffer, and the injected solvent doesn't have those components, the injection will produce a peak due to the sharp change in refractive index. A detector that compensates for RI artifacts may prevent this from working.

Methanol and other alcohols have a higher viscosity than water/acetonitrile solutions. When injected, the system will show a pressure surge as the sample travels through the system. The length of that surge will give you the dwell volume.

All this assumes the injection doesn't interact with the column media at all.

Frankly, I've never had a problem using methanol to determine a system's dwell volume. I never used such a determination to validate a method, however.

How do you know that this injection disturbance gives you a meaningful number? Is it excluded from the pores? Does the retention change whne you throw some salt or buffer into the mobile phase? Which peak do you use? The first negative one? Or the last positive one? Or the value when your zig-zag crosses the baseline? What is the physical meaning of these values? This question is especially important, if the phenomenon goes away when your sytsem has a functioning pulse dampener.
I have some very serious doubts about your pressure measurement. The measurement is over, when the pressure returns to normal, right? I think that this is the point when your sample is diluted enough not to cause any pressure disturbances.

Honestly, these type of measurements are figments of somebody's imagination, but not real measurements of anything real.

For resolution and capacity factor determination, does dead volume include the time/volume from the injector to the column to the detector, or just the column volume?

If it's just the column volume, would you need to do the pressure change once with the whole system and once without the column? Then take the difference?

For other's info:

I use the pressure fluctuation method. I take the time point between the negative and positive peak, where it crosses the baseline.

Reply to A.Mouse.
If analysts use different methods to determine the dead volume, and thereby achieve different results, than these results are not reproducible. Different labs will show different results.

P.D.: I agree with you: people need to agree on a good and useful method to determine the dead volume for measuring the retention factor. What you use should be part of the method. It is not acceptable, if different labs use different methods, and I can not see that this would happen in a well organized environment.
Michael Goodwin: Most people will include the time/volume from injection to detection into the calculation. Unless you are doing physical chemistry, the amount by which this number is falsified by the instrument volume is rather irrelevant and rarely causes any difficulties.

Thanks for clarifying.

However, do keep in mind just like there are different roads I can take home from work, there are different methods I can use to get the dead volume.

I disagree that their needs to be one standardized method for all laboratories. Many analytical labs to date have been successful in their endeavers without a standardized dead volume method.

I would agree that such a method used be stated within their operating procedures.

Michael Goodwin: I did not suggest that everybody should use the same method for determining the dead volume. For the practical work that most of us do, the only thing that is relevant is a reliable method.
When I said that the same method should be used in different labs, I meant that if a method is transferred from one lab to another, the method for determining the dead volume should go with the method. If you want reproducible results for retention data, this is important. If you use different good methods for different assays, I do not have a problem either. But I do have a problem, if one lab is using a method with one dead volume marker and another lab is using the same method with another marker. This is as much ground for trouble as no determination at all.

I misread. My apologies.

which solvent can versatile?

sorry which solvent can b considered as versatile