Acquity UPLC and conventional columns

[indium]

Is there any problem with using a 150 x 4.6 mm, 3um or 150 x 3 mm, 3 um HPLC column in the Waters Acquity UPLC system? Since we mainly have "conventional" HPLC systems (max. 6000 psi), I was wondering how easy it would be to move back and forth between the UPLC systen and a conventional HPLC instruments? Let say the above columns were to be used with a flow rate of 1 mL/min (4.6 mm) or 0.42 mL/min (3mm) and a 50 uL injection of a 0.5 mg/mL solution of an analyte. Would modifications of the UPLC system be required?

[Uwe Neue]

The UPLC system is capable of running the flowrates that you intend to use with your classical columns. While you will not get the performance advantage of using the UPLC columns, you can still use the classical columns.

[ravi]

however, going from an optimised UPLC system to an HPLC will require modification

[AA]

As was said in the previous post, traditional HPLC columns can be used in the UPLC system. A word of warning though, be sure to set the high pressure cutoff in the software to protect those HPLC columns. The ACQUITY system is quite capable of pumping those tradition columns past the point of breakage. It is not usually a problem with 5 um, 4.6 mm id's, but may be with 3 um. As you drop in diameter with the traditional columns the posibility of damaging these comulns increases.

I am unsure what you are trying to say in your second post. The ACQUITY system is optimized for UPLC work without modification.

AA

[indium]

By modifications, I was thinking about flow cell changes in particular. I assume the Acquity system has a flow cell designed for smaller injection volumes and lower on-column amounts that the 1 and 2 mm id columns would require. If a 4.6 mm id column is used for the same analysis then a larger on-column amount or larger inj volume would be required for an equivalent analysis.

[Uwe Neue]

The detector flow cell is not a problem. However, the injection volume is limited due to the higher performance characteristics of the normally used UPLC columns.

[ravi]

It will be difficult to properly compare results from and UPLC and HPLC in a quantitative way. Be aware of the flow cell size, dead volume, injection volume. If you're hoping for a validated method, some re-validation would be required to ensure proper transfer.

[Uwe Neue]

I do not see what the fundamental difficulty is for a comparison of a standard HPLC with standard detector cells to a UPLC system with a UPLC detector. You inject, and you get peaks. The peaks on the UPLC system might be a bit narrower, but maybe not, depending if you have a lot of extra-column effects on the HPLC system or not.

[indium]

I am not sure if I was clear in explaining my question. Since we have only one Acquity system and many conventional LC systems, we would like to use standard HPLC columns in the Acquity system (for now) so the methods can be run on conventional LC systems. I was wondering if there would be any practical issues using a conventional HPLC column (e.g. 150 x 4.6 mm, 3 um) with the Acquity UPLC system.

[Kostas Petritis]

Indium,

No you won't have any practical issues other than:

The injection volume is limited (Uwe can specify the exact number).

Most conventional columns are packed at about 8000-10000 psi. If your backpressure will increase more than the pressures your column was packed (i.e. 14000 psi) then you might observe the creation of some void volume in the head of the column with all the consequences. This can be avoided by setting a high pressure limit as AA mentioned.

There is a pressure (I can not remember the exact number) from which 4.6 mm ID columns start to heat up. Smaller ID columns (i.e. 2.1 and less do not have such a problem)up to 20000 psi.

Other than that there are no more issues I can think of.

[BenNC]

My understanding was that the pump on the Acquity will only go up to 2 mL/min so you may be limited there.

Also, the Acquity uses high pressure mixing. If your other systems use low pressure mixing, then you may see a difference if you try to tranfer a gradient method between the two instruments.

[ravi]

As well, it depends on the use of the method. You may need to do a quick linearity comparison between the HPLC and the UPLC. If you're injecting the same amount you could easily saturate your detector sooner. Conversely, if your conc'n is very low then you may not see it with the shorter path length detection cell (and lower injection volume).

Therefore, as mentioned earlier, you may need to re-evaluate your injection volume. Also, if you have closely eluting peaks in your UPLC system, the resolution will suffer in the HPLC (since the path length and the tubing diameter most probably is not the same).

Doing a quantitative method transfer will require a preliminary bridging study (very short) to verify that what you're getting on your UPLC is the same as what you're seeing on your HPLC.

[Lime]

Indium ,

You may use HPLC columns ( only C18 ones ) in UPLC systems. however you cannot use a UPLC column ( which has smaller particles and narrower diameter ) in HPLC system because of high back pressure.

hope helps ..

Lime

[Kostas Petritis]

Lime,

You are not limited only in C18 columns with the UPLC. Any stationary phase that is mechanically resistant in high back pressures can be used (i.e. silica based ion exchange etc).

And the internal diameter of the column doesn't have to do anything with the back pressure as you always adjust the flow rate to the same linear velocity.

I guess that about 3 cm UPLC columns could be used with conventional HPLC system.