UPLC Pump Acceleration Destroys Conventional HPLC Columns

Lusby

<p><span>Hi All,</span></p><p></p><p><span><span>In recent times we have been converting our laboratory over to Acquity from Alliance instruments, making the assumption that any 'legacy' methods that used to be run on an Alliance could also be run on an Acquity. This is not turning out to be the case....</span></span></p><p></p><p><span>There is a feature in Alliance methods where the acceration of the pump can be set, i.e you can tell the pump how long to take to accelerate to 10 mL/min. This allows the pump to apply pressure on a column gradually, and hence not create voids in the column packing. Unfortunately the Acquity Methods do not have this feature. </span></p><p></p><p><span><span>When the flow is set on an Acquity, the pump accelerates to that flow rate so quickly that the resulting massive increase in pressure destroys many conventional HPLC columns through the formation of a void in the column (even if they do not reach the maximum pressure for that column). I realise that i could program the flowrate to increase slowly in my gradient table (provided the method uses a gradient), but this is not a good fix, and there is still a risk of an operator forgetting to set the flow rate in increments (like we used to do manually 10-20 years ago) and destroying a $1000 column in seconds.</span></span></p><p></p><p><span><span>Am i missing something?, is there a way to configure pump acceleration on Acquity instruments?.</span></span></p><p></p><p><span><span>regards,</span></span></p><p></p><p><span><span>Jason.</span><span> </span></span></p>

lizh

Jason:

I do apologize about your column and we will look into your suggestion further for the nect revision of the software - as it is a good one.

I hope that other users on the community would confirm if they would like to see this feature.

And on a more practical note, you can do a flow ramp using just two lines of the gradient table and utilizing the curves. Please ensure that the column pressure limit is set to a value that is lower and I will copy your email to others who have experience in running in HPLC mode and see if we can maje further suggestions.

Liz

Jukka

This feature would be nice.

ACQUITYEPM

Greetings Jason,

The original intent of the ACQUITY UPLC system was to support the, newly developed and tightly packed, small diameter particle columns. These columns are much less prone to voiding and collapsing. Further, the original product definition and scope of ACQUITY did not include separation techniques such as gel permeation or ion exchange. These types of columns, along with some traditional HPLC columns, have a higher tendancy to void under sudden pressure pulses. In contrast, the Alliance system had to accommodate all previous separation techniques and the flow ramping was included in the design with this in mind as Alliance was more evolutionary and less revolutionary in nature. Now, with the broad acceptance of ACQUITY (and UPLC) there is a growing desire to migrate all types of separations to the ACQUITY platform. With this comes a need for a less immediate initiation of flow to protect some of the less robust column types. We have, certainly, discussed this amongst ourselves (Waters) and your commentary may be the impetus needed for us to implement the feature.

In the meantime, you are not missing anything. Some type of flow profile should be instituted on your ACQUITY system to protect the, conventional, HPLC columns. This can either be performed manually or through some "automatic" means with the use of a startup instrument method.

Thanks for your observation and we will take it under serious consideration,

pcb

Lusby

Thanks for the reply.

I appreciate Waters consideration of this feature. I look forward to it being implemented

harringg

To clarify. If I was to take a 2.1x50, 3.5 um Symmetry column and run the same gradient profile on the UPLC as I'm running on the Alliance 2695, same flow rate (0.2 ml/min), it will in fact damage the column by the very nature of the current firmware in the UPLC and the pump? I'm not talking about increasing the flow rate to decrease run time on the column, but take an apples-to-apples method and run it on the UPLC. This will damage the column?

Thanks,

Grant

lizh

Absolutely, not, users routinely run HPLC columns on the UPLC, especially for method transfers. In fact, we also have customers running "fragile "protein SEC columns The maximum backpressure limit does have to be changed to reflect the column's limit, but that's it. The UPLC has a smaller dispersion and this can affect the separation as the gradient profile while set up identically will reach the column faster and will be less diluted by a significant factor ( 100 uL compated to 650 uL).

Best regards,Liz

harringg

Thanks for the clarification. The original post confused me, as I thought we could run the UPLC like a 2695. Realizing that it defeats the purpose of having a UPLC, but if we needed to for a project, we could.

You mention too "fragile" columns. We have an IC-Pak Anion HR 4.6x75 (WAT026765) that we would like to run on the UPLC/TQD, we are currently running it on the 2695/QTof. With the proper pressure limit settings, is there any reason that can't be run on the UPLC?

Thank you.

lizh

Absolutely not, just wash out the buffers afterwards!

Liz

harringg

Thanks for your help.

ACQUITYEPM

Greetings,

Yes, Liz is correct and I hope I did not mislead anyone with my previous comments. The initial issue was that, unlike Alliance, ACQUITY does not have any native flow ramp function. This could (and apparently can) be bothersome with traditional HPLC columns, especially in the case where the application flow rate is high (1 mL/min. and above). In that case, Alliance will take a few seconds to "ramp up" to the flow rate whereas ACQUITY will, immediately, initiate the full flow rate. This can create the phenomenon known as a "pneumatic (actually hydraulic) hammer" that transmits a shock wave against the head of the column. Under these circumstances it would be wise to, manually, step up through a couple of intermediate flow rates before attaining the final, desired, flow rate. Alternately, a flow gradient Instrument Method could be created in Empower and used as a Condition Column line at the start of a Sample Set. However, there is absolutely nothing about ACQUITY that precludes the use of traditional HPLC columns.

Regards,

pcb

massmats

Good morning,

I have seen people routinely doing a manual flow ramp on the UPLC, also when running UPLC columns. If they are running at flows generating pressures around 12 - 13000psi, sometimes it will reach above 15000 momentarily at startup. Therefore I agree that an automatic flow ramp would be a great feature in the next ICOP release, especially in the "System Start Up" sequence in the UPLC Console.

Mats

harringg

ACQUITY_EPM

Thanks for the clarificaiton and pointing that specific difference between the two systems. With our ICPak column even on the 2695 we have a gradient ramp to starting conditions and we will for sure use one on the UPLC.

This has been a valuable discussion (one to go in our care and use lab notebook) and thank you to everyone for your contributions.

Thanks,
Grant

lizh

I think that in fact we do need to add a flow ramp and I have noted this in our database as a customer request that we wil implement in next software version. We just missed the latest version, but we shall get this in the next.

Liz

ACQUITYEPM

Greetings Mats,

The phenomenon you speak of, under UPLC conditions, is a little different. The very small particle diameters (1.7 um) of the UPLC columns causes another effect. These particles present a, significant, amount of resistance to the solvent flowing through them. So much so that there is actual, and measurable, frictional work (as defined by physics) done within the column. This work manifests itself as heat. This adiabatic heating is the reason why the backpressure on a UPLC system will decrease or "sag" during the first few minutes of flow. The internal column heating will lower the viscosity of the solvent and reduce the pressure drop across the column. Once the internal temperature of the column comes to equilibrium, the pressure will stabilize. Of course, this also occurs within traditional 3.5 um to 5 um particle size columns, but the impact is quite small and, essentially, negligible. Externally heating the column does mitigate this situation, but the internal friction and heating always takes place. The higher the requested flow results in higher initial backpressure and causes a greater frictional heating effect. Whenever I'm planning on working above 12,000 psi, I always step up to the flow rate. Otherwise, as you say, the, internally, "cold" column may cause an overpressure.

Regards,

pcb