Using the UPLC as an HPLC

pat060157
pat060157
in Instrumentation and Mass Spectrometry
<p><span>What would be the pitfalls for using uplc (with no alterations) as a "regular" hplc with conventional columns. </span></p>

Comments

  • lizh
    lizh

    Hello:

    I am sure additional comments will come.

    If you compare one LC to another or an LC to a UPLC you can list the differences, amongst other factors, here are three that you must consider. Essentially, what do you do today when you transfer a method from a Shimadzu system to an Agilent system? What factors do you take into account to be successful? Here are three key considerations:

    (1) Allowable system backpressure limit differences

    (2) System volume differences

    (3) Column Heater/Temperature control differences

    One of the important item is to ensure that the backpressure limit in the pump instrument method editor is set to 6,000 psi, or the appropriate pressure setting to prevent over pressurization of the HPLC column.

    The other factor is to assess the ration of system volume to column volume. This must be preserved. The column volume will no change, but the system volume of the ACQUITY UPLC is far lower than a typical HPLC. It is likely when transferring an HPLC method developed on an HPLC column that a gradient hold will be required to delay the beginning of the gradient until the number of column volumes vs. system volumes has been preserved.

    When transferring a method between systems and column temperature coontrol is used the efficiency of the heating/cooling can vary and as is well know temperature will affect a separation. It may be necessary to lower or raise the temperature to account for this.

    The injection volume required may be higher than the configured loop in the ACQUITY and a loop change and possibly a syringe change may be required to reach the necessary HPLC injection volume range.

    For ACQUITY UPLC the existing ACQUITY detectors can be used for HPLC. It may be necessary to adjust the filter time constant and data acquisition rate can be reduced.

    Attached is a guide that can be helpful.

    Best regards,

  • BJ
    BJ

    Another thing to consider: it is a different injection system.

    Most of the time this won't cause any problem, but we have seen cases where 1 peak was giving 100%,

    while the recovery for another peak in the same chromatogram was only 80%.

    But I think Liz has covered most of the pitfalls.

    BJ

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