Scale UPLC method to 50-80ul flows

fmichopo
fmichopo
in Instrumentation and Mass Spectrometry
<p><span>Hi all</span></p><p></p><p><span>I do have a UPLC method on BEH column (2.1*100) at 0.4ml/min flow. I do want to scale this method to 50 or 80uL per minute flows. </span></p><p></p><p><span>Does anyone know whether the 1*100 BEH column would hepl me to achieve this or shall i go to cappillary scale chromatography? </span></p><p></p><p><span>Can UPLC reliably deliver those flows? </span></p><p></p><p><span>Thanks in advance</span></p>

Comments

  • lizh
    lizh

    Hello:

    As a point of interest, I would like to why you want to scale to 1 mm. Is this for MS?

    The ACQUITY UPLC system is designed for 1 mm columns and their flows, and this example should scale to about 100 uL/min. However, in practice 1 mm columns need consideration as the volume of the column is so small (about 39 uL in this case, so much smaller than the system) so you need to make sure that your dispersion is limited. At maximum a 2 uL loop should be used and limit your tubing lengths and enaure that the smallest ID is used after the column.

    Off the top of my head - I can remember an application run at 47 uL/min, this was so the user could string lots of columns together to attain maximum resolution, so in practice it was a good test, low flow rate, gradient and highest posible back pressure.

    Best regards,

    Liz

  • fmichopo
    fmichopo

    Hi Liz

    The issue is that my method is developed for a UPLC-ICP system and as you know ICP cant tolerate high organic flow rates. So at the moment i split the flow 1/4 in order to analyse my samples without significant plasma upsetting.

    So I thought reducing the flow down to 50ul/min I could directly introduce the UPLC efluent to ICP, gain sensitivity and the same time maintain the chromatographic resolution. I know the void volume of the ICP is a real issue to take advantage of the high UPLC resolution but on the other hand the analysis time is significant shorter. This is the reason why we use UPLC technology. Dispersion is something that we cant really avoid due to interface chamber design.

    Can UPLC pump realiable deliver 50ul/min flow?

    Thanks a lot

    Filippos

  • atus
    atus

    Hi,

    I have upgraded my nanoUPLC to handle 1mm ID columns. The system is running at 45ul/min with BEH C18 1.0x150mm column for Peptide Mapping (4 hr gradient). Runs are reproducible with TUV detector monitoring at 214nm. I also ran BEH 1x150mm column on the regular UPLC without any modification with a PDA detector. The UV maps were reproducible. However, significant dispersion observed with extra tubing connection to the Mass Spec. Watch out for the back pressure limit of the UV flow cell with the tubing ID. Even 64um ID tubing has significant peak distortion. I am hoping to move the upgraded nanoUPLC as close to the ion source as possible to minimize the dispersion.

    Hope this help a bit.

    Albert

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