How long to wait between gradient injections?

<p><span>When I am working with a gradient instrument method, how long, if at all, should I delay injecting the next sample? Should I wait until the pressure difference is within a certain psi value?</span></p><p></p><p><span>I know with the gradient there will be a gradual change in pressure during the run. My instrument method does have the gradient returning to the original composition before the injection ends. I'm concerned the pressure or mobile phase in the system may not be re-equilibrated enough before the next injection and may affect the chromatography. There may be a several hundred psi difference in the pressure ripple as the next injection starts. Do I need to be concerned about this?</span></p>


  • lizh

    Thank you for this question.

    Do not wait. Ensure you have accounted for the 3 system + 5 column volumes and if your compounds are known, ensure that you have evaluated the minimum hold at final gradient conditions necessary to stop your peaks moving around. Otherwise allow the system to manage the injection process and you will not have to extend the cycle time. The ACQUITY UPLC system designed to do manage just that and all injections get equal pump strokes from start to finish.

    The system has to manage injections going from atmospheric pressure to system pressure and back again without impact on the column - which is the most sensitive to pressure change, and will loose effiecency.The system also removes the injection pulse associated with un-pressurized pressure loop brought online. You may see the ripple, but the reproducible peak RSD's and column lifetimesshould demonstrate the process is effective.

    Additionally, the Sample Manager and Binary Solvent Manager are synchronized. Actually all the modules are communicating. The BSM is aware of when the injector will inject and will catch up to it, and make an injection off a full stroke. The system works to maximize RSD's and minimize cycletime. For a high pressure system to deliver smooth flow and react to gradient viscocity changes requires fast compressions which creates heat, the solvent sheds this heat and contracts and a pressure drops. The address this the BSM uses active pressure control during Pump “Transfer” to compensates for solvent contraction after compression is done.We use the active transfer to eliminates uncertainty of which pump (A vs. B) may overlay injection with a pressure-controlled Transfer

    Hope that this reassures you,