Chromatography Forum

We are running into issues between these 2 systems, reading through other posts it seems as though the only differences between the 1100 series and 1200 series are that the 1200 has a higher rating for pressure and compressibility. With the 1100 our chromatograms look just as they are supposed to. Our issue that we are seeing is with the 1200 the elution time is about 20% later than with the 1100 and much broater peaks. Same column was used, same flow cell, same flow rate, gradient, etc.

Sidenote: a 10uL injection on the 1200 gives us a perfect defined chromatogram, but at 20uL we are seeing a lot of fronting, suggesting sample overloading? Our method is validated for 20uL which worked fine with out 1100's but we are doing away with the 1100's. Any suggestions as to what the differences/issues we are seeing between these systems is greatly appreciated!

You're assuming that both systems are functioning correctly, which may not be true. If it were my problem, I'd start by re-running PQ on both systems to verify gradient delay volumes (which can affect retention times). Something as simple as plumbing an injection valve backwards (pump and column lines switched) can make a big difference. Also, if your diluent is stronger than your mobile phase, you can sometimes see weird instrument-to-instrument variations due to different mixing volumes.

Agree with Tom's post. Are your lengths and inner diameters of your tubing exactly the same? This is especially important for gradient elutions.

Have you verified the flow rate, for example even with graduated cylinder over 20 minutes.

20ul is a quite large sample size for most modern-type columns; if the peak shape improves significantly when reducing the injection volume when injecting from a strong solvent into lower-organic mobile phase, then you have the solvent effect of the injected solvent. I assume that your column is 4.6mm i.d. and not a more-modern smaller i.d./smaller particle size one.

Our typical injection size is 5ul and we typically use 3mm i.d. columns for our in-house cGMP-validated methods.