By Anonymous on Monday, February 23, 2004 - 08:13 am:
We use a lot of 2.0 mm x 50 mm columns for TIS (ESI) LC-MS (PE 200 pump and Sciex MS). It usually takes half an hour or longer to get stable RT, even if we use premixed MP (typically aq. Methanol or acetonitrile with 1-2 mM volatile buffer). Could anyone explain me why it takes so long? Of course, the system is primed with the MP before equilibration.
Thanks in advance.
By MG on Monday, February 23, 2004 - 11:20 am:
I don't think this is related to the column internal diameter, assuming you are running at a sufficient flow rate (0.2 mL/min or higher). I've run plenty of gradient methods with 4-6 min equilibration time for this column dimension, with the gradient starting at 5 to 10% organic. I did have this problem once while running a 10% methanol isocratic method. Someone else suggested that this was too much water for an ordinary C18 to handle continuously, and that I should switch to an "AQ" type column, but I never tried it as the retention times would eventually stabilize.
Could you provide more details on your conditions? Flow rate, exact mobile phase composition(s), gradient, column, etc.
By Anonymous on Tuesday, February 24, 2004 - 03:38 am:
Pump: PE 200 with online degasser
Autosampler: PE 200
Column: Zorbax SB AQ 50x2mm 3.5 um
MP: 45% methanol/55% 2mM ammonium acetate pH 4.5
MP is premixed, 0.35ml/min
The compounds are in neutral form under these conditions.
TurboIonSpray interface
RT shifts from 2.4 to 2.0 minutes within about 60 minutes.
In this lab, drifting RT happens with PE 200 LC-MS systems with 2.0 mm ID columns (namely flow rate of 0.2 to 0.35 ml/min, happens also to other methods with low flow rate). When we use 4.6 mmID columns, flow rate is usually about 1.0ml/min and we never see drifting RT.
Regards and thanks!
By Consumer Products Guy on Tuesday, February 24, 2004 - 06:36 am:
We use 2.1 mm i.d. columns for most of our pharmaceutical work, and retention times are perfect (Agilent HPLCs). We also use them for non-pharmaceutical and gradients, and they work fine.
By MG on Tuesday, February 24, 2004 - 12:23 pm:
If those are the "PE-200 Micro" pumps that were commonly sold with the PE-Sciex instruments a few years ago, I am using the same kind of pumps for most of my work! (and on 2mm columns too). Could it be some sort of temperature variation? Methanol/water at those ratios would be warmer than room temperature immediately after mixing. Sorry I can't be of more help.