Unknown Peak dilemma
Posted: Fri Aug 03, 2012 5:49 pm
While doing some development work, noticed various peaks within blank solution (junk?) appearing and several gaussian peaks at the same retention time as one of the analytes. These peaks are showing up during the organic ramp sequence from about 6 minutes to about 16 minutes. I can work around it with a large injection volume but it may affect peak purity later in validation.
Here are the chrom. parameters.
Mobile Phase A: 0.1% Perchloric Acid in H20
Mobile Phase B: 80:10:10 (MeCN:MeOH:MobilePhaseA)
System
Detector: PDA
Time Flow %A %B
1 0.01 1.20 98.0 2.0
2 2.00 1.20 98.0 2.0
3 15.00 1.20 0.0 100.0
4 18.00 1.20 0.0 100.0
5 19.00 1.20 98.0 2.0
6 30.00 1.20 98.0 2.0
7 31.00 0.10 80.0 20.0
Using a C18 w/ TMS endcapping Column 75mm X 4.6mm, 2.6 micron Core-shell Silica
Troubleshooting performed to narrow down my cause:
-tried different manufacturer perchloric and hplc grade water.
-cleaned glassware thoroughly (leaching)
-using different lines on hplc
-full loop purges/system washes overnight
-switched to brand new column, same result
-direct inject through detector flow cell (no peaks visible)
Some observations to note:
Peaks are largest after a long equilibration period, get smaller with each sequential injection. My assumption is that something is forming at the head of the column and is sticking to the silica beds, but what could it be and where is it coming from?
If I missed anything please ask and any insight would be much appreciated. Thank you.
Here are the chrom. parameters.
Mobile Phase A: 0.1% Perchloric Acid in H20
Mobile Phase B: 80:10:10 (MeCN:MeOH:MobilePhaseA)
System
Detector: PDA
Time Flow %A %B
1 0.01 1.20 98.0 2.0
2 2.00 1.20 98.0 2.0
3 15.00 1.20 0.0 100.0
4 18.00 1.20 0.0 100.0
5 19.00 1.20 98.0 2.0
6 30.00 1.20 98.0 2.0
7 31.00 0.10 80.0 20.0
Using a C18 w/ TMS endcapping Column 75mm X 4.6mm, 2.6 micron Core-shell Silica
Troubleshooting performed to narrow down my cause:
-tried different manufacturer perchloric and hplc grade water.
-cleaned glassware thoroughly (leaching)
-using different lines on hplc
-full loop purges/system washes overnight
-switched to brand new column, same result
-direct inject through detector flow cell (no peaks visible)
Some observations to note:
Peaks are largest after a long equilibration period, get smaller with each sequential injection. My assumption is that something is forming at the head of the column and is sticking to the silica beds, but what could it be and where is it coming from?
If I missed anything please ask and any insight would be much appreciated. Thank you.