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RT advancement in sugars RP HPLC
Posted: Mon May 21, 2012 1:16 pm
by dineshgbabu
Hi
I was performing sugar analysis on our Agilent 1260 HPLC. Today I got the sugar peaks with advanced RT. What might be the reason and how can I overcome it? I am worried....
Waiting for a reply....
Dinesh babu.
Re: RT advancement in sugars RP HPLC
Posted: Mon May 21, 2012 5:02 pm
by tom jupille
What might be the reason and how can I overcome it?
We are not mind readers!
If you told us what you were doing (what column packing, what dimensions and flow rate, what mobile phase, what sugars, etc.) someone might actually be able to help.
Re: RT advancement in sugars RP HPLC
Posted: Tue May 22, 2012 11:34 am
by dineshgbabu
Yes, sorry.
I was using C18 column as provided by the company for carbohydrate analysis. The flow rate is 1.4ml/min and the mobile phase was 75% ACN in water.
Is there any other details that could be helpful? Please let me know.
Dinesh
Re: RT advancement in sugars RP HPLC
Posted: Tue May 22, 2012 1:46 pm
by bisnettrj2
Do you pre-mix your mobile phase and run isocratically?
Re: RT advancement in sugars RP HPLC
Posted: Tue May 22, 2012 2:07 pm
by carls
With these conditions (75% ACN) this must be an amino column?
What is your sample matrix?
Re: RT advancement in sugars RP HPLC
Posted: Tue Jun 19, 2012 1:41 pm
by AUREND
Yes, sorry.
I was using C18 column as provided by the company for carbohydrate analysis. The flow rate is 1.4ml/min and the mobile phase was 75% ACN in water.
Is there any other details that could be helpful? Please let me know.
Dinesh
Hi
I usually work at a flow rat of 1.0 ml/min. Maybe it can help you.
Byes
Re: RT advancement in sugars RP HPLC
Posted: Tue Jun 19, 2012 2:57 pm
by Vlad Orlovsky
Sugars are not retained by reversed-phase mechanism. You might get some minimal retention at 90-95% ACN but this will be attributed to polarity of residual silanols (HILIC). Flow rate does not change void volume or k' prime. You need to use HILIC column (bare silica, aminopropyl, etc.). Also you need to make sure that you have appropriate detection technique and that what ever you see in chromatogram is indeed your sugar or not some artifact of detection
Re: RT advancement in sugars RP HPLC
Posted: Tue Jul 03, 2012 7:41 am
by thekartis
i do face the same problem my MP is 79 ACN + 21 water with 2 ml flow rate, RI detector same Agilent 1260 HPLC instrument...
Column is Waters 10 micron amino column with 4.6 X 250 mm...
was not able to sort the problem
Re: RT advancement in sugars RP HPLC
Posted: Tue Jul 03, 2012 2:48 pm
by Gaetan Glauser
Re: RT advancement in sugars RP HPLC
Posted: Thu Jul 05, 2012 5:24 am
by thekartis
thanks a lot for pointing out..
Actually i have raised a complaint to waters when i brought this column.
since our customer was using the same column for more than 4 years their RT was at 2.8 min, whereas in my brand new column i was getting RT at 12-13 min...
so the waters have replaced the column with the brand new one, even in that i was getting RT at 12-13 min..
since the results were fine...the issue has been closed..
now u r link has cleared the mystery...
thanks a lot once again