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Weird peak shape all of a sudden...
Posted: Tue Oct 15, 2024 5:20 pm
by Pear
Hi,
Looking for some help with a strange peak shape that's appeared on one of our HPLC's.
The peak looks like it's splitting in 2 at the top / shouldering? So far I have:
- Changed the UV bulb
- Changed all solvents
- Purged the system
- Changed the column
- Remade the samples using new standards from a completely different lot number
- Remade the samples using standards from a different supplier
And yet this odd shape remains. Anybody have any ideas?
For reference, this is what a previous peak of the same concentration looks like:
Appreciate any help.
EDIT:
I have just run 5 injections and 4 of them were shouldered and 1 was well resolved. The system was serviced 2 weeks ago by the manufacturer and this is the first time using it since. Could it possibly be the new pump seals aren't seated properly and letting air bubbles form?
Thanks
Re: Weird peak shape all of a sudden...
Posted: Wed Oct 16, 2024 12:54 pm
by Consumer Products Guy
If this happened to me, I'd "for fun" try halving the injection volume, just to see if peak shape got better, worse, or stayed the same.
Re: Weird peak shape all of a sudden...
Posted: Wed Oct 16, 2024 1:54 pm
by Multidimensional
Could be co-elution, poor chromatography method (no retention), too low a sample rate, wrong injection solvent etc. Why speculate and make it hard? The easy way involves you providing some basic info.
Please provide:
Column Type (Name and type),
Column Dimensions and particle size,
Flow Rate,
Detection Settings,
Sampling Rate,
injection solvent,
Injection volume,
Mobile phase composition (and gradient conditions, if applicable):
Re: Weird peak shape all of a sudden...
Posted: Wed Oct 16, 2024 2:01 pm
by TylerSmith123
Hi Pear,
This is anecdotal somewhat, but I recently had to change my seals because I was seeing issues that are similar to yours. My issues were slightly more pronounced, but I run Prep LC and most of my peaks are so large that I cannot see splitting occurring instantaneously. However, my issue was the seals wearing and after replacement everything was much smoother. If this issue manifested following PM, I would suspect that perhaps the seals are seated improperly. Even in your baseline, it seems that there is a rhythmic disturbance which may be air getting in. If you're comfortable doing the work you could replace the seal with the old one, or another you have handy. But I'm not certain what system you're running, if it's a binary pump, then you will likely wish to isolate the pump with the issue (assuming these are new seals). No need to give yourself extra work. Check those seals and then get back to us.
TS
Re: Weird peak shape all of a sudden...
Posted: Wed Oct 16, 2024 2:14 pm
by bcd_GCLCMSMS
If this appeared suddenly after running for a long period of stable chromatography, it may also be seals or a bad check valve.
Re: Weird peak shape all of a sudden...
Posted: Wed Oct 16, 2024 3:20 pm
by Pear
If this appeared suddenly after running for a long period of stable chromatography, it may also be seals or a bad check valve.
Hi Pear,
This is anecdotal somewhat, but I recently had to change my seals because I was seeing issues that are similar to yours. My issues were slightly more pronounced, but I run Prep LC and most of my peaks are so large that I cannot see splitting occurring instantaneously. However, my issue was the seals wearing and after replacement everything was much smoother. If this issue manifested following PM, I would suspect that perhaps the seals are seated improperly. Even in your baseline, it seems that there is a rhythmic disturbance which may be air getting in. If you're comfortable doing the work you could replace the seal with the old one, or another you have handy. But I'm not certain what system you're running, if it's a binary pump, then you will likely wish to isolate the pump with the issue (assuming these are new seals). No need to give yourself extra work. Check those seals and then get back to us.
TS
Both of you were absolutely bang on.
New seals removed, reseated and connected back up. Peaks are sharp again.
Thank you all.