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- Posts: 6
- Joined: Tue Jul 12, 2022 12:02 pm
I looked for someone to have posted a similar topic but I couldn't find any using the search function.
My issue is as follows:
I am working on a method for amine analysis by HILIC-MS/MS. Two of my analytes are spermidine and spermine. These two analytes arequite tough to get nice peak shapes, so that i have adjusted my conditions accordingly. Unfortunately, even with those conditions I can see those compounds carrying over into the next runs. I have confirmed that it is those analytes in the following runs by retention time and ms/ms.
First off, conditions are as follows:
Eluent A: 90% ACN, 10% H2O, 100 mM ammonium formate, 4.5% formic acid, pH 2.5
Eluent B: 100% H2O, 100 mM ammonium formate, 4,5% formic acid
0->3 min: 100% A
3->7.5 min: ramp to 42% A
7.5->8 min: hold at 42% A
8->10 min: ramp to 100% A
10->12 min: hold at 100% A
Flow: 0.5 ml/min
columns: I have tested this method on both an Agilent Poroshell 120 HILIC-Z (10 cm) and a Thermo Fisher Accucore HILIC (10 cm). Both columns show the same carryover issues.
I also want to provide some further explanation of the issue. English is not my mothertongue, so please excuse any wording and logic errors.
After injecting a sample containing spermidine and spermine, I will detect a fraction (~10%) of the sample response in every further run (even water and blanks) I do. I first realised this was an issue already a few hundred runs into method development. By now I have carryover in every run, so that I always detect a small amount of spermidine and spermine. It isn't as much as I will inject with any sample containing the analytes but it is enough to change the quantitative results significantly.
I have already tried extensive troubleshooting by excluding certain parts of the LC for the same blanks. Thus, I can exclude the Eluent Pump, sample manager and MS as root cause. I am almost 100% certain, that the reason for carryover lies within the capillaries leading to and the column itself.
I have already tried cleaning those with varying cleaning solutions used as eluent:
pure water, water:IPA 50:50, ammonia pH10, ammonia pH11, acetic acid pH2.5.
None of them have significantly lowered the amount of carryover, even though i have done 10 runs with every cleaning solution.
I am very grateful for any input and ideas on how to deal with this issue.
Thank you in advance,
Dominik