Hello Research Gate Community,
I'm currently trying to establish methods for detecting oligonucleotides on our LC/MS system. As eluent I picked Water and Methanol each buffered with HFIP/TEA (1%/0.1%). Irregardless of my flow (0.5 ml/min - 0.25 ml/min) and ionisation polarity the MS detector is saturated with signals from the buffering additives (HFIP in negative mode and TEA in positive mode) even in literature I've read that way higher concentrations of the used additives are common.
Is there a way to reduce the ionisation of the buffering compounds? Is maybe the ionisation unit in the used spectrometer too sensitive? If yes how can I adjust it?
Unfortunately we have not too much experience when it comes to doing LC/MS with buffered eluents.
Best regards Ferenc
Hi Ferenc,
maybe this link will be helpful to you:
https://www.researchgate.net/post/HPLC_column_for_ssRNA_1000-4000bp_IP-RP
Regards
Markus
Hi all,
in the meantime I've found a solution myself:
since small molecules like buffer additives will saturate your detecter if your machine's ionisation unit is powerfull enough it is mandatory to adjust the voltages in the ionisation unit (usually over the MS interface) in favor of big molecules. Since we're having a maxis impact from Bruker this information was provided by bruker itsself.
Further, adjust the detecting range of the detector to at least 2x of the expected mass of the buffer substances. This advice was given to me by the head of the institutes MS platform. In the end i adjusted to 500 m/z as lower threshhold.
The result was very low background noise and clear signals of the applied oligos which showed ionization states from 3- to 12-.
Another tip if your device is not exclusively used for buffered LC/MS: If flushing back to an unbuffered or even weakly buffered eluent (like Water/ACN/HCOOH in our case) make sure to completely remove the buffer from all LC parts by flushing or exchanging plastic parts since the buffer might give detector saturation again when using methods with lower mass threshhold (e.g. 100 compared to 500 for oligos) again.
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