LC-MS: excessive contamination ion source
Posted: Sat Dec 01, 2007 12:51 pm
Hi all,
I have been trying to quantify a mixture of antibiotics in human plasma using UPLC-MS/MS. During method development, I am always left with excessive (somewhat crystalline) contamination of the ESI-chamber and cone. I have tried rinsing all tubing with IPA/MeOH/AcN/H2O and preparing fresh mobile phase before each run. I have avoided the use of non-volatile buffers and have diluted plasma to minimize the level of endogenous salts. What do you think: is it my sample prep? Should I go over to SPE or should this protein prec have worked? Are there any other causes I haven't thought of?
Sample prep:
Tenfold dilution of plasma with aqueous 0.1% FA in water, followed by precipitation with 10% TCA, containing IS (1 part dil plasma to 1 part 10%TCA). after that left for 20 min at 4C, followed by 5 min centrifugation at 14000rpm and transfer of supernatant to autosamplervial.
I've tried different protein precipitation methods, but TCA gives the best chromatograms; a higher conc is not an option since ion suppression starts to kick in.
LC: BEH-column with in-line filterunit (0.2um), flow=0.5mL/min, column temp=50C
Gradient: A = 0.1% FA in water, B = MeOH, 10% to 90% B over a period of appr. 2.5 min. Injection volume = 30 ul
Thanks in advance
Maurice
I have been trying to quantify a mixture of antibiotics in human plasma using UPLC-MS/MS. During method development, I am always left with excessive (somewhat crystalline) contamination of the ESI-chamber and cone. I have tried rinsing all tubing with IPA/MeOH/AcN/H2O and preparing fresh mobile phase before each run. I have avoided the use of non-volatile buffers and have diluted plasma to minimize the level of endogenous salts. What do you think: is it my sample prep? Should I go over to SPE or should this protein prec have worked? Are there any other causes I haven't thought of?
Sample prep:
Tenfold dilution of plasma with aqueous 0.1% FA in water, followed by precipitation with 10% TCA, containing IS (1 part dil plasma to 1 part 10%TCA). after that left for 20 min at 4C, followed by 5 min centrifugation at 14000rpm and transfer of supernatant to autosamplervial.
I've tried different protein precipitation methods, but TCA gives the best chromatograms; a higher conc is not an option since ion suppression starts to kick in.
LC: BEH-column with in-line filterunit (0.2um), flow=0.5mL/min, column temp=50C
Gradient: A = 0.1% FA in water, B = MeOH, 10% to 90% B over a period of appr. 2.5 min. Injection volume = 30 ul
Thanks in advance
Maurice