Quantitative peptide bioanalysis using column-switching nano liquid chromatography/mass spectrometry

✍ Scribed by Oosterkamp, A. J.; Gelpí, E.; Abian, J.

• Publisher: John Wiley and Sons
• Year: 1998
• Tongue: English
• Weight: 298 KB
• Volume: 33
• Category: Article
• ISSN: 1076-5174
• DOI: 10.1002/(sici)1096-9888(1998100)33:10<976::aid-jms710>3.0.co;2-7

✦ Synopsis

Many endogenous peptides are circulating in bodily Ñuids at the low pmol l-1 range, placing high demands on the bioanalytical procedure. In order to analyze these minute concentrations in complex matrices, a miniaturized liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) bioanalysis method was developed using custom-made nanoLC columns (75 lm i.d.) and a micro-electrospray interface (lESI). To be able to analyze large sample volumes in order to cope with low biological analyte concentrations, the nanoLC/ESI-MS method was coupled to an on-line preconcentration (PC) system based on a strong anion-exchange material. This method was used to analyze endothelin peptides (ETs) in complex matrices, which are potent vasoconstrictors of M r Á 2500 Da. The ET isoforms could be simultaneously analyzed with detection limits down to 30 pmol l-1 in cell supernatants (1.5 fmol on column). The method was linear from 50 to 2000 pmol l-1 with correlation coefficients of 0.99 for two of the three endothelin isoforms. Several other parameters, such as matrix e †ects and recovery, were also investigated.