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Improving the resolution of neuropeptides in rat brain with on‐line HILIC‐RP compared to on‐line SCX‐RP
Author(s) -
Mihailova Albena,
Malerød Helle,
Wilson Steven Ray,
Karaszewski Bartosz,
Hauser Roman,
Lundanes Elsa,
Greibrokk Tyge
Publication year - 2008
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.200700257
Subject(s) - hydrophilic interaction chromatography , chemistry , chromatography , elution , two dimensional chromatography , mass spectrometry , resolution (logic) , high performance liquid chromatography , artificial intelligence , computer science
Our two already established on‐line 2‐D LC systems, a strong cation exchange‐RP chromatography (SCX‐RP) system and a hydrophilic interaction LC (HILIC)‐RP 2‐D LC system, were compared to explore which system is best suited for our further studies of differences in cerebral neuropeptide expression as a function of hypoxia‐caused stress. The same mass spectrometer and database search parameters were applied in both systems. In total, 19 first dimension fractions were collected with the novel on‐line HILIC‐RP system, including a Hypercarb SPE column that was applied to trap the compounds not retained on a Kromasil C 18 enrichment column. In contrast, six fractions were collected in the SCX‐RP method, due to practical limitations of this traditional on‐line 2‐D LC system. With the on‐line HILIC‐RP system three times more peaks were detected. It was observed that most of the compounds eluted in the first two fractions in the SCX‐RP method, while in the 2‐D HILIC‐RP method there seemed to be no correlation between peaks detected and fraction number. Thus, from this systematic study it seems that on‐line HILIC‐RP chromatography is the method of choice for comparative “peptidomics” of cerebral neuropeptides in future studies.