A selective report on topics at Pittcon 2015

Groskreutz, Stephen R.; Horner, Anthony R.; Varner, Erika L.; Yin, Bing; Michael, Adrian C.; Weber, Stephen G.

doi:10.1016/j.trac.2015.04.005

Cited by 1 publication

(1 citation statement)

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“…Formic acid is typically used to replace TFA, but it gives lower chromatographic resolution due to its higher pKa [20]. A compromise is difluoroacetic acid (DFA), which has a lower pKa than formic acid and less surface activity than TFA [21]. To gauge how much chromatographic efficiency is lost in using modifiers other than TFA, RPLC with UV detection was used for the histone separation.…”

Section: Resultsmentioning

confidence: 99%

Chromatographic efficiency and selectivity in top-down proteomics of histones

Zhou¹,

Zhang²,

Fornelli

et al. 2017

Journal of Chromatography B

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Histones are involved in epigenetic control of a wide variety of cellular processes through their multiple post-translational modifications. Their strongly cationic nature makes them challenging to separate with reversed-phase liquid chromatography coupled to mass spectrometry (RPLC-MS), where trifluoroacetic acid is avoided due to adduct formation. Columns with higher resolution are needed. In this work, RPLC-MS is performed on a histone sample using difluoroacetic acid and a 20-minute gradient. Columns with C18 surfaces are compared for two different types of particle morphologies: 1) fully porous particles of 5 μm in diameter, 2) superficially porous particles of 3 μm in diameter with a shell of 0.2 μm. The resolution for the histone separation is better for the latter column, but only when the modifier is trifluoroacetic acid, which is used with UV absorbance detection. When difluoroacetic acid is used for LCMS, the peaks broaden enough to erase the advantage in efficiency for the superficially porous particles. The fully porous and superficially porous cases show similar performance in RPLC-MS, with slightly higher resolution for the fully porous particles. The expected advantage of the shorter diffusion distances for the superficially porous particles is shown to be outweighed by the lower selectivity of its bonded phase.

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Section: Resultsmentioning

confidence: 99%