Improved Spectral Quality in LC-MS Peptide Analysis Using Ultra Small Particle Chromatographic Packings

Library Number:
Part Number:
Thomas E. Wheat;Ziling Lu;Beth L. Gillece-Castro;Eric S. Grumbach;Paul R. Rainville;Uwe D. Neue;Jeffrey R. Mazzeo [Waters]
ASMS 2005
Content Type:
Content Subtype:
Phosphorylase Tryptic Digest, Enolase Tryptic Digest
Related Products:
Proteins are commonly identified and characterized using peptide mapping with mass spectral identification. Although Peptide Mass Fingerprinting of the unfractionated digest can be used for identification, the most complete structural information becomes available when the digest is fractionated. The most complete separation yields the best spectra. Recent advances in column chemistry and instrumentation permit the routine use of very small particle packings to enhance separations. The basis of this technique is described in the van Deemter equation and is explicitly related to diffusion. To apply ultra high resolution techniques to peptides, we have investigated operating conditions that could influence the application of UPLC to peptide mapping. We have compared the packing material used for UPLC for to those more commonly used for peptide LC/MS to measure effect on resolution and selectivity. Those parameters that influence diffusion-related chromatographic equilibria, specifically linear velocity and gradient slope, have been measured. The suitability of the material with different modifiers to increase electrospray signal is demonstrated. The possibility of using intrinsically higher resolution to reduce run is explored.

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