Direct quantitation of peptide mixtures without standards using clusters formed by electrospray ionization mass spectrometry.
In electrospray ionization mass spectrometry, ion abundances depend on a number of different factors, including analyte surface activity, competition between analytes for charge, analyte concentration, as well as instrumental factors, including mass-dependent ion transmission and detection. Here, a novel method for obtaining quantitative information about solution-phase concentrations of peptide mixtures is described and demonstrated for five different peptide mixtures with relative concentrations ranging from 0.05% to 50%. In this method, the abundances of large clusters containing anywhere from 0 to 13 impurity molecules are measured and directly related to the relative solution-phase concentration of the peptides. For clusters containing approximately 15 or more peptides, the composition of the clusters approaches the statistical value indicating that these clusters are formed nonspecifically and that any differences in ion detection or ionization efficiency are negligible at these large cluster sizes. This method is accurate to within approximately 20% or better, even when the relative ion intensities of the protonated monomers can differ by over an order of magnitude compared to their solution-phase concentrations. Although less accurate than other quantitation methods that employ internal standards, this method does have the key advantages of speed, simplicity, and the ability to quantitate components in solution even when the identities of the components are unknown.
Department of Chemistry, University of California, Berkeley, California 94720-1460, USA.
Ryan D
RD
Leib
Tawnya G
TG
Flick
Evan R
ER
Williams
D001920:Bradykinin / Q000032:analysis / Q000737:chemistry; D004743:Enkephalin, Leucine / Q000032:analysis / Q000737:chemistry; D010455:Peptides / Q000032:analysis* / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D010455:Peptides; D004743:Enkephalin, Leucine; D001920:Bradykinin