Peak Coalescence Research Articles

Peak coalescence phenomena in enantioselective chromatography

Peak coalescence phenomena in enantioselective chromatography

Peak coalescence phenomena in enantioselective chromatography

Peak coalescence phenomena in enantioselective chromatography

Minimizing Peak Coalescence: High-Resolution Separation of Isotope Peaks in Partially Deamidated Peptides by Matrix-Assisted Laser Desorption/Ionization Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

Resolution of greater than 100 000 is routinely achieved by MALDI-FT-ICR, based on measured peak widths. However, the ability to separate peaks that require this resolution is difficult to obtain in practice due to peak coalescence, a result of coupling of the cyclotron motion of ions with similar frequencies. This phenomenon is accentuated for high space charge, high trapping plate voltages, and high mass. Very low trapping plate voltages with properly chosen transient measurement times are shown here to yield ultrahigh-resolution separation of closely spaced peaks in peptide mixtures. Measurements of the isotope peaks for partially deamidated preparations of substance P or partially reduced/partially deamidated Ala-Gly-[Arg]8-vasopressin show as many as six isotope peaks at one nominal mass. In one example, the 13C isotope peak was separated from the 15N isotope, a separation that required a resolution in excess of 180 000. Measurements were made with an external source MALDI-FT-ICR mass spectrometer with a 4.7 T magnet. These data suggest the need for high-resolution measurements for the determination of exact masses for peptide mixtures.

Advantages of High Magnetic Field for Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

It is well-known that mass resolving power in Fourier transform ion cyclotron resonance mass spectrometry (FTICR MS) can increase linearly with increasing applied magnetic field induction, B. Here, we show that eight other FTICR primary performance parameters theoretically also increase linearly (quadrupolar axialization efficiency, data acquisition speed), quadratically (upper mass limit, maximum ion kinetic energy, maximum number of trapped ions, maximum ion trapping duration, two-dimensional FTICR mass resolving power), or inverse-quadratically (peak coalescence tendency) with increasing B. The origin of (and conditions for) the magnetic field dependence of each of these parameters are presented and discussed. These fundamental advantages lead to corollary improvement in other FTICR performance parameters: e.g. signal-to-noise ratio, dynamic range, mass accuracy, ion remeasurement efficiency and mass selectivity for MS/MS. Finally, we note that these various advantages may be exploited in combination, so as to produce even higher enhancement in a particular parameter: e.g. signal-to-noise ratio can improve by more than a factor of B2 if mass resolving power is fixed at the same value as at lower magnetic field.

Characterization of supported-palladium catalysts by deuterium NMR spectroscopy

Alumina-supported Pd, Rh and Pd–Rh bimetallic alloys have been prepared by the method of incipient wetness. Sorption of deuterium by these samples was investigated by means of 2H NMR spectroscopy and uptake measurements at room temperature. Deuterium chemisorbed on palladium and rhodium atoms was characterized by 2H NMR peaks with chemical shifts of –12 and –170 ppm, respectively, from monometallic samples under a deuterium pressure of 1 Torr or less. These two characteristic peaks remained for the bimetallic samples and their relative intensities unambiguously revealed the surface composition of the bimetallic crystallites. Deuterium atoms were absorbed or weakly chemisorbed at greater pressures of deuterium. The weakly sorbed deuterium atoms exchanged with the strongly chemisorbed deuterium atoms, indicated by the coalescence of the 2H NMR characteristic peaks of the bimetallic samples. The position of the coalesced peak depended on the composition of the alloy crystallites.

Chiral Separation of Lorazepam on Ovomucoid-Bonded Columns: Peak Coalescence Due to Racemization

Abstract Racemic lorazepam (LZ) was separated on an ovomucoid (OVM)-bonded column by changing the column temperature. Chromatographic peak coalescence, appearing as a plateau between the resolved peaks, was observed above column temperatures of 15°C. Each LZ enantiomer was isolated on another chiral column using a non-aqueous eluent. Racemization of each LZ enantiomer in two different solutions and peak coalescence on these chiral columns were investigated. These results revealed that the peak coalescence should be due to racemization of LZ on the OVM-bonded column. By reducing the column temperature to 7°C, the enantiomeric composition of LZ before chromatography could be determined on the OVM-bonded column.

Hindered migration of amino acids toward their isoelectric positions in gel electrofocusing, and facilitation of the migration at high ionic strength

Amino acids with a largep I -p K p difference are known to be poor carrier ampholytes in electrofocusing, exhibiting isoelectric zones with poor conductivity across as many as 4 pH units. Accordingly, radioactive amino acids of this type, e.g., glycine, are found to be distributed over the entire pH gradient formed by Ampholine in electrofocusing gels, while radioactive amino acids like histidine or glutamic acid with small p I - p K p differences form single peaks at or near their p I's. When poor carrier ampholyte amino acids are subjected to gel electrofocusing in 0.1 m KCl, their distribution sharpens into single peaks, at or near the p I, indistinguishable from those of the good carrier ampholyte amino acids. At an intermediate stage of peak coalescence of the original broad distributions of poor carrier ampholyte amino acids, in 0.01 m KCl, acidic and basic peaks of amino acid can be observed, possibly analogous to acidie and basic distributions previously observed with labeled Ampholine. The rate of peak coalescence of anionic amino acids seems higher than that of the cationic species. The mechanism by which high ionic strength facilitates the condensation of poor carrier ampholyte amino acids at their p I remains unknown. Possibly, the current within zones of poor carrier ampholyte amino acids is insufficient, or poor carrier ampholyte amino acids are not sufficiently charged, to allow for electrophoretic migration of the bulk of loaded amino acid to its isoelectric position, unless the current density is increased by electrofocusing at high ionic strength. Alternatively, 0.1 m KCl may interfere with electrovalent interactions between amino acids and isoelectric carrier ampholyte zones, analogous to the action of urea in preventing the interaction between polyanions and carrier ampholytes.

Formyl-, aroyl, and acyl-alkylidenetriphenylphosphoranes: conformational analysis by proton nuclear magnetic resonance spectroscopy

The conformations of eleven oxoalkylidenetriphenylphosphoranes have been assigned from their 1H n.m.r. spectra. Mixtures of (Z)cisoid and (E)transoid conformers were detected in four compounds. The Z-geometry of β-oxomethylenetriphenylphosphoranes is favoured by the presence of β-substituents and the E-geometry is favoured by the presence of α-substituents. The conformational preferences are rationalised in terms of steric and electrostatic interactions. The absence of peak coalescence in the p.m.r. spectra up to 150° shows that interconversion of the conformers is very slow relative to the n.m.r. time scale.