Combined Multiharmonic Frequency Analysis for Improved Dynamic Energy Measurements and Accuracy in Charge Detection Mass Spectrometry

Abstract

The ability to determine ion energies in electrostatic ion-trap-based charge detection mass spectrometry (CDMS) experiments is important for the accurate measurement of individual ion m/z, charge, and mass. Dynamic energy measurements throughout the time an ion is trapped take advantage of the relationship between ion energy and the harmonic amplitude ratio (HAR) composed from the fundamental and second harmonic amplitudes in the Fourier transform of the ion signal. This method eliminates the need for energy-filtering optics in CDMS and makes it possible to measure energy lost in collisions and changes in ion masses due to dissociation. However, the accuracy of the energy measurement depends on the signal-to-noise ratio (S/N) of the amplitudes used to determine the HAR. Here, a major improvement to this HAR-based dynamic energy measurement method is achieved using HARs composed of higher-order harmonics in addition to the fundamental and second harmonic to determine ion energies. This combined harmonic amplitude ratios for precision energy refinement (CHARPER) method is applied to the analysis of a 103 nm polystyrene nanoparticle ion (359.7 MDa, m/z = 308,300) and the energy resolution (3140) and effective mass resolution (730) achieved are the best yet demonstrated in electrostatic ion-trap-based CDMS. The CHARPER method applied to an ensemble of several thousand adeno-associated virus ion signals also results in higher mass resolution compared to the basic HAR method, making it possible to resolve additional features in the composite mass histogram.

Characterization of Mass, Diameter, Density, and Surface Properties of Colloidal Nanoparticles Enabled by Charge Detection Mass Spectrometry

“…Because mass is proportional to the cube of the diameter, the uncertainty in the diameter of each individual nanoparticle is only ∼0.3% (∼0.3 nm) in the CDMS analysis using a spherical model. When longer trapping periods are used, CDMS mass uncertainties as low as ∼0.1% can be achieved for nanoparticles of similar size and charge to those analyzed in this work, which translates to subangstrom (∼0.03 nm) diameter precision. In practice, this level of precision is not likely to be useful because it represents a length scale shorter than the shortest chemical bond.…”

“…The signal is amplified by a charge-sensitive preamplifier and subsequent bandpass filter stage before it is digitized at 1 MHz. Signals are analyzed using a program that performs unapodized short-time Fourier transforms (STFT), picks and fits the fundamental and multiple harmonic frequency peaks for each ion signal in each STFT segment and uses the resultant centroid frequencies and amplitudes to dynamically determine the mass, charge, and energy of each ion throughout the entire trapping period. ,, Multiple ions are routinely trapped and analyzed simultaneously; , frequency overlap events that resulted in ions being discarded made up <1% of all ion signals. The mass resolution of the CDMS used in this work depends on the charge states of the analytes and the length of the trapping period, which establishes the measurement time.…”

“…Charge detection mass spectrometry (CDMS) is an alternative method to characterize synthetic nanoparticles that directly measures masses and charges of individual ionized nanoparticles − instead of inferring or imaging the physical dimensions such as the diameter. Mass is a fundamental property of molecules, molecular complexes, and nanoparticles, and the extent of charging in the electrospray ionization (ESI) process typically used to generate ions for CDMS analyses can provide information about shape and physical properties of the molecular surface. − In CDMS, similar to microscopy techniques, individual nanoparticle measurements are compiled into histograms to obtain the mass and charge distribution of a sample.…”

“…CDMS is distinct from conventional mass spectrometry methodologies where only mass-to-charge ratios ( m / z ) are measured for large ensembles of analytes that are typically <1 MDa. Previous work in CDMS has probed the masses of large nanoparticles and polymers − ,− but the technique has only recently been applied directly to determining nanoparticle size distributions using recently developed instrumentation with high resolution and high throughput …”

Protein Complex Heterogeneity and Topology Revealed by Electron Capture Charge Reduction and Surface Induced Dissociation

“…Several approaches have been presented in recent years to overcome these issues. These include charge manipulation approaches and charge detection mass spectrometry (CDMS), in which the charge and mass-to-charge ratio are simultaneously measured. − CDMS overcomes the need for resolved charge states and provides insights into heterogeneous glycoproteins and large heterogeneous systems. − Charge manipulation approaches typically involve reducing the charge either in solution or the gas-phase. This moves the charge state distribution to lower charges where the spacing between charge states is larger and enables resolution of heterogeneous systems. , …”