Transient-mediated simulations of FTMS isotopic distributions and mass spectra to guide experiment design and data analysis

Fourier transform mass spectrometry (FTMS) applications require accurate analysis of extremely complex mixtures of species in a wide mass and charge state ranges. To optimize the related FTMS data analysis accuracy, parameters for data acquisition and the allied data processing should be selected rationally and their influence on the data analysis outcome is to be understood. Thus, to facilitate this selection process and to guide the experiment design and data processing workflows, we implemented the underlying algorithms in a software tool with a graphic user interface, FTMS Isotopic Simulator. This tool accurate-ly computes FTMS data via time-domain data (transient) simulations for user-defined molecular species of interest and FTMS instruments, including diverse Orbitrap FTMS models, followed by user-specified FT processing steps. Herein we describe implementation and benchmarking of this tool for analysis of a wide range of compounds, as well as compare simulated and experimentally-generated FTMS data. In particular, we discuss the use of this simulation tool for narrowband and broadband, low- and high-resolution analysis of small mol-ecules, peptides and proteins, up to the level of their isotopic fine structures. By demonstrating the allied FT processing artifacts, we raise awareness of a proper selection of FT processing parameters for modern applications of FTMS, including intact mass analysis of proteoforms and top-down proteomics. Over-all, the described transient-mediated approach to simulate FTMS data has proven useful for supporting contemporary FTMS applications. We also find its utility in fundamental FTMS studies and creating didactic materials for FTMS teaching.

Distinguishing analyte from noise components in mass spectra of complex samples: where to cut the noise?

Luca Fornelli, Anton Kozhinov, Yury Tsybin, Konstantin Zhurov

Fourier transform mass spectrometry (FTMS) enables comprehensive analysis of complex molecular mixtures. Given the broad intensity ranges of components in the mass spectra, it is imperative to accurately determine a noise threshold level above which peak assignments will be made. Conventionally, to find the threshold level, the "N sigma" approach or an equivalent rule is used. However, the "N sigma" approach cannot be applied to mass spectra stored with partially removed noise (reduced-profile mode). It is also not directly applicable to mass spectra acquired in the absorption mode with removed negative spectral amplitudes. Moreover, N value selection is normally made based on a rule of thumb, meaning that the calculated threshold level may be biased. Here, we present a noise thresholding method which addresses these limitations for analysis of mass spectra of complex molecular mixtures. The introduced data-dependent thresholding method involves analysis of the distribution of logarithmic intensity of all peaks, including noise and analyte, for a given mass spectrum. Selected method applications include FTMS analysis of crude oil fractions as well as tandem MS analysis of intact proteins.

Amer Chemical Soc2014

Least-Squares Fitting of Time-Domain Signals for Fourier Transform Mass Spectrometry

Tagir Aushev, Anton Kozhinov, Yury Tsybin

To advance Fourier transform mass spectrometry (FTMS)-based molecular structure analysis, corresponding development of the FTMS signal processing methods and instrumentation is required. Here, we demonstrate utility of a least-squares fitting (LSF) method for analysis of FTMS time-domain (transient) signals. We evaluate the LSF method in the analysis of single- and multiple-component experimental and simulated ion cyclotron resonance (ICR) and Orbitrap FTMS transient signals. Overall, the LSF method allows one to estimate the analytical limits of the conventional instrumentation and signal processing methods in FTMS. Particularly, LSF provides accurate information on initial phases of sinusoidal components in a given transient. For instance, the phase distribution obtained for a statistical set of experimental transients reveals the effect of the first data-point problem in FT-ICR MS. Additionally, LSF might be useful to improve the implementation of the absorption-mode FT spectral representation for FTMS applications. Finally, LSF can find utility in characterization and development of filter-diagonalization method (FDM) MS.

Springer2014

Structural Analysis of Complex Molecular Systems by High-Resolution and Tandem Mass Spectrometry

Yury Tsybin

In this chapter, we describe the advances in high-resolution and tandem mass spectrometry applied for structural analysis of complex molecular systems. Focus is made on Fourier transform mass spectrometers (FTMS) because of their ability to routinely provide resolving power (resolution), which is above 100 000, and correspondingly high mass accuracy, which is higher than 2 ppm. The advantages of using these analytical capabilities have been demonstrated for analysis of extremely complex mixtures of molecules of diverse nature and size. What are the recent and envisioned steps in FTMS method and technique development? What are the novel application areas that await these developments? We describe the state-of-the-art in ion cyclotron resonance (ICR) and Orbitrap FTMS and showcase the advantages of FTMS applications for petroleomics and intact protein mass measurements. We briefly overview the current methods of ion activation and dissociation as most widely employed in tandem mass spectrometry (MS/MS) for peptide and protein structure analysis. Finally, we discuss the analytical limitations of the state-of-the-art FTMS and offer a brief introduction to some of the possible solutions to these limitations. Particularly, we describe the achieved and envisioned benefits of advanced signal processing methods aimed specifically at FTMS improvement.

Wiley-VCH Verlag GmbH & Col. KGaA2014