Lipidome atlas of the adult human brain.

Lipids are the most abundant but poorly explored components of the human brain. Here, we present a lipidome map of the human brain comprising 75 regions, including 52 neocortical ones. The lipidome composition varies greatly among the brain regions, affecting 93% of the 419 analyzed lipids. These differences reflect the brain's structural characteristics, such as myelin content (345 lipids) and cell type composition (353 lipids), but also functional traits: functional connectivity (76 lipids) and information processing hierarchy (60 lipids). Combining lipid composition and mRNA expression data further enhances functional connectivity association. Biochemically, lipids linked with structural and functional brain features display distinct lipid class distribution, unsaturation extent, and prevalence of omega-3 and omega-6 fatty acid residues. We verified our conclusions by parallel analysis of three adult macaque brains, targeted analysis of 216 lipids, mass spectrometry imaging, and lipidome assessment of sorted murine neurons.

Development and Complex Application of Methods for the Identification of Mutations in the FAD3A and FAD3B Genes Resulting in the Reduced Content of Linolenic Acid in Flax Oil.

Flax is grown worldwide for seed and fiber production. Linseed varieties differ in their oil composition and are used in pharmaceutical, food, feed, and industrial production. The field of application primarily depends on the content of linolenic (LIN) and linoleic (LIO) fatty acids. Inactivating mutations in the FAD3A and FAD3B genes lead to a decrease in the LIN content and an increase in the LIO content. For the identification of the three most common low-LIN mutations in flax varieties (G-to-A in exon 1 of FAD3A substituting tryptophan with a stop codon, C-to-T in exon 5 of FAD3A leading to arginine to a stop codon substitution, and C-to-T in exon 2 of FAD3B resulting in histidine to tyrosine substitution), three approaches were proposed: (1) targeted deep sequencing, (2) high resolution melting (HRM) analysis, (3) cleaved amplified polymorphic sequences (CAPS) markers. They were tested on more than a thousand flax samples of various types and showed promising results. The proposed approaches can be used in marker-assisted selection to choose parent pairs for crosses, separate heterogeneous varieties into biotypes, and select genotypes with desired homozygous alleles of the FAD3A and FAD3B genes at the early stages of breeding for the effective development of varieties with a particular LIN and LIO content, as well as in basic studies of the molecular mechanisms of fatty acid synthesis in flax seeds to select genotypes adequate to the tasks.

Structure-Preserving and Perceptually Consistent Approach for Visualization of Mass Spectrometry Imaging Datasets.

Mass spectrometry imaging (MSI) has become an important tool for 2D profiling of biological tissues, allowing for the visualization of individual compound distributions in the sample. Based on this information, it is possible to investigate the molecular organization within any particular tissue and detect abnormal regions (such as tumor regions) and many other biologically relevant phenomena. However, the large number of compounds present in the spectra hinders the productive analysis of large MSI datasets when utilizing standard tools. The heterogeneity of samples makes exploratory visualization (a presentation of the general idea of the molecular and structural organization of the inspected tissues) challenging. Here, we explore the application of various dimensionality reduction techniques that have been used extensively in the visualization of hyperspectral images and the MSI data specifically, such as principal component analysis, independent component analysis, non-negative matrix factorization, t-distributed stochastic neighbor embedding, and uniform manifold approximation and projection. Further, we propose a new approach based on a combination of structure preserving visualization with nonlinear manifold embedding of normalized spectral data. This way, we aim to preserve as much spatially overlapping signals as possible while augmenting them with information on compositional (spectral) variation. The proposed approach can be used for exploratory visualization of MSI datasets without prior deep chemical or histological knowledge of the sample. Thus, different datasets can be visually compared employing the proposed method. The proposed approach allowed for the clear visualization of the molecular layer, granular layer, and white matter in chimpanzee and macaque cerebellum slices.

Genetic diversity of SAD and FAD genes responsible for the fatty acid composition in flax cultivars and lines.

BACKGROUND: Flax (Linum usitatissimum L.) is grown for fiber and seed in many countries. Flax cultivars differ in the oil composition and, depending on the ratio of fatty acids, are used in pharmaceutical, food, or paint industries. It is known that genes of SAD (stearoyl-ACP desaturase) and FAD (fatty acid desaturase) families play a key role in the synthesis of fatty acids, and some alleles of these genes are associated with a certain composition of flax oil. However, data on genetic polymorphism of these genes are still insufficient. RESULTS: On the basis of the collection of the Institute for Flax (Torzhok, Russia), we formed a representative set of 84 cultivars and lines reflecting the diversity of fatty acid composition of flax oil. An approach for the determination of full-length sequences of SAD1, SAD2, FAD2A, FAD2B, FAD3A, and FAD3B genes using the Illumina platform was developed and deep sequencing of the 6 genes in 84 flax samples was performed on MiSeq. The obtained high coverage (about 400x on average) enabled accurate assessment of polymorphisms in SAD1, SAD2, FAD2A, FAD2B, FAD3A, and FAD3B genes and evaluation of cultivar/line heterogeneity. The highest level of genetic diversity was observed for FAD3A and FAD3B genes - 91 and 62 polymorphisms respectively. Correlation analysis revealed associations between particular variants in SAD and FAD genes and predominantly those fatty acids whose conversion they catalyze: SAD - stearic and oleic acids, FAD2 - oleic and linoleic acids, FAD3 - linoleic and linolenic acids. All except one low-linolenic flax cultivars/lines contained both the substitution of tryptophan to stop codon in the FAD3A gene and histidine to tyrosine substitution in the FAD3B gene, while samples with only one of these polymorphisms had medium content of linolenic acid and cultivars/lines without them were high-linolenic. CONCLUSIONS: Genetic polymorphism of SAD and FAD genes was evaluated in the collection of flax cultivars and lines with diverse oil composition, and associations between particular polymorphisms and the ratio of fatty acids were revealed. The achieved results are the basis for the development of marker-assisted selection and DNA-based certification of flax cultivars.

Hydrogen/Deuterium Exchange Aiding Compound Identification for LC-MS and MALDI Imaging Lipidomics.

We present a novel approach for the increasing reliability of compound identification for LC-MS and MALDI imaging lipidomics. Our approach is based on the characterization of compounds not only by the elution time, accurate mass, and fragmentation spectra but also by the number of labile hydrogens that can be measured using the hydrogen/deuterium (H/D) exchange approach. The number of labile hydrogens (those from -OH and -NH groups) serves as an additional structural descriptor used when performing a database search. For LC-MS experiment, the H/D exchange was performed in the heating capillary of the modified electrospray ionization (ESI) source, while for MALDI imaging, the exchange was performed in the ion funnel at 10 Torr pressure. It was observed that such an approach allowed one to achieve a considerable degree of deuteration, enough to unambiguously distinguish between different classes of lipids. The proposed analytical approach may be successfully used for the identification not only of lipids but also of peptides and metabolites. A special software for the automatic filtration of molecules based on the number of functional groups was also developed.

Multi-electrode Harmonized Kingdon Traps.

Based on the analysis of the results of the study of various designs of multi-electrode harmonized Kingdon traps, we propose a new type of trap with two merged internal electrodes that has the ability to capture and accumulate ions formed inside. We also investigated the influence of inaccuracies in the manufacture of the electrodes on the field inside such trap. The four-electrode trap, which actually degenerates into a two-electrode device with traces of two other electrodes present at the ends of the internal electrodes (their splitting) has been found as the less sensitive to inaccuracies caused by manufacturing and cutting the ends of trap electrodes. We show that a mass spectrometer with a relatively high resolving power can be created on the basis of such a trap. The creation of the traps requires the manufacture of complex electrodes with demanded accuracy of their surfaces. This becomes possible with the advent of 3D printers. Graphical Abstract Multi electrode harmonize Kingdon trap with fused elctrode pares.

Influence of solvent on the yield and chemical composition of liquid products of hydrothermal liquefaction of Arthrospira platensis as revealed by Fourier transform ion cyclotron resonance mass spectrometry.

Yields of liquid products (bio-oil and residual organics in aqueous solution) of hydrothermal liquefaction (at 300℃ and 60 min) of own cultivated Arthrospira platensis using different solvents have been determined. Nonpolar hexane, moderately polar dichloromethane, and relatively more polar acetonitrile have been used. High-resolution mass spectrometry based on linear quadrupole ion trap and Fourier transform mass spectrometer (LTQ FT) has been used for liquid sample characterization. Concentration of bio-oil in a unit of solvent volume after solvent extraction took the following arrangement: acetonitrile > dichloromethane > hexane. Concentration of residual organics in a unit of aqueous solution after solvent extraction took the following arrangement: hexane >dichloromethane > acetonitrile. Yield of total organics was arranged as follows: hexane > dichloromethane > acetonitrile. Content of carbon in bio-oil was increased and the content of oxygen was decreased with the increasing of solvent polarity. From mass spectrometric analysis it was established that in the positive electrospray ionization (ESI) mode the compounds containing two nitrogen atoms dominate and the considerable portion of the compounds containing single nitrogen atom are also presented for both bio-oil and residual organics samples. In the negative ESI mode the compounds containing four oxygen atoms dominated except bio-oil obtained using hexane where the compounds containing two oxygen atoms dominated. Bio-oil fraction had highly saturated compounds with low double bond equivalent values while the compounds of the residual organics fraction had large double bond equivalent values.

Effect of ion clouds micromotion on measured signal in Fourier transform ion cyclotron resonance: Computer simulation.

Particle-in-cell-based realistic simulation of Fourier transform ion cyclotron resonance experiments could be used to generate ion trajectories and a signal induced on the detection electrodes. It has been shown recently that there is a modulation of "reduced" cyclotron frequencies in ion cyclotron resonance signal caused by Coulomb interaction of ion clouds. In this work it was proposed to use this modulation in order to determine frequency difference between an ion of known m/z and all other ions generating signal in ion cyclotron resonance cell. It is shown that with an increase of number of ions in ion cyclotron resonance trap, the modulation index increases, which lead to a decrease in the accuracy of determination of peak intensities by super Fourier transform resolution methods such as filter diagonalization method.

The investigation of the bio-oil produced by hydrothermal liquefaction of Spirulina platensis using ultrahigh resolution Fourier transform ion cyclotron resonance mass spectrometry.

We report the investigation of the hydrothermal liquefaction products of the Spirulina platensis microalgae by using the Fourier transform ion cyclotron resonance mass spectrometry. The hydrothermal liquefaction produced two fractions: one with boiling temperature below 300℃ and the dense residue that remained in the reactor. It was observed that N2 and N classes of compounds that dominate in the positive ESI Fourier transform ion cyclotron resonance spectra for both fractions, and that the light fraction is considerably more saturated then the heavy one. The performed hydrogen/deuterium exchange reaction indicated the presence of the onium compounds in the bio-oil.

Fourier transform ion cyclotron resonance (FT ICR) mass spectrometry: Theory and simulations.

Fourier transform ion cyclotron resonance (FT ICR) mass spectrometer offers highest resolving power and mass accuracy among all types of mass spectrometers. Its unique analytical characteristics made FT ICR important tool for proteomics, metabolomics, petroleomics, and investigation of complex mixtures. Signal acquisition in FT ICR MS takes long time (up to minutes). During this time ion-ion interaction considerably affects ion motion and result in decreasing of the resolving power. Understanding of those effects required complicated theory and supercomputer simulations but culminated in the invention of the ion trap with dynamic harmonization which demonstrated the highest resolving power ever achieved. In this review we summarize latest achievements in theory and simulation of FT ICR mass spectrometers.

Effect of magnetic field inhomogeneity on ion cyclotron motion coherence at high magnetic field.

A three-dimensional code based on the particle-in-cell algorithm modified to account for the inhomogeneity of the magnetic field was applied to determine the effect of Z(1), Z(2), Z(3), Z(4), X, Y, ZX, ZY, XZ(2) YZ(2), XY and X(2)-Y(2) components of an orthogonal magnetic field expansion on ion motion during detection in an FT-ICR cell. Simulations were performed for magnetic field strengths of 4.7, 7, 14.5 and 21 Tesla, including experimentally determined magnetic field spatial distributions for existing 4.7 T and 14.5 T magnets. The effect of magnetic field inhomogeneity on ion cloud stabilization ("ion condensation") at high numbers of ions was investigated by direct simulations of individual ion trajectories. Z(1), Z(2), Z(3) and Z(4) components have the largest effect (especially Z(1)) on ion cloud stability. Higher magnetic field strength and lower m/z demand higher relative magnetic field homogeneity to maintain cloud coherence for a fixed time period. The dependence of mass resolving power upper limit on Z(1) inhomogeneity is evaluated for different magnetic fields and m/z. The results serve to set the homogeneity requirements for various orthogonal magnetic field components (shims) for future FT-ICR magnet design.

Twelve million resolving power on 4.7 T Fourier transform ion cyclotron resonance instrument with dynamically harmonized cell--observation of fine structure in peptide mass spectra.

Resolving power of about 12,000 000 at m/z 675 has been achieved on low field homogeneity 4.7 T magnet using a dynamically harmonized Fourier transform ion cyclotron resonance (FT ICR) cell. Mass spectra of the fine structure of the isotopic distribution of a peptide were obtained and strong discrimination of small intensity peaks was observed in case of resonance excitation of the ions of the whole isotopic cluster to the same cyclotron radius. The absence of some peaks from the mass spectra of the fine structure was explained basing on results of computer simulations showing strong ion cloud interactions, which cause the coalescence of peaks with m/z close to that of the highest magnitude peak. The way to prevent peak discrimination is to excite ion clouds of different m/z to different cyclotron radii, which was demonstrated and investigated both experimentally and by computer simulations.

From Supercomputer Modeling to Highest Mass Resolution in FT-ICR.

Understanding of behavior of ion ensembles inside FT-ICR cell based on the computer simulation of ion motion gives rise to the new ideas of cell designs. The recently introduced novel FT-ICR cell based on a Penning ion trap with specially shaped excitation and detection electrodes prevents distortion of ion cyclotron motion phases (normally caused by non-ideal electric trapping fields) by averaging the trapping DC electric field during the ion motion in the ICR cell. Detection times of 5 min resulting in resolving power close to 40,000,000 have been reached for reserpine at m/z 609 at a magnetic field of only 7 Tesla. Fine structures of resolved 13Cn isotopic cluster groups could be measured for molecular masses up to 5.7 kDa (insulin) with resolving power of 4,000,000 at 7 Tesla. Based on resolved fine structure patterns atomic compositions can be directly determined using a new developed algorithm for fine structure processing. Mass spectra of proteins and multimers of proteins reaching masses up to 186 kDa (enolase tetramer) could be measured with isotopic resolution. For instance, at 7 Tesla resolving power of 800,000 was achieved for enolase dimer (96 kDa) and 500,000 for molecular masses above 100 kDa. Experimental data indicate that there is practically no limit for the resolving power of this ICR cell except by collisional damping in the ultrahigh vacuum chamber.

Dynamically harmonized FT-ICR cell with specially shaped electrodes for compensation of inhomogeneity of the magnetic field. Computer simulations of the electric field and ion motion dynamics.

The recently introduced ion trap for FT-ICR mass spectrometers with dynamic harmonization showed the highest resolving power ever achieved both for ions with moderate masses 500-1000 Da (peptides) as well as ions with very high masses of up to 200 kDa (proteins). Such results were obtained for superconducting magnets of very high homogeneity of the magnetic field. For magnets with lower homogeneity, the time of transient duration would be smaller. In superconducting magnets used in FT-ICR mass spectrometry the inhomogeneity of the magnetic field in its axial direction prevails over the inhomogeneity in other directions and should be considered as the main factor influencing the synchronic motion of the ion cloud. The inhomogeneity leads to a dependence of the cyclotron frequency from the amplitude of axial oscillation in the potential well of the ion trap. As a consequence, ions in an ion cloud become dephased, which leads to signal attenuation and decrease in the resolving power. Ion cyclotron frequency is also affected by the radial component of the electric field. Hence, by appropriately adjusting the electric field one can compensate the inhomogeneity of the magnetic field and align the cyclotron frequency in the whole range of amplitudes of z-oscillations. A method of magnetic field inhomogeneity compensation in a dynamically harmonized FT-ICR cell is presented, based on adding of extra electrodes into the cell shaped in such a way that the averaged electric field created by these electrodes produces a counter force to the forces caused by the inhomogeneous magnetic field.

Performance of Orbitrap mass analyzer at various space charge and non-ideal field conditions: simulation approach.

The orbital trap mass analyzer provides a number of unique analytical features along with inevitable limitations as an electrostatic instrument operating in high space charge regimes resulting in systematic measured frequency errors as an effect of stored ion clouds on the trap field and each other effect of non-ideal machining the trap electrodes, effect of injection slot, effect of real versus theoretical trap dimensions, etc. This paper deals with determining the influence of the space charge effect and imperfection of the electrostatic field on the motion of ion ensembles in the orbital trap. We examine effects of theoretically modeled non-harmonicity of the electrostatic potential and the number of confined ions on stability of coherent ion motion in the trap that determines the frequency shifts of axial ion oscillation. Three different Orbitrap geometries were considered: geometry close to preproduction Orbitrap, close to standard Orbitrap, close to high field Orbitrap. Frequency shifts for m/z = 500 and for charge state +23 of cytochrome c isotopic cluster particles with 10(4)-6*10(6) elemental charges in the trap were considered. Refined spectra were calculated using the filter diagonalization method proposed by Mandelshtam et al. and applied to mass spectrometry by O'Connor and Aizikov.

Fourier transform ion cyclotron resonance mass resolution and dynamic range limits calculated by computer modeling of ion cloud motion.

Particle-in-Cell (PIC) ion trajectory calculations provide the most realistic simulation of Fourier transform ion cyclotron resonance (FT-ICR) experiments by efficient and accurate calculation of the forces acting on each ion in an ensemble (cloud), including Coulomb interactions (space charge), the electric field of the ICR trap electrodes, image charges on the trap electrodes, the magnetic field, and collisions with neutral gas molecules. It has been shown recently that ion cloud collective behavior is required to generate an FT-ICR signal and that two main phenomena influence mass resolution and dynamic range. The first is formation of an ellipsoidal ion cloud (termed "condensation") at a critical ion number (density), which facilitates signal generation in an FT-ICR cell of arbitrary geometry because the condensed cloud behaves as a quasi-ion. The second phenomenon is peak coalescence. Ion resonances that are closely spaced in m/z coalesce into one resonance if the ion number (density) exceeds a threshold that depends on magnetic field strength, ion cyclotron radius, ion masses and mass difference, and ion initial spatial distribution. These two phenomena decrease dynamic range by rapid cloud dephasing at small ion density and by cloud coalescence at high ion density. Here, we use PIC simulations to quantitate the dependence of coalescence on each critical parameter. Transitions between independent and coalesced motion were observed in a series of the experiments that systematically varied ion number, magnetic field strength, ion radius, ion m/z, ion m/z difference, and ion initial spatial distribution (the present simulations begin from elliptically-shaped ion clouds with constant ion density distribution). Our simulations show that mass resolution is constant at a given magnetic field strength with increasing ion number until a critical value (N) is reached. N dependence on magnetic field strength, cyclotron radius, ion mass, and difference between ion masses was determined for two ion ensembles of different m/z, equal abundance, and equal cyclotron radius. We find that N and dynamic range depend quadratically on magnetic field strength in the range 1-21 Tesla. Dependences on cyclotron radius and Δm/z are linear. N depends on m/z as (m/z)(-2). Empirical expressions for mass resolution as a function of each of the experimental parameters are presented. Here, we provide the first exposition of the origin and extent of trade-off between FT-ICR MS dynamic range and mass resolution (defined not as line width, but as the separation between the most closely resolved masses).