Collision-activated Dissociation Spectra Research Articles

Chiral and Molecular Recognition through Protonation between Aromatic Amino Acids and Tripeptides Probed by Collision-Activated Dissociation in the Gas Phase.

Chiral and molecular recognition through protonation was investigated through the collision-activated dissociation (CAD) of protonated noncovalent complexes of aromatic amino acid enantiomers with l-alanine- and l-serine-containing tripeptides using a linear ion trap mass spectrometer. In the case of l-alanine-tripeptide (AAA), NH3 loss was observed in the CAD of heterochiral H+(d-Trp)AAA, while H2O loss was the main dissociation pathways for l-Trp, d-Phe, and l-Phe. The protonation site of heterochiral H+(d-Trp)AAA was the amino group of d-Trp, and the NH3 loss occurred from H+(d-Trp). The H2O loss indicated that the proton was attached to the l-alanine tripeptide in the noncovalent complexes. With the substitution of a central residue of l-alanine tripeptide to l-Ser, ASA recognized l-Phe by protonation to the amino group of l-Phe in homochiral H+(l-Phe)ASA. For the protonated noncovalent complexes of His enantiomers with tripeptides (AAA, SAA, ASA, and AAS), protonated His was observed in the spectra, except for those of heterochiral H+(d-His)SAA and H+(d-His)AAS, indicating that d-His did not accept protons from the SAA and AAS in the noncovalent complexes. The amino-acid sequences of the tripeptides required for the recognition of aromatic amino acids were determined by analyses of the CAD spectra.

Collisional Activation Dissociation Mass Spectrometry Studies of Oligosaccharides Conjugated with Na+-Encapsulated Dibenzo-18-Crown-6 Ether

To determine the influence of the cationization agent on the collision activated dissociation (CAD) fragmentation behavior of oligosaccharides, the CAD spectra of the singly protonated, sodiated oligosaccharides and singly sodiated and dibenzo-18-crown-6 ether conjugated oligosaccharides were carefully compared. Each of these three different species showed quite different fragmentation spectra. The comparison of singly protonated and sodiated oligosaccharide CAD spectra revealed that different cationization agents affected the cationization agent adduction sites as well as the fragmentation sites within the oligosaccharides. When the mobility of Na+ was limited by the dibenzo-18-crown-6 ether encapsulation agent, the examined linear oligosaccharides showed fragmentation patterns quite different from the unmodified ones. For the dibenzo-18-crown-6 ether conjugated oligosaccharides, the charge-remote fragmentation pathways were more likely to be activated than the chargedirected pathways. This work demonstrates that dibenzo-18-crown-6 ether conjugation can potentially provide a route to selectively activate the charge-remote fragmentation pathways, albeit to a limited extent, in tandem mass spectrometry studies.

Enantioselective Collision-Activated Dissociation of Gas-Phase Tryptophan Induced by Chiral Recognition of Protonated L-Alanine Peptides.

Enantioselective dissociation in the gas phase is important for enantiomeric enrichment and chiral transmission processes in molecular clouds regarding the origin of homochirality in biomolecules. Enantioselective collision-activated dissociation (CAD) of tryptophan (Trp) and the chiral recognition ability of L-alanine peptides (L-Ala n ; n=2-4) were examined using a linear ion trap mass spectrometer. CAD spectra of gas-phase heterochiral H+(D-Trp)(L-Ala n ) and homochiral H+(L-Trp)(L-Ala n ) noncovalent complexes were obtained as a function of the peptide size n. The H2O-elimination product was observed in CAD spectra of both heterochiral and homochiral complexes for n=2 and 4, and in homochiral H+(L-Trp)(L-Ala3), indicating that the proton is attached to the L-alanine peptide, and H2O loss occurs from H+(L-Ala n ) in the noncovalent complexes. H2O loss did not occur in heterochiral H+(D-Trp)(L-Ala3), where NH3 loss and (H2O+CO) loss were the primary dissociation pathways. In heterochiral H+(D-Trp)(L-Ala3), the protonation site is the amino group of D-Trp, and NH3 loss and (H2O+CO) loss occur from H+(D-Trp). L-Ala peptides recognize D-Trp through protonation of the amino group for peptide size n=3. NH3 loss and (H2O+CO) loss from H+(D-Trp) proceeds via enantioselective CAD in gas-phase heterochiral H+(D-Trp)(L-Ala3) at room temperature, whereas L-Trp dissociation was not observed in homochiral H+(L-Trp)(L-Ala3). These results suggest that enantioselective dissociation induced by chiral recognition of L-Ala peptides through protonation could play an important role in enantiomeric enrichment and chiral transmission processes of amino acids.

High-energy collision-activated and electron-transfer dissociation of gas-phase complexes of tryptophan with Na+, K+, and Ca2+

The structure and reactivity of gas-phase complexes of tryptophan (Trp) with Na+, K+, and Ca2+ were examined by high-energy collision-activated dissociation (CAD) and electron transfer dissociation (ETD) using alkali metal targets. In the CAD spectra of M+Trp (M = Na and K), neutral Trp loss was the primary dissociation pathway, and the product ion of collision-induced intracomplex electron transfer from the indole π ring of Trp to the alkali metal ion was observed, indicating a charge-solvated structure in which Trp is non-zwitterionic. The NH3 loss observed in the CAD spectrum of Ca2+Trp2 is ascribed to a CZ (mixed charge-solvated/zwitterionic)-type structure, in which one Trp is non-zwitterionic and the other Trp adopts a zwitterionic structure with an NH 3 + moiety. The H atom and NH3 losses observed in the ETD spectrum of Ca2+Trp2 indicate the formation of a hypervalent radical in the complex, R–NH3, via electron transfer from the alkali metal target to the NH 3 + group of the CZ-type structure. Ca2+ attachment to Trp cluster induces the zwitterionic structure of Trp in the gas phase, and an electron transfer to the zwitterionic Trp forms the hypervalent radical as a reaction intermediate.

Tandem mass spectrometric analysis of isosorbide‐1,4‐cyclohexane‐dicarboxylic acid polyester oligomer cations using ion‐trap mass spectrometry

Isosorbide is a promising biomass-derived molecule that can be used as a replacement for fossil resource-derived diol monomers used in polyester synthesis. Due to its increased use in sustainable development, it is useful to understand the tandem mass spectrometric (MS/MS) fragmentation pathways of the isosorbide-based copolymer as an aid to interpreting the MS/MS spectra of other isosorbide-containing copolymers. Collision-activated dissociation (CAD) experiments were performed on the sodiated/protonated molecules, [(AB)(n)A+Na(or H)](+), n = 2-5, of isosorbide (A)-1,4-cyclohexanedicarboxylic acid (B) oligomers formed by ion-trap electrospray ionization (ESI). Product ions arose from cleavage of the bonds between isosorbide and 1,4-cyclohexanedicarboxylic acid. In the MS/MS spectra, f(n)'' product ions were most abundant, followed by e(n) ions. McLafferty rearrangement appeared to provide the most facile pathway to yield the abundant f(n)'' and e(n) ions. In addition, a(n), b(n)'', f(n)''u(n)'', and en (+) ions were observed. Inductive cleavage and β-elimination were suggested to be the pathways involved in generating e(n)(+)- and e(n)/b(n)''-type ions, respectively. Based on the obtained CAD spectra, the alternating sequences of two copolymer building blocks, A and B, were unambiguously determined. The fragmentation pathways leading to the observed product ion types were also established.

Characterization of Modified RNA by Top‐Down Mass Spectrometry

Characterization of Modified RNA by Top‐Down Mass Spectrometry

Electron Detachment Dissociation of Underivatized Chloride-Adducted Oligosaccharides

Chloride anion attachment has previously been shown to aid determination of saccharide anomeric configuration and generation of linkage information in negative ion post-source decay MALDI tandem mass spectrometry. Here, we employ electron detachment dissociation (EDD) and collision activated dissociation (CAD) for the structural characterization of underivatized oligosaccharides bearing a chloride ion adduct. Both neutral and sialylated oligosaccharides are examined, including maltoheptaose, an asialo biantennary glycan (NA2), disialylacto-N-tetraose (DSLNT), and two LS tetrasaccharides (LSTa and LSTb). Gas-phase chloride-adducted species are generated by negative ion mode electrospray ionization. EDD and CAD spectra of chloride-adducted oligosaccharides are compared to the corresponding spectra for doubly deprotonated species not containing a chloride anion to assess the role of chloride adduction in the stimulation of alternative fragmentation pathways and altered charge locations allowing detection of additional product ions. In all cases, EDD of singly chloridated and singly deprotonated species resulted in an increase in observed cross-ring cleavages, which are essential to providing saccharide linkage information. Glycosidic cleavages also increased in EDD of chloride-adducted oligosaccharides to reveal complementary structural information compared to traditional (non-chloride-assisted) EDD and CAD. Results indicate that chloride adduction is of interest in alternative anion activation methods such as EDD for oligosaccharide structural characterization.

Electron Induced Dissociation of Singly Deprotonated Peptides

Dissociation of singly charged species is more challenging compared with that of multiply charged precursor ions because singly charged ions are generally more stable. In collision activated dissociation (CAD), singly charged ions also gain less kinetic energy in a fixed electric field compared with multiply charged species. Furthermore, ion-electron and ion-ion reactions that frequently provide complementary and more extensive fragmentation compared with CAD typically require multiply charged precursor ions. Here, we investigate electron induced dissociation (EID) of singly deprotonated peptides and compare the EID fragmentation patterns with those observed in negative ion mode CAD. Fragmentation induced upon electron irradiation and collisional activation is not specific and results in the formation of a wide range of product ions, including b-, y-, a-, x-, c-, and z-type ions. Characteristic amino acid side chain losses are detected in both techniques. However, differences are also observed between EID and CAD spectra of the same species, including formation of odd-electron species not seen in CAD, in EID. Furthermore, EID frequently results in more extensive fragmentation compared with CAD. For modified peptides, EID resulted in retention of sulfonation and phosphorylation, allowing localization of the modification site. The observed differences are likely due to both vibrational and electronic excitation in EID, whereas only the former process occurs in CAD.

Mass Spectrometry Analysis of 2-Nitrophenylhydrazine Carboxy Derivatized Peptides

Peptides with two or more basic residues, including those with post-translational modifications (PTMs), such as methylation and phosphorylation, can be highly hydrophilic and, therefore, are often difficult to be retained on a reversed-phase (RP) column. In addition, these highly hydrophilic peptides may carry two or more positive charges, which often fragment poorly upon collisionally activated dissociation (CAD), resulting in few sequence-specific ions. C-terminal rearrangement may also occur during CAD. Furthermore, some PTMs are labile and tend to be lost when subjected to CAD as is the case with phosphorylation on serine or threonine. To overcome the difficulties of separation, detection, and fragmentation of highly hydrophilic peptides, we report here the effect of carboxy group derivatization with 2-nitrophenylhydrazine (this strategy will be called NPHylation for simplicity). NPHylation significantly increases the hydrophobicity of the peptides, eliminates C-terminal rearrangement in all cases, and offers enhanced sensitivity in some cases. In addition, the CAD spectra of the resulting NPHylated peptides carry more sequence-specific ions due to significant reduction of sequence scrambling as observed for peptide EHAGVISVL. Furthermore, the different carboxy derivatives of this peptide undergo sequence scrambling to varying degrees, which clearly demonstrates that the C-terminus has a profound effect on peptide fragmentation. Finally, sequence scrambling is a charge dependent phenomenon, which affects CAD of doubly charged peptides far more than their singly charged counterparts.

Site-specific Phosphorylation of CXCR4 Is Dynamically Regulated by Multiple Kinases and Results in Differential Modulation of CXCR4 Signaling

The chemokine receptor CXCR4 is a widely expressed G protein-coupled receptor that has been implicated in a number of diseases including human immunodeficiency virus, cancer, and WHIM syndrome, with the latter two involving dysregulation of CXCR4 signaling. To better understand the role of phosphorylation in regulating CXCR4 signaling, tandem mass spectrometry and phospho-specific antibodies were used to identify sites of agonist-promoted phosphorylation. These studies demonstrated that Ser-321, Ser-324, Ser-325, Ser-330, Ser-339, and two sites between Ser-346 and Ser-352 were phosphorylated in HEK293 cells. We show that Ser-324/5 was rapidly phosphorylated by protein kinase C and G protein-coupled receptor kinase 6 (GRK6) upon CXCL12 treatment, whereas Ser-339 was specifically and rapidly phosphorylated by GRK6. Ser-330 was also phosphorylated by GRK6, albeit with slower kinetics. Similar results were observed in human astroglia cells, where endogenous CXCR4 was rapidly phosphorylated on Ser-324/5 by protein kinase C after CXCL12 treatment, whereas Ser-330 was slowly phosphorylated. Analysis of CXCR4 signaling in HEK293 cells revealed that calcium mobilization was primarily negatively regulated by GRK2, GRK6, and arrestin3, whereas GRK3, GRK6, and arrestin2 played a primary role in positively regulating ERK1/2 activation. In contrast, GRK2 appeared to play a negative role in ERK1/2 activation. Finally, we show that arrestin association with CXCR4 is primarily driven by the phosphorylation of far C-terminal residues on the receptor. These studies reveal that site-specific phosphorylation of CXCR4 is dynamically regulated by multiple kinases resulting in both positive and negative modulation of CXCR4 signaling.

Enrichment and Site Mapping of O-Linked N-Acetylglucosamine by a Combination of Chemical/Enzymatic Tagging, Photochemical Cleavage, and Electron Transfer Dissociation Mass Spectrometry

Numerous cellular processes are regulated by the reversible addition of either phosphate or O-linked beta-N-acetylglucosamine (O-GlcNAc) to nuclear and cytoplasmic proteins. Although sensitive methods exist for the enrichment and identification of protein phosphorylation sites, those for the enrichment of O-GlcNAc-containing peptides are lacking. Reported here is highly efficient methodology for the enrichment and characterization of O-GlcNAc sites from complex samples. In this method, O-GlcNAc-modified peptides are tagged with a novel biotinylation reagent, enriched by affinity chromatography, released from the solid support by photochemical cleavage, and analyzed by electron transfer dissociation mass spectrometry. Using this strategy, eight O-GlcNAc sites were mapped from a tau-enriched sample from rat brain. Sites of GlcNAcylation were characterized on important neuronal proteins such as tau, synucleins, and methyl CpG-binding protein 2.

Low-energy collisionally activated dissociation of pentose–borate complexes

Pentose–borate 1:1 complexes were generated in the ESI source of a triple quadrupole and ion trap mass spectrometer by electrospray ionization of Na 2B 4O 7 and pentose (arabinose, lyxose, ribose, xylose) 2:1 solution in CH 3CN/H 2O. The study of their low-energy collisionally activated dissociation (CAD) demonstrated that ribose and lyxose are preferentially complexed at the C 2–C 3 cis-diol function whereas arabinose and xylose are esterified at the C 1–C 2 hydroxyl groups. No evidence was found of the stronger affinity for ribose to borate. The ribose probiotic rule can be explained by considering its peculiar capability, among the investigated pentoses, to almost totally complex the borate anion at the C 2–C 3 hydroxyl group, thus enabling the subsequent stages of nucleotide assembly, such as phosphorylation and linkage to the nucleobases. Finally, the differences observed in the pentose–borate complex CAD spectra can be used for the mass spectrometric discrimination of isomeric pentoses in complex mixtures.

Post-Acquisition ETD Spectral Processing for Increased Peptide Identifications

Tandem mass spectra (MS/MS) produced using electron transfer dissociation (ETD) differ from those derived from collision-activated dissociation (CAD) in several important ways. Foremost, the predominant fragment ion series are different: c- and z(*)-type ions are favored in ETD spectra while b- and y-type ions comprise the bulk of the fragments in CAD spectra. Additionally, ETD spectra possess charge-reduced precursors and unique neutral losses. Most database search algorithms were designed to analyze CAD spectra, and have only recently been adapted to accommodate c- and z(*)-type ions; therefore, inclusion of these additional spectral features can hinder identification, leading to lower confidence scores and decreased sensitivity. Because of this, it is important to pre-process spectral data before submission to a database search to remove those features that cause complications. Here, we demonstrate the effects of removing these features on the number of unique peptide identifications at a 1% false discovery rate (FDR) using the open mass spectrometry search algorithm (OMSSA). When analyzing two biologic replicates of a yeast protein extract in three total analyses, the number of unique identifications with a approximately 1% FDR increased from 4611 to 5931 upon spectral pre-processing--an increase of approximately 28.6%. We outline the most effective pre-processing methods, and provide free software containing these algorithms.

Tandem Mass Spectrometry Characteristics of Silver-Cationized Polystyrenes: Backbone Degradation via Free Radical Chemistry

The [M + Ag](+) ions of polystyrene (PS) oligomers are formed by matrix-assisted laser desorption/ionization, and their fragmentation characteristics are determined by tandem mass spectrometry experiments in a quadrupole/time-of-flight mass spectrometer. Collisionally activated dissociation (CAD) of [M + Ag](+) starts with random homolytic C-C bond cleavages in the PS chain, which generate radical ions carrying either the initiating (a(n*), b(n*)) or the terminating (y(n*), z(n*)) chain end and primary (a(n*), y(n*)) or benzylic (b(n*), z(n*)) radical centers. The fragments ultimately observed arise by consecutive, radical-induced dissociations. The primary radical ions mainly decompose by monomer evaporation and, to a lesser extent, by beta-H(*) loss. The benzylic radical ions primarily decompose by 1,5-H rearrangement (backbiting) followed by beta C-C bond scissions; this pathway leads to either closed-shell fragments with CH(2) end groups, internal fragments with 2-3 repeat units, or truncated benzylic b(n*)/z(n*) radical ions that can undergo anew backbiting. The same internal fragments are produced in all backbiting steps; hence, these fragments and small benzylic radical ions (which cannot undergo backbiting) dominate the low-mass region of the CAD spectra, while the less abundant closed-shell fragments with CH(2) end groups (a(n)/y(n)) dominate the medium- and high-mass regions. The latter fragments are suitable for determining the individual initiating and terminating end groups, whereas the internal ions could be valuable in sequence analyses of styrene copolymers.

Collisionally Activated Dissociation and Electron Capture Dissociation Provide Complementary Structural Information for Branched Permethylated Oligosaccharides

Doubly charged sodiated and permethylated linear malto-oligosaccharides ({Glc}6-{Glc}9), branched N-linked glycans (high-mannose type GlcNAc2Man5-9, and complex asialo- and disialylated-biantennary glycans) were analyzed by tandem mass spectrometry using collisionally-activated dissociation (CAD) and "hot" electron capture dissociation (ECD) available in a custom-built ESI FTICR mass spectrometer. For linear permethylated malto-oligosaccharides, both CAD and "hot" ECD produced glycosidic cleavages (B, Y, C, and Z ions), cross-ring cleavages (A- and X-type), and internal cleavages (B/Y and C/Y type) to provide sequence and linkage information. For the branched N-linked glycans, CAD and "hot" ECD provided complementary structural information. CAD generated abundant B and Y fragment ions by glycosidic cleavages, whereas "hot" ECD produced dominant C and Z ions. A-type cross-ring cleavages were present in CAD spectra. Complementary A- and X-type cross-ring fragmentation pairs were generated by "hot" ECD, and these delineated the branching patterns and linkage positions. For example, 0, 4An and 3, 5An ions defined the linkage position of the major branch as the 6-position of the central core mannose residue. The internal fragments observed in CAD were more numerous and abundant than in "hot" ECD spectra. Since the triply charged (sodiated) molecular ion of the permethylated disialylated-biantennary N-linked glycan has relatively high abundance, it was isolated and fragmented in a "hot" ECD experiment and extensive fragment ions (glycosidic and complementary pairs of cross-ring cleavages) were generated to fully confirm the sequence, branching, and linkage assignments for this glycan.

Secondary ozonides of endo-cyclic alkenes analyzed by atmospheric sampling Townsend discharge ionization mass spectrometry

Secondary ozonides (SOZ) of cyclohexene, 1-methylcyclohexene, 4-isopropyl-1-methylcyclohexene and d-limonene were cryo-synthesized by ozonolysis in pentane and purified on a silica gel column. The mass spectra obtained by atmospheric sampling Townsend discharge ionization (ASTDI) and collision activated dissociation (CAD) of the protonized SOZ showed characteristic losses evident of the ozonide structure. Oxygen was eliminated as, e.g., O and O 2, and loss of (HCHO + HCHO) or (O + CO 2) corresponded to the SOZ base-peak for the substituted cyclohexenes by ASTDI-MS. The CAD spectra of the protonized species by use of methane as chemical ionization gas, showed consecutive losses of three oxygen atoms. Elimination of hydroxy-methyl hydroperoxide (HMHP) was particular important for the protonized SOZ, unlike consecutive loss of (HCHO + HCHO) or (O + CO 2). In addition, the spectra of d-limonene were characterized by an unique loss of H 2O 2. These losses appear to be useful for identification of SOZ in gas-phase ozonolysis mixtures of endo-cyclic alkenes, which makes ASTDI an alternative to other on-line techniques for analysis of SOZ in ozonolysis mixtures.

Dissociation mechanisms of neutral methyl stearate and its hydrogen atom adduct formed from the respective positive ions by electron transfer

The mechanism of dissociation of neutral methyl stearate and its hydrogen atom adduct was investigated by charge inversion mass spectrometry using an alkali metal target. Migrations of functional groups in fatty acid ester ions are often observed during the dissociation of the cations in collisionally activated dissociation (CAD). In the charge inversion spectrum, the main dissociation channels of methyl stearate molecule are the loss of a CH3 radical or a H atom. To identify the source of the CH3 radical and the H atom, the charge inversion spectra of partially deuterated methyl stearate (C17H35COOCD3) were measured. The loss of CH3 occurred through elimination from the methoxy methyl group and that of H occurred through elimination from the hydrocarbon chain of the fatty acid group. In the protonated ester, a simultaneous loss of CH3 (from the methoxy methyl group) and a H atom or a H2 molecule was observed. The charge inversion process gave the dissociation fragments with almost no migration of atoms. Only a few peaks that were structure sensitive were observed in the higher mass region in the charge inversion spectra; these peaks were associated with dissociations of energy-selected neutral species, unlike the case of CAD spectra in which they result from dissociation of ions. Charge inversion mass spectrometry with alkali metal targets provided direct information on the dissociation mechanism of methyl stearate and its hydrogen atom adduct without any migration of functional groups.

A collisionally activated dissociation (CAD) and computational investigation of doubly and singly charged DMSO complexes of Cu2+

The singly and doubly charged Cu(II)–DMSO complexes formed by electrospray have been examined by CAD and computation. The CAD spectra were obtained as a function of collision energy. The doubly charged ions, [Cu(DMSO)n]2+, were observed only for n ≥ 2. For n = 3, dissociation leads mainly to [Cu(DMSO)2]+ + DMSO+, with only a trace of [Cu(DMSO)2]2+. Although [Cu(DMSO)]2+ was never detected, computation shows that the n = 1 complex exists in a potential well. Loss of DMSO+ is computed to be exothermic for n = 1–3, the exothermicity decreasing as n increases. The singly charged complexes in the ESI spectra were [CuX(DMSO)n]+ (X = Cl, Br, NO3, HSO4, n = 1 or 2). The CAD spectra showed competition between electron transfer from anion to metal followed by loss of X and loss of DMSO+. Experiment and computation show that for [CuX(DMSO)]+, loss of X is the preferred decomposition at low collision energy. NBO analysis shows that electron transfer to Cu from DMSO decreases in [Cu(DMSO)n]2+ as n increases, the bonding becoming more electrostatic and less covalent. In [CuX(DMSO)n]+, the negative charge on X is much less than unity with most of the difference appearing on the DMSO ligand(s).Key words: copper–DMSO complexes, electrospray, CAD, structures.

Manipulating the Fragmentation Patterns of Phosphopeptides via Gas-Phase Boron Derivatization: Determining Phosphorylation Sites in Peptides with Multiple Serines

Trivalent boron species readily react with protonated phosphopeptides to give addition products with the loss of boron ligands. In the present study, trimethoxyborane (TMB), diisopropoxymethylborane (DIPM), and diethylmethoxyborane (DEMB) were allowed to react with four phosphopeptides, VsSF, LSsF, LsGASA, and VSGAsA (lower-case s indicates phosphoserine). Each of the phosphopeptides contains one serine that is phosphorylated and one that is not. Under collision-activated dissociation (CAD) conditions, the boron-derivatized peptides give fragmentation patterns that differ significantly from that of the protonated phosphopeptide. The patterns vary, depending on the number of labile (i.e., alkoxy) ligands on the boron. In general, boron derivatization increases the yield of phosphate-containing sequence ions, but dramatic effects are only seen with certain reagent/peptide combinations. However, the suite of reagents provides a means of altering and increasing the information content of phosphopeptide CAD spectra.

Automated assignment of high‐resolution collisionally activated dissociation mass spectra using a systematic bond disconnection approach

AbstractAn essential component of the process of characterising chemical unknowns via mass spectrometry is the analysis of collisionally activated dissociation (CAD) mass spectra. Existing tools for the automated assignment of CAD spectra typically use a rule‐based approach which identifies those bonds that are likely to break. While valuable, the failure of explicitly rule‐based approaches to suggest rationalisations for a significant proportion of observed product ions led us to develop an alternative approach (elucidation of product ion connectivity, EPIC) based on high‐resolution mass spectrometry, systematic bond disconnection of the precursor structure, and ranking of the resulting substructures. We exemplify this approach with a reanalysis of published MS/MS data for two compounds taken from the literature. Copyright © 2005 John Wiley & Sons, Ltd.