2,5—双（4—羟基苯亚甲基）环戊酮中双电荷离子存在的串联质谱证据

应用串联质谱的碰撞诱导解离和联动扫描技术，研究了２，５－双（４－羟基苯亚甲基）环戊酮的质谱解离特征，提供了双电荷离子存在的实验证据。进一步对双电荷离子（ｍ／ｚ１４６）的碰撞诱导解离碎裂进行了讨论。     

Peptide and protein characterization by high-rate electron capture dissociation Fourier transform ion cyclotron resonance mass spectrometry

The analytical utility of the electron capture dissociation (ECD) technique, developed by McLafferty and co-workers, has substantially improved peptide and protein characterization using Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS). The limitations of the first ECD implementations on commercial instruments were eliminated by the employment of low-energy electron-injection systems based on indirectly heated dispenser cathodes. In particular, the ECD rate and reliability were greatly increased, enabling the combination of ECD/FTICR-MS with on-line liquid separation techniques. Further technique development allowed the combination of two rapid fragmentation techniques, high-rate ECD and infrared multiphoton dissociation (IRMPD), in a single experimental configuration. Simultaneous and consecutive irradiations of trapped ions with electrons and photons extended the possibilities for ion activation/dissociation and led to improved peptide and protein characterization. The application of high-rate ECD/FTICR-MS has demonstrated its power and unique capabilities in top-down sequencing of peptides and proteins, including characterization of post-translational modifications, improved sequencing of peptides with multiple disulfide bridges and secondary fragmentation (w-ion formation). Analysis of peptide mixtures has been accomplished using high-rate ECD in bottom-up mass spectrometry based on mixture separation by liquid chromatography and capillary electrophoresis. This paper summarizes the current impact of high-rate ECD/FTICR-MS for top-down and bottom-up mass spectrometry of peptides and proteins.     

Ion trap tandem mass spectrometric determination of F2-isoprostanes

A new tandem mass spectrometry (MS/MS) method based on the use of an ion trap mass spectrometer for the identification and quantitation of F(2)-isoprostanes has been developed. It consists of two solid-phase extractions and two derivation steps followed by gas chromatography/negative ion chemical ionization tandem mass spectrometry (GC/NICI-MS/MS) analysis. This method is highly selective and sensitive and it has been successfully applied to biological samples.     

Gallium metal affinity capture tandem mass spectrometry for the selective detection of phosphopeptides in complex mixtures

Metal affinity capture tandem mass spectrometry (MAC-MSMS) is evaluated in a comparative study of a lysine-derived nitrilotriacetic acid (N(alpha), N(alpha)-bis-(carboxymethyl)lysine, LysNTA) and an aspartic-acid-related iminodiacetic acid (N-(4-aminobutyl)aspartic acid, AspIDA) as selective phosphopeptide detection reagents. Both LysNTA and AspIDA spontaneously form ternary complexes with Ga(III) and phosphorylated amino acids and phosphopeptides upon mixing in solution. Collision-induced dissociation of positive complex ions produced by electrospray produces common fragments (LysNTA + H)(+) or (AspIDA + H)(+) at m/z 263 and 205, respectively. MSMS precursor scans using these fragments as reporter ions allow one to selectively detect multiple charge states of phosphopeptides in mixtures. It follows from this comparative study that LysNTA is superior to AspIDA in detecting phosphopeptides, possibly because of the higher coordination number and greater stability constant for Ga(III)-phosphopeptide complexation of the former reagent. In a continuing development of MAC-MSMS for proteomics applications, we demonstrate its utility in a post-column reaction format. Using a simple post-column-reaction 'T' and syringe pump to deliver our chelating reagents, alpha-casein tryptic phosphopeptides can be selectively analyzed from a solution containing a twofold molar excess of bovine serum albumin. The MAC-MSMS method is shown to be superior to the commonly used neutral loss scan for the common loss of phosphoric acid.     

Ion chromatography tandem mass spectrometry for simultaneous confirmation and determination of indandione rodenticides in serum

This paper describes a simple method for the simultaneous determination and confirmation of the indandione rodenticides in serum. After samples were extracted with 10% (v/v) methanol in acetonitrile and cleaned by solid‐phase extraction, chromatographic separation was performed on an IonPac® AS11 analytical column (250 × 4.0 mm) using gradient KOH eluent with 10% (v/v) methanol as organic modifier. Confirmation was depended on the extensive fragmentation of the indandione molecule under MS/MS conditions which provides sufficient structural information. Quantification was performed by negative electrospray ionization in multiple reaction monitoring mode. All the method parameters were validated. It was confirmed that this method could be used in clinical diagnosis and forensic toxicology. Copyright © 2009 John Wiley & Sons, Ltd.     

Structural characterization of glycoporphyrins by electrospray tandem mass spectrometry

Porphyrin derivatives having a galactose or a bis(isopropylidene)galactose structural unit, linked by ester or ether bonds, were characterized by electrospray tandem mass spectrometry (ES-MS/MS). The electrospray mass spectra of these glycoporphyrins show the corresponding [M + H](+) ions. For the glycoporphyrins with pyridyl substituents and those having a tetrafluorophenyl spacer, the doubly charged ions [M + 2H](2+) were also observed in ES-MS with high relative abundance. The fragmentation of both [M + H](+) and [M + 2H](2+) ions exhibited common fragmentation pathways for porphyrins with the same sugar residue, independently of the porphyrin structural unit and type of linkage. ES-MS/MS of the [M + H](+) ions of the galactose-substituted porphyrins gave the fragment ions [M + H - C(2)H(4)O(2)](+), [M + H - C(3)H(6)O(3)](+), [M + H - C(4)H(8)O(4)](+) and [M + H - galactose residue](+). The fragmentation of the [M + 2H](2+) ions of the porphyrins with galactose shows the common doubly charged fragment ions [porphyrin + H](2+), [M + 2H - C(2)H(4)O(2)](2+), [M + 2H - C(4)H(8)O(4)](2+), [M + 2H - galactose residue](2+) and the singly charged fragment ions [M + H - C(3)H(6)O(3)](+) and [M + H - galactose residue](+). The fragmentation of the [M + H](+) ions of glycoporphyrins with a protected galactosyl residue leads mainly to the ions [M + H - CO(CH(3))(2)](+), [M + H - 2CO(CH(3))(2)](+), [M + H - 2CO(CH(3))(2) - CO](+), [M + H - C(10)H(16)O(4)](+) and [M + H - protected galactose](+). The doubly charged ions [M + 2H](2+) fragment to give the doubly charged ions [porphyrin + H](2+) and the singly charged ions [M + H - protected galactose residue](+) and [M + H - CO(CH(3))(2)](+). For the porphyrins where the sugar structural unit is linked by an ester bond, [M + 2H](2+), ES-MS/MS showed a major and typical fragmentation corresponding to combined loss of a sugar structural unit and further loss of water, leading to the ion [M + 2H - sugar residue - H(2)O](2+), independently of the structure of the sugar structural unit. These results show that ES-MS/MS can be a powerful tool for the characterization of the sugar structural unit of glycoporphyrins, without the need for chemical hydrolysis.     

Charge permutation reactions in tandem mass spectrometry

Central to the tandem mass spectrometry experiment is the process that gives rise to product ions, i.e. the reaction intermediate to stages of mass analysis. Changes in mass or charge of the parent ion (or both) are generally readily detected by all forms of tandem mass spectrometry. Charge changing, or charge permutation, reactions have a long history in mass spectrometry. However, with the advent of new ionization methods, such as electrospray ionization, and the expansion of tandem mass spectrometry instrumentation to include ion trapping instruments, the past decade has seen a major increase in the types of charge permutation reactions that can be studied. Most charge permutation reactions involve electrons or protons as the charge mediating agents. This report, therefore, provides an overview of charge permutation reactions involving protons or electrons. Particular emphasis is placed on processes that involve interactions of precursor ions with gaseous neutral species, electrons or oppositely charged ions. Charge permutation reactions involving electron gain/loss are described first according to a rough order of the energy required for the reaction beginning with the most endoergic reactions and ending with the most exoergic reactions. An analogous approach is then taken with charge permutation reactions involving proton gain/loss. Important charge permutation reactions discussed herein, among others, include those referred to as charge inversion, charge stripping, electron capture dissociation, collision-induced ionization and charge separation. These reaction types, and others described herein, are the subjects of active research and are also finding use in many current areas of application. Copyright © 2004 John Wiley & Sons, Ltd.     

Multi-parameter investigation of tandem mass spectrometry in a linear ion trap using response surface modelling

The feasibility of experimental design in combination with subsequent response surface modelling was illustrated for the prediction and interpretation of tandem mass spectrometric (MS/MS) fragmentation data using a linear quadrupole ion trap under various experimental conditions. The instrumental parameters included were (i) the pressure of the collision gas, (ii) the collision energy, (iii) the fill time of the linear ion trap and (iv) the scan rate. The spectral intensity and width of five fragment ions of the doubly charged neuro-active peptide bombesin were used for evaluation, and all experiments were performed so as to resemble the results obtained from a liquid chromatographic peak. The reported results show how fairly simple mathematical tools can be utilized successfully to describe fundamental mechanisms associated with multiple collisional activation and collision-induced dissociation processes without an extensively controlled experimental environment. Most beneficial, using the suggested approach, is the ability to study interaction (synergistic) effects between various parameters. As was realized from the results, many interaction effects are indeed significant. For example, the effect on the signal intensity of different collision gas pressure settings is strongly dependent on the settings of the other parameters. The described approach can easily be adopted for optimization purposes of any MS/MS experiment.     

'Top down' protein characterization via tandem mass spectrometry

Technological and scientific advances over the past decade have enabled protein identification and characterization strategies to be developed that are based on subjecting intact protein ions and large protein fragments directly to tandem mass spectrometry. These approaches are referred to collectively as 'top down' to contrast them with 'bottom up' approaches whereby protein identification is based on mass spectrometric analysis of peptides derived from proteolytic digestion, usually with trypsin. A key step in enabling top down approaches has been the ability to assign tandem mass spectrometer product ion identities, which can be done either via high resolving power or through product ion charge state manipulation. The ability to determine product ion charge states has permitted studies of the reactions, including dissociation, ion-molecule reactions, ion-electron reactions and ion-ion reactions of high-mass, multiply charged protein ions. Electrospray ionization combined with high magnetic field strength Fourier transform ion cyclotron resonance has proven to be particularly powerful for detailed protein characterization owing to its high mass resolution and mass accuracy and its ability to effect electron capture-induced dissociation. Other types of tandem mass spectrometers are also beginning to find increasing use in top down protein identification/characterization studies. Charge state manipulation via ion-ion reactions in electrodynamic ion traps, for example, enables top down strategies to be considered using instruments with relatively modest mass resolution capabilities. Precursor ion charge state manipulation techniques have also recently been demonstrated to be capable of concentrating and charge-state purifying proteins in the gas phase. Advances in technologies applied to the structural analysis of whole protein ions and in understanding their reactions, such as those described here, are providing new options for the study of complex protein mixtures.     

Discriminating alkylbenzene isomers with tandem mass spectrometry using a dielectric barrier discharge ionization source

Soft ambient ionization sources generate reactive species that interact with analyte molecules to form intact molecular ions, which allows rapid, sensitive, and direct identification of the molecular mass. We used a dielectric barrier discharge ionization (DBDI) source with nitrogen at atmospheric pressure to detect alkylated aromatic hydrocarbon isomers (C₈H₁₀ or C₉H₁₂). Intact molecular ions [M]^•+ were detected at 2.4 kV_pp, but at increased voltage (3.4 kV_pp), [M + N]⁺ ions were formed, which could be used to differentiate regioisomers by collision-induced dissociation (CID). At 2.4 kV_pp, alkylbenzene isomers with different alkyl-substituents could be identified by additional product ions: ethylbenzene and -toluene formed [M-2H]⁺, isopropylbenzene formed abundant [M-H]⁺, and propylbenzene formed abundant C₇H₇⁺. At an operating voltage of 3.4 kV_pp, fragmentation of [M + N]⁺ by CID led to neutral loss of HCN and CH₃CN, which corresponded to steric hindrance for excited state N-atoms approaching the aromatic ring (C-H). The ratio of HCN to CH₃N loss (interday relative standard deviation [RSD] < 20%) was distinct for ethylbenzene and ethyltoluene isomers. The greater the number of alkyl-substituents (C-CH₃) and the more sterically hindered (meta > para > ortho) the aromatic core, the greater the loss of CH₃CN relative to HCN was.     

Application of liquid chromatography-ion trap mass spectrometry to the characterization of the 16-membered ring macrolide josamycin propionate

Coupled liquid chromatography and ion trap mass spectrometry (LC/MS) was used for the characterization of the semi-synthetic 16-membered ring macrolide josamycin propionate. On-line identification of impurities in this antibiotic complex was performed with an ion trap mass spectrometer without recourse to time-consuming isolation and purification procedures. Ion trap mass spectrometry is ideally suited to identification of impurities because it provides MSn capability, enabling multiple stages of mass spectrometry to obtain the maximum amount of structural information for a given molecule. The ion trap was used with an electrospray ionization source operated in the positive ion mode or with an atmospheric pressure chemical ionization source operated in the negative ion mode. The identity of the unknown compounds was deduced using the MS/MS and MSn collision-induced dissociation spectra of reference substances or structural analogs as interpretative templates, combined with knowledge about the nature of functional group fragmentation behavior. Given the importance attached to the identification of impurities of unknown identity in pharmaceutical substances, this study is useful for companies producing josamycin propionate. The knowledge of the fragmentation behavior is also of importance in further research on other 16-membered macrolides.     

Speciation of organotin compounds by tandem mass spectrometry

The collison-induced dissociation of a range of organotin compounds has been examined by tandem (MS–MS) mass spectrometry in order to investigate the potential of the technique for speciation without prior chromatographic separation. Tables of parent–daughte ion relationships are prepared for a group of six closely related compounds. The results obtained are sufficiently characteristic to distinguish between the isolated compounds and to identify the individual components of a simple mixture. This procedure has potential applications for less volatile compounds and for quantitative analysis.     

Mass spectrometry in the structural analysis of flavonoids

Flavonoids are very common and widespread secondary plant metabolites. They have a wide range of biological and physiological activities and serve as chemotaxonomic marker compounds. Therefore, they have been extensively investigated both in the past and during recent years. The interest in them is still increasing. In the search for new compounds, and also in quality control, there is a need to have reliable methodology for the analysis of flavonoids. Mass spectrometry can make an invaluable contribution because of its high sensitivity, possibilities of coupling with liquid chromatography and the availability of powerful tandem mass spectrometric techniques. A review of currently available mass spectrometric methodology used in the structure elucidation of flavonoids is presented. Sample preparation, liquid chromatographic/mass spectrometric analysis and tandem mass spectrometric procedures for the characterization of flavonoid aglycones, O-glycosides, C-glycosides and acylated glycosides are considered.     

Determination of binding sites in carboplatin-bound cytochrome c using electrospray ionization mass spectrometry and tandem mass spectrometry

Interaction of carboplatin with cytochrome c (Cyt. c) has been investigated by electrospray ionization mass spectrometry (ESI-MS) and tandem mass spectrometry (MS/MS). ESI-MS studies revealed that the ring-opened adducts of carboplatin with Cyt. c were formed in the stoichiometric ratio of 1:1 and 2:1 at pH 5.0 and 37 degrees C and in the stoichiometric ratio of 1:1 only at pH 7.0 and 37 degrees C. It was also found that Cyt. c could be cleaved by carboplatin at pH 2.5 and 50 degrees C. The cleaved fragments of Cyt. c were determined by ESI-MS and MS/MS analysis to be Glu66 approximately Met80, Ac-Gly01 approximately Met65, Glu66 approximately Glu104, Ac-Gly01 approximately Met80 and Ile81 approximately Glu104. The carboplatin prefers to anchor to Met65 first, then to Met80. To further confirm the binding site of Met, AcMet-Gly was used as the model molecule to investigate its interaction with carboplatin and its hydrolysis reaction. On the basis of species detected during the reaction monitored by ESI-MS, a possible pathway of the cleavage reaction was proposed.     

Investigation of activation energies for dissociation of host‐guest complexes in the gas phase using low‐energy collision induced dissociation

A low‐energy collision induced dissociation (CID) (low‐energy CID) approach that can determine both activation energy and activation entropy has been used to evaluate gas‐phase binding energies of host‐guest (H‐G) complexes of a heteroditopic hemicryptophane cage host (Zn (II)@1) with a series of biologically relevant guests. In order to use this approach, preliminary calibration of the effective temperature of ions undergoing resonance excitation is required. This was accomplished by employing blackbody infrared radiative dissociation (BIRD) which allows direct measurement of activation parameters. Activation energies and pre‐exponential factors were evaluated for more than 10 H‐G complexes via the use of low‐energy CID. The relatively long residence time of the ions inside the linear ion trap (maximum of 60 s) allowed the study of dissociations with rates below 1 s^?1. This possibility, along with the large size of the investigated ions, ensures the fulfilment of rapid energy exchange (REX) conditions and, as a consequence, accurate application of the Arrhenius equation. Compared with the BIRD technique, low‐energy CID allows access to higher effective temperatures, thereby permitting one to probe more endothermic decomposition pathways. Based on the measured activation parameters, guests bearing a phosphate (―OPO₃^2?) functional group were found to bind more strongly with the encapsulating cage than those having a sulfonate (―SO₃^?) group; however, the latter ones make stronger bonds than those with a carboxylate (―CO₂^?) group. In addition, it was observed that the presence of trimethylammonium (―N(CH₃)₃⁺) or phenyl groups in the guest's structure improves the strength of H‐G interactions. The use of this technique is very straightforward, and it does not require any instrumental modifications. Thus, it can be applied to other H‐G chemistry studies where comparison of bond dissociation energies is of paramount importance.     

Versatility of tandem mass spectrometry for focused analysis of oxylipids

Liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) in the multiple reaction monitoring (MRM) scan mode has been the primary MS method applied for the target identification of specific and minor oxylipids in complex matrices, such as eicosanoids and docosanoids, which are potent lipid mediators derived from polyunsaturated fatty acid oxygenation. However, the high specificity of MRM can limit the detection of species with m/z MRM transitions not covered by the method. In addition to MRM, tandem‐quadrupole mass analyzers enable other experiments to be conducted, by fragmenting ions via collision‐induced dissociation process (CID). This paper presents the potential of tandem mass spectrometry for the focused analysis of oxylipids. We have successfully developed an LC‐MS/MS method for the identification of precursor ions of m/z 115, a diagnostic product ion of 5‐hydroxy‐ and 5‐epoxy‐fatty acids. As a proof of concept, the developed method was used to discover several oxylipids oxidized at C5 derived from arachidonic acid (C20 : 4) oxygenation in a hypothalamus rat extract that were not identified using the target MRM methodology. The proposed focused MS/MS‐based approach in a tandem mass analyzer has proven to be a powerful strategy to accelerate the identification of oxylipids with structural similarities and assist the field of lipidomic research. Copyright © 2015 John Wiley & Sons, Ltd.     

Direct glycan structure determination of intact N-linked glycopeptides by low-energy collision-induced dissociation tandem mass spectrometry and predicted spectral library searching

Intact glycopeptide MS analysis to reveal site-specific protein glycosylation is an important frontier of proteomics. However, computational tools for analyzing MS/MS spectra of intact glycopeptides are still limited and not well-integrated into existing workflows. In this work, a new computational tool which combines the spectral library building/searching tool, SpectraST (Lam et al. Nat. Methods2008, 5, 873–875), and the glycopeptide fragmentation prediction tool, MassAnalyzer (Zhang et al. Anal. Chem.2010, 82, 10194–10202) for intact glycopeptide analysis has been developed. Specifically, this tool enables the determination of the glycan structure directly from low-energy collision-induced dissociation (CID) spectra of intact glycopeptides. Given a list of possible glycopeptide sequences as input, a sample-specific spectral library of MassAnalyzer-predicted spectra is built using SpectraST. Glycan identification from CID spectra is achieved by spectral library searching against this library, in which both m/z and intensity information of the possible fragmentation ions are taken into consideration for improved accuracy. We validated our method using a standard glycoprotein, human transferrin, and evaluated its potential to be used in site-specific glycosylation profiling of glycoprotein datasets from LC-MS/MS. In addition, we further applied our method to reveal, for the first time, the site-specific N-glycosylation profile of recombinant human acetylcholinesterase expressed in HEK293 cells. For maximum usability, SpectraST is developed as part of the Trans-Proteomic Pipeline (TPP), a freely available and open-source software suite for MS data analysis.     

Manipulating internal energy of protonated biomolecules in electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry

The internal energy of protonated leucine enkephalin has been manipulated in electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry with two newly designed pump-probe experiments. Blackbody infrared radiation was applied to pump an ion population into a well-defined internal energy distribution below the dissociation threshold. Following this pumping stage, the internal energy distribution was probed using on-resonance collisional activation to dissociate the ions. These pump-probe experiments were carried out in two different ways: (a) using on-resonance collisional activation with variable kinetic energies to dissociate the ions at a constant initial ion temperature (determining the precursor ion survival percentage as a function of kinetic energy) and (b) using on-resonance collisional activation with a constant kinetic energy to dissociate the ions at variable initial ion temperatures (to investigate the ion survival yield-initial ion temperature dependence). Using this approach, a detailed study of the effects of the initial ion temperature, the probing kinetic energy and the internal energy loss rate on the effective conversion efficiency of (laboratory-frame) kinetic energy to internal energy was conducted. This conversion efficiency was found to be dependent on the initial ion temperature. Depending on the experimental conditions the conversion efficiency (for collisions with argon) was estimated to be about 4.0 +/- 1.7%, which agrees with that obtained from a theoretical modeling. Finally, the reconstructed curves of the ion survival yield versus the mode of the (final) total internal energy distribution of the activated ion population (after pump and probe events) at different pump-probe conditions reveal the internal energy content of the activated ions.     

Protein identification by peptide mass fingerprinting and peptide sequence tagging with alternating scans of nano-liquid chromatography/infrared multiphoton dissociation Fourier transform ion cyclotron resonance mass spectrometry

We have developed a method for protein identification with peptide mass fingerprinting and sequence tagging using nano liquid chromatography (LC)/Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS). To achieve greater sensitivity, a nanoelectrospray (nano-ES) needle packed with reversed-phase medium was used and connected to the nano-ES ion source of the FTICR mass spectrometer. To obtain peptide sequence tag information, infrared multiphoton dissociation (IRMPD) was carried out in nano-LC/FTICR-MS analysis. The analysis involves alternating nano-ES/FTICR-MS and nano-ES/IRMPD-FTICR-MS scans during a single LC run, which provides sets of parent and fragment ion masses of the proteolytic digest. The utility of this alternating-scan nano-LC/IRMPD-FTICR-MS approach was evaluated by using bovine serum albumin as a standard protein. We applied this approach to the protein identification of rat liver diacetyl-reducing enzyme. It was demonstrated that this enzyme was correctly identified as 3-alpha-hydroxysteroid dehydrogenase by the alternating-scan nano-LC/IRMPD-FTICR-MS approach with accurate peptide mass fingerprinting and peptide sequence tagging.     

离子色谱-质谱联用技术测定饮用水及环境水样中的痕量高氯酸盐

建立了一种测定痕量高氯酸盐的离子色谱-质谱联用方法.选用高容量、强亲水性的阴离子交换柱IonPac AS20(2 mm)进行分离,以淋洗液自动发生器在线产生的KOH为淋洗液,采用等浓度淋洗.在不添加有机溶剂的情况下,淋洗液经过抑制器抑制后直接进入电喷雾-串联质谱(ESI-MS-MS),以负离子模式进行检测.同时采用多元反应监测(MRM)模式对高氯酸盐进行监控,以100.8/84.9、 98.8/66.9和100.8/68.9为监控离子对,以98.8/82.9离子对的峰面积进行定量.高氯酸盐质量浓度在0.05～50 μg/L范围内具有良好的线性(r＝0.9985),检出限(S/N=3)为0.01 μg/L.将该方法用于饮用水以及地下水、雪水等环境水样中高氯酸盐的分析,并进行了加标回收实验,回收率在86%～110%之间,将实际自来水样品连续11次进样,所得高氯酸盐的峰面积的相对标准偏差(RSD)为1.6%.