Structural Distinction of Diacyl-, Alkylacyl,and Alk-1-Enylacyl Glycerophosphocholines as [M – 15]– Ions by Multiple-Stage Linear Ion-Trap Mass Spectrometry with Electrospray Ionization

We describe a linear ion-trap (LIT) multiple-stage (MSⁿ) mass spectrometric approach towards differentiation of alkylacyl, alk-1-enylacyl- and diacyl-glycerophoscholines (PCs) as the [M – 15]^– ions desorbed by electrospray ionization (ESI) in the negative-ion mode. The MS⁴ mass spectra of the [M – 15 – R₂′CH = CO]^– ions originated from the three PC subfamilies are readily distinguishable, resulting in unambiguous distinction of the lipid classes. This method is applied to two alkyl ether rich PC mixtures isolated from murine bone marrow neutrophils and kidney, respectively, to explore its utility in the characterization of complex PC mixture of biological origin, resulting in the realization of the detailed structures of the PC species, including various classes and many minor isobaric isomers.

Elucidation of O-Phosphoryl and N-Phosphoryl Amino Acids by Electrospray Ionization Tandem Mass Spectrometry

Mass spectroscopic characteristics of phosphoryl amino acids were studied in detail by positive and negative electrospray ionization mass spectrometry (ESI-MS) in conjunction with tandem mass spectrometry (MS/MS). Besides N-diisopropyloxyphosphoryl amino acids (N-DIPP-AA), O-phospho- and O-diisopropyloxyphosphoryl amino acids (O-DIPP-AA) were studied and compared to N-DIPP-AA. The fragmentation pathways of [M H]^ ,[M Na]^ and [M-H]^- ions of phosphoryl amino acids were summarized. In addition to several similar patterns, each of them showed its characteristic fragmention.     

Analysis of the Aconitine Alkaloids in Chuanwu by Electrospray Ionization ／ Tandem Mass Spectrometry

An electrospray ionization / tandem mass spectrometric (ESUMS/MS) method was developed for the simultaneous identification and analysis of three aconitine alkaloids [mesacontine (MA), hypaconitine (HA), and aconitine (A)] as intact molecules at low nanogram level in Chinese traditional medicine Chuanwu decoction as well as in human whole blood extract without chromatographic separation.     

Novel Rearrangement of Small Peptides in Electrospray Ionization Tandem Mass Spectrometry

Electrospray ionization mass spectrometry (ESI-MS) is a powerful method for sequencing peptides. A novel fragmentation pattern with the loss of a neutral fragment of 45 Da was observed with the dipeptides, tripeptides,tetrapeptides and pentapeptides containing phenylalanine or histidine residues. A novel rearrangement reaction with the extrusion of a formamide piece was studied and the rearrangement mechanism was proposed and confirmed by deuterium labeling experiments with ESI-MS^n and high-resolution mass spectrometry. These findings are potentially helpful in identifying the specific sequence pattern in the peptide sequencing.     

Electrospray Ionization Mass Spectrometry of Hexanitrohexaazaisowurtzitane (CL‐20)

Abstract

Hexanitrohexaazaisowurzitane (CL‐20) is a newly developed propellant and has been examined using negative ion electrospray mass spectrometry. The primary ions observed included adduct ions (M+Cl)^– and (M+ONO₂)^?.     

New Fragmentation Pathways for Cephalosporins by Electrospray Ionization Quadrupole Time-of-flight Mass Spectrometry

Introduction Cephalosporins are antibiotics of β-lactam family. They have a broad spectrum of antibiotic activity due to their ability to inhibit bacterial cell wall synthesis of different Gram-positive and Gram-negative bacteria. Cephalosporins are used orally or parenterally to treat a wide variety of infections throughout the body and are often prescribed to fight penicillin resistant microorganisms.     

The Study on the Noncovalent Complexes of Silver Ion with Xanthone Diglycosides by Electrospray Ionization with Tandem Mass Spectrometry

Xanthones are one species of C6-C1-C6 structure flavonoids, showing the various pharmacological activities, such as hepatocytes1 and antioxidative2 etc.. As one of the softest ionization technique, atmospheric pressure ionization mass spectrometry (API-MS…     

Characterization of the Retro-[3+2] Reaction of Protonated 2,3′-Bisindolylmethanes by Electrospray Ionization–Tandem Mass Spectrometry

Twelve 2,3′-bisindolylmethanes with various substituents were investigated using electrospray ionization quadrupole time-of-flight tandem mass spectrometry in positive ion mode. A retro-[3+2] reaction was observed in the collision-induced dissociation spectra of protonated 2,3′-bisindolylmethanes for the first time. The mechanism of retro-[3+2] reaction was concerted or stepwise. For the concerted pathway, carbon–carbon bonds of a protonated compound simultaneously cracked and the m/z 208 ion ([C₁₅H₁₀D₂N]⁺) was observed with hydrogen–deuterium exchange labeling. The stepwise pathway goes through 1,3-hydrogen migration twice and the m/z 208 ion ([C₁₅H₁₀D₂N]⁺) and m/z 207 ion ([C₁₅H₁₁DN]⁺) were detected with deuterium labeling. In the deuterium-labeled tandem mass spectrum for one compound, only the peak at m/z 208 was present at high abundance, suggesting that the concerted pathway is more likely. In addition, the substituents have no obvious trends on the ratios of the product intensity to the base intensity, further supporting the concerted pathway.     

Probing Conformational Changes of Ubiquitin by Host–Guest Chemistry Using Electrospray Ionization Mass Spectrometry

We report mechanistic studies of structural changes of ubiquitin (Ub) by host–guest chemistry with cucurbit[6]uril (CB[6]) using electrospray ionization mass spectrometry (ESI-MS) combined with circular dichroism spectroscopy and molecular dynamics (MD) simulation. CB[6] binds selectively to lysine (Lys) residues of proteins. Low energy collision-induced dissociation (CID) of the protein-CB[6] complex reveals CB[6] binding sites. We previously reported (Anal. Chem. 2011, 83, 7916–7923) shifts in major charge states along with Ub-CB[6] complexes in the ESI-MS spectrum with addition of CB[6] to Ub from water. We also reported that CB[6] is present only at Lys₆ or Lys₁₁ in high charge state (+13) complex. In this study, we provide additional information to explain unique conformational change mechanisms of Ub by host–guest chemistry with CB[6] compared with solvent-driven conformational change of Ub. Additional CID study reveals that CB[6] is bound only to Lys₄₈ and Lys₆₃ in low charge state (+7) complex. MD simulation studies reveal that the high charge state complexes are attributed to the CB[6] bound to Lys₁₁. The complexation prohibits salt bridge formation between Lys₁₁ and Glu₃₄ and induces conformational change of Ub. This results in formation of high charge state complexes in the gas phase. Then, by utilizing stronger host–guest chemistry of CB[6] with pentamethylenediamine, refolding of Ub via detaching CB[6] from the protein is performed. Overall, this study gives an insight into the mechanism of denatured Ub ion formation via host-guest interactions with CB[6]. Furthermore, this provides a direction for designing function-controllable supramolecular system comprising proteins and synthetic host molecules.      

Monitoring the Hydrolysis of p—Nitrophenyl Acetate Catalyzed by Seryl—histidine with Electrospray Ionization Mass Spectrometry

ＩｎｔｒｏｄｕｃｔｉｏｎＩｎｔｈｅｐａｓｔｄｅｃａｄｅ ,ｉｔｗａｓｆｏｕｎｄｔｈａｔｔｗｏａｍｉｎｏａｃｉｄｒｅｓｉｄｕｅｓ ,ＳｅｒａｎｄＨｉｓ ,ｗｏｒｋａｓｔｈｅａｃｔｉｖｅｓｉｔｅｓｉｎｔｈｅｓｅｒ ｉｎｅｐｒｏｔｅａｓｅ .1 3 Ｉｎｏｕｒｐｒｅｖｉｏｕｓｗｏｒｋ ,ａｄｉｐｅｐｔｉｄｅｓｅｒｙｌ ｈｉｓｔｉｄｉｎｅ (Ｓｅｒ Ｈｉｓ)ｗａｓｆｏｕｎｄｔｏｈａｖｅｔｈｅｐｒｏｔｅｉｎａｎｄｎｕ ｃｌｅｏｓｉｄｅｓｃｌｅａｖａｇｅａｃｔｉｖｉｔｙ .4 7ｐ Ｎｉｔｒｏｐｈｅｎｙｌａｃｅｔａｔｅ (ｐ ＮＰＡ)ｗ…     

Analysis of Norditerpenoid Alkaloids. Extracted from Aconitum sinomantanum Nakai by Electrospray Ionization Tandem Mass Spectrometry

Introduction AconitumsinomantanumNakai(GaowutouinChi nese)isdistributedinthenorthwesternareaofChina,anditsroots(RAS)areexternallyusedasakindof folkmedicineinChina.Themainactiveconstituentsof RASarenorditerpenoidalkaloids.Themajoralkaloid,lappaconitine,was…     

High Pressure (>1 atm) Electrospray Ionization Mass Spectrometry

High pressure electrospray ionization mass spectrometry has been performed by pressurizing a custom made ion source chamber with compressed air to a pressure higher than the atmospheric pressure. The ion source was coupled to a commercial time-of-flight mass spectrometer using a nozzle-skimmer arrangement. The onset voltage for the electrospray of aqueous solution was found to be independent on the operating pressure. The onset voltage for the corona discharge, however, increased with the rise of pressure following the Paschen’s law. Thus, besides having more working gas for the desolvation process, gaseous breakdown could also be avoided by pressurizing the ESI ion source with air to an appropriate level. Stable electrospray ionization has been achieved for the sample solution with high surface tension such as pure water in both positive and negative ion modes. Fragmentation of labile compounds during the ionization process could also be reduced by optimizing the operating pressure of the ion source.     

Rapid Determination of Coumatetralyl in Human Serum by High‐Performance Liquid Chromatography Coupled with Electrospray Ionization Tandem Mass Spectrometry

Abstract

A rapid, sensitive, and selective high‐performance liquid chromatography‐tandem mass spectrometric method (HPLC‐MS‐MS) for the determination of coumatetralyl in human serum using warfarin as an internal standard has been developed and validated. Coumatetralyl and the internal standard were extracted from the human serum samples by liquid‐liquid extraction with ethyl acetate, followed by separation on a XDB C₁₈ reversed‐phase column (150 mm×2.1 mm i.d., 5 µm) using a mobile phase consisting of acetic acid‐ammonium acetate (5 mmol/L, pH=4.5)/methanol (20:80, v/v) at a constant flow rate of 0.40 mL/min. Coumatetralyl and the internal standard were ionized by negative ion pneumatically assisted electrospray and detected in the multiple‐reaction monitoring mode using precursor→product ion combinations at m/z 291→247 and 307→161, respectively. The calibration curve was linear (r²=0.9945) in the concentration range of 0.5～100.0 ng/mL, with a lower limit of quantification of 0.5 ng/mL in human serum. Intra‐ and inter‐day relative standard deviations were less than 6.3 and 11.0%, respectively. The mean extraction recovery was 87.9% for coumatetralyl and 90.1% for the internal standard. This method is found to be able to determine trace coumatetralyl in human serum and can be used for the diagnosis of poisoned human beings.     

Electrospray Ionization Mass Spectra of Amino Acid Methyl Ester 5′-Phosphoramidates of 2′,3′-Isopropylideneuridine

IntroductionIn recent years, nucleosides and their analogshave been extensively studied as potential anticancerand antiviral agents[1—3]. For example, several purineand pyridine bases and nucleoside analogs are used aschemotherapeutic arsenal. For their biological activity,these analogs should be intracellularly metabolized to5′-mononucleotides by kinase-mediated phosphoryla-tion[4]. To overcome the problemof drug resistance andto improve the membrane penetration, a series of aminoacid phosp…     

Electrospray Ionization Mass Spectra of Amino Acid Methyl Ester 5′-Phosphoramidates of 2′,3′-Isopropylideneuridine

Amino acid methyl ester phosphates were synthesized and determined by using positive-ion mode electrospray ionization mass spectrometry(ESIMS) in combination with multistage tandem mass spectrometry. The fragmentation pathways were investigated, and it was observed that most fragment ions contained the phosphoryl group. It was interesting to observe that the fragmentation pathways of the protonated molecule show some differences when compared with those of the sodium ion adduct. The methoxy group of amino acid methyl ester can migrate from the carbonyl group to the phosphoryl group in the sodium ion adduct.     

Improved Accuracy of Low Affinity Protein–Ligand Equilibrium Dissociation Constants Directly Determined by Electrospray Ionization Mass Spectrometry

There is continued interest in the determination by ESI-MS of equilibrium dissociation constants (K_D) that accurately reflect the affinity of a protein–ligand complex in solution. Issues in the measurement of K_D are compounded in the case of low affinity complexes. Here we present a K_D measurement method and corresponding mathematical model dealing with both gas-phase dissociation (GPD) and aggregation. To this end, a rational mathematical correction of GPD (f_sat) is combined with the development of an experimental protocol to deal with gas-phase aggregation. A guide to apply the method to noncovalent protein–ligand systems according to their kinetic behavior is provided. The approach is validated by comparing the K_D values determined by this method with in-solution K_D literature values. The influence of the type of molecular interactions and instrumental setup on f_sat is examined as a first step towards a fine dissection of factors affecting GPD. The method can be reliably applied to a wide array of low affinity systems without the need for a reference ligand or protein.     

Studies on Fragmentation Pathways of N-Ethoxy （phenyl） phosphoryl Amino Acids by Electrospray Ionization Tandem Mass Spectrometry

IntroductionPhosphorylation often acts as a molecular switchcontrolling the protein activity in different pathways asin metabolism,signal transduction,cell division,andso on.Therefore,N-phosphoryl amino acids play aspecial and important role in biological…     

Sequence pattern Correlation of Amino Acid in Collision—induced Dissociation Electrospray Ionization Mass Spectrometry

A novel approach of seuqence pattern correlation has been applied to predict an expected amino acid sequence from CID ESI-MS spectra.The proposed approach deduces sequence patterns with no help form known protein database such that it is useful to identify an unknown petide or new protein.The algorithm applies a cross-correlation to match an experimental CID spectrum with predicted sequence pattern generated form fragmentation information.The fragmentation knowledge of both y-series and other non y-series are utilized to generate the predicted sequence patterns.In contrast to the normal de novo approach,the proposed approach is insensitive to mass tolerance and non-sussceptive to spectral integrality with no need for selection of a starting point.     

Determination of Phenylethanoid Glycosides in Lagotis brevituba Maxim. by High-Performance Liquid Chromatography–Electrospray Ionization Tandem Mass Spectrometry

High performance liquid chromatography coupled with electrospray ionization quadrupole-time-of-flight tandem mass spectrometry was used to profile the phenylethanoid glycosides in Lagotis brevituba. A total of twenty-three phenylethanoid glycosides were characterized by comparing the retention time and fragmentation with standards, accurate mass measurements, and fragmentation at low and high collision energies. Most phenylethanoid glycosides were reported in L. brevituba for the first time. This established method may be employed for comprehensive quality control of L. brevituba.     

Dissociation Kinetics of the Streptavidin–Biotin Interaction Measured Using Direct Electrospray Ionization Mass Spectrometry Analysis

Dissociation rate constants (k _off ) for the model high affinity interaction between biotin (B) and the homotetramer of natural core streptavidin (S₄) were measured at pH 7 and temperatures ranging from 15 to 45 °C using electrospray ionization mass spectrometry (ESI-MS). Two different approaches to data analysis were employed, one based on the initial rate of dissociation of the (S₄? + ?4B) complex, the other involving nonlinear fitting of the time-dependent relative abundances of the (S₄? + ?iB) species. The two methods were found to yield k _off values that are in good agreement, within a factor of two. The Arrhenius parameters for the dissociation of the biotin–streptavidin interaction in solution were established from the k _off values determined by ESI-MS and compared with values measured using a radiolabeled biotin assay. Importantly, the dissociation activation energies determined by ESI-MS agree, within 1 kcal?mol^–1, with the reported value. In addition to providing a quantitative measure of k _off , the results of the ESI-MS measurements revealed that the apparent cooperative distribution of (S₄? + ?iB) species observed at short reaction times is of kinetic origin and that sequential binding of B to S₄ occurs in a noncooperative fashion with the four ligand binding sites being kinetically and thermodynamically equivalent and independent.