Interactions of platinum(II) complexes with sulfur-containing peptides studied by electrospray ionization mass spectrometry and tandem mass spectrometry

Reactions of two platinum(II) complexes, cis-[Pt(NH3)2(H2O)2]2+ (Pt1) and cis-[Pt(en)(H2O)2]2+ (Pt2), with several sulfur-containing peptides, have been investigated by electrospray ionization mass spectrometry (ESI-MS) and tandem mass spectrometry (MS/MS). The species produced in the reactions were detected with ESI-MS, and MS/MS analysis was performed to probe structural information. Collision-induced dissociation revealed different dissociation pathways for the main reaction products of the two platinum(II) complexes with the same peptides. The major difference is the prominent loss of ammonia ligand for complexes of Pt1 due to the strong trans effect of sulfur, whereas the loss of ethylenediamine (en) ligand from Pt2 complexes is less favored, reflecting the chelating effect of the bidentate ligand. Despite the differences in dissociation patterns, Pt1 and Pt2, in general, form structurally similar complexes with the same peptides. In the reactions with Met-Arg-Phe-Ala they both produce a N,S-chelate ring through the N-terminal NH2 and sulfur of the Met residue, and in the reactions with Ac-Met-Ala-Ser they bind to the sulfur of Met and deprotonate an amide nitrogen upstream from the anchor site. Both of them are able to promote hydrolysis of the peptides. In reactions with glutathione they both form four-membered Pt2S2 rings and Pt-S-Pt bonding through the bridging thiolate ligand, although the reaction rate is much slower for Pt2 due to steric hindrance of the en ligand.     

Investigation of disulfonamide ligands derived from o-phenylenediamine and their Pb(II) complexes by electrospray ionization mass spectrometry

An electrospray ionization mass spectrometry (ESI-MS) method, in both positive and negative ion modes, was developed for characterization of disulfonamide ligands derived from o-phenylenediamine and their Pb(II) complexes. For the ligands, negative ion mode ESI-MS in methanolic solutions gave simple and easily interpretable mass spectra. However, the spectra of Pb complexes were not readily interpretable under the same conditions. Protonated ligands and their Pb(II) complexes were observed in methanolic solutions by ESI-MS in positive ion mode. The formation of Na(+), K(+), or NH(4) (+) adducts was also observed, complicating the mass spectra and decreasing the signal intensity. In order to optimize the detection of the ligands and the Pb complexes, a method was developed by adding NaOAc in the solutions. The presence of 0.2 mM NaOAc simplified the mass spectra of the ligands and the Pb complexes, and significantly increased sensitivity in both negative and positive ion modes. This modification makes ESI-MS in both modes suitable for characterization of sulfonamide ligands and their Pb complexes, thus providing a potentially powerful tool for evaluating formation of metal complexes and screening combinatorial ligand libraries.     

Coordination chemistry of chromium-Salen complexes studied by electrospray ionization mass spectrometry

The gas-phase coordination behavior of the [Cr(III)(Salen)]PF(6) complex at the free axial positions has been studied in the presence of amines as ligands (propylamine and a series of diamines) under electrospray ionization conditions. The [Cr(III)(Salen)](+) complex formed stable five- and six-coordinated complex ions, [Cr(III)(Salen)(L)](+) and [Cr(III)(Salen)(L)(2)](+), respectively, where L = solvent molecule or amine. When diamines were used as ligands, abundant [Cr(III)(Salen)(L)](+) ions were observed in which two axial positions of the [Cr(III)(Salen)](+) species are occupied by the two amino groups of the diamine ligand. The relative abundances of ligated complex ions, fragment ions, and solvent adducts of fragment ions in the ESI mass spectra, were found to depend on the cone voltage used to record the spectrum. The ESI mass spectra of [Cr(III)(Salen)](+) in the presence of diamines as ligands, and experiments on ligand-pickup in the collision cell, clearly demonstrated that the [Cr(III)(Salen)(L)](+) complex ion is stable for 1,2-diaminoethane and 1,3-diaminopropane. The stability of [Cr(III)(Salen)(L)](+) ions gradually decreased from 1,4-diaminobutane to 1,6-diaminohexane, and then showed a slight increase for 1,7-diaminoheptane and 1,8-diaminooctane. The collision-induced dissociation spectra of [Cr(III)(Salen)(L)](+) ions support the above observations.     

Complexation between the fungicide tebuconazole and copper(II) probed by electrospray ionization mass spectrometry

Electrospray ionization mass spectrometry (ESI-MS) is used to probe the complex formation between tebuconazole (1) and copper(II) salts, which both are commonly used fungicides in agriculture. Experiments with model solutions containing 1 and CuCl(2) reveal the initial formation of the copper(II) species [(1)CuCl](+) and [(1)(2)CuCl](+) which undergo reduction to the corresponding copper(I) ions [(1)Cu](+) and [(1)(2)Cu](+) under more drastic ionization conditions in the ESI source. In additional experiments, copper/tebuconazole complexes were also detected in samples made from soil solutions of various origin and different amount of mineralization. The direct sampling of such solutions via ESI-MS is thus potentially useful for understanding of the interactions between copper(II) salts and tebuconazole in environmental samples.     

Characterization of ageing products of ester-based synthetic lubricants by liquid chromatography with electrospray ionization mass spectrometry and by electrospray ionization (tandem) mass spectrometry

Ageing products of a commercial jet engine oil based on pentaerythritol tetraesters which were formed upon operation in an aviation turbine were detected by electrospray ionization mass spectrometry (ESI-MS) and characterized by LC-ESI-MS. The fatty acid composition of these ageing products was investigated by ESI-MS-MS analysis. The ammonium adducts of the newly formed pentaerythritol tetraester degradation products were found to be suitable parent ions for further structure elucidation work. ESI-MS, LC-ESI-MS and ESI-MS-MS proved to be versatile tools to study the chemical composition (distribution of homologues) as well as the mechanism of ageing of ester based lubricants on a molecular level. Due to its high sensitivity, ESI-MS can also be used to characterize and identify trace levels of ester-based lubricants.     

Low temperature aqueous electrospray ionization mass spectrometry of noncovalent complexes

In the present study we describe conditions that permit the characterization of noncovalent protein–substrate complexes in aqueous solution by microspray electrospray ionization-mass spectrometry (ESI-MS), using a heated transfer capillary at low temperature (45 °C). Specifically, we examined the binding of calmodulin to two polypeptides; the calmodulin-binding domain of calmodulin-dependent protein kinase II (CamK-II) and melittin. Calmodulin, a well known calcium-binding protein, binds to a number of small amphipathic peptides in a calcium-dependent manner. Our results directly show that both peptides form equimolar complexes with calmodulin only in the presence of calcium. The stoichiometry necessary for the formation of each complex was 1:1:4 for calmodulin:peptide (melittin or CamK-II):Ca²⁺, respectively. Furthermore, it is demonstrated that the detection of the complex in ESI-MS is source temperature dependent.     

Determination of micelle mass by electrospray ionization mass spectrometry

By electrospray ionization (ESI) mass spectrometry, micelle solutions of sodium cholate were investigated in detail in the presence and absence of ethanol. The average aggregation number could be evaluated from the spectra acquired under conditions where soft collisions adequate to measure the micelle solution were induced, and the value agreed well with that obtained previously by other methods. From the dependence on ethanol content, it was also found that the average aggregation number in aqueous solution without organic solvent could be reliably estimated. The ESI method proved to be a useful tool for determining the micelle mass in the original aqueous phase.     

Detection of Al(III) and Ga(III) complexes with morin by electrospray ionization mass spectrometry.

The Al(III) and Ga(III) complexes formed by morin (M) in aqueous solution were investigated by means of electrospray ionization mass spectrometry (ESI-MS). In the full scan mass spectra, Al:M showed 1:2 and 2:3 stoichiometric ratios. When (S)-N-acetylserine methyl ester (Ser), as a partial mimic of the serine residue in silk, was added to Al:M and Ga:M complexes in aqueous solution, the mass spectra of Ser:Al:M showed 1:1:1 and 1:1:2 stoichiometric ratios. The patterns of the mass spectra of Ga:M and Ser:Ga:M complexes were similar to those for the corresponding Al(III) complexes. Calculated heats of formation of potential structures of the complexes, with and without bound water, were obtained using semiempirical PM3 calculations.     

Multiply charged ions of ruthenium(II), rhodium(III) and cobalt(III) complexes in electrospray ionization mass spectrometry

Multiply charged ions from electrospray ionization (ESI) were observed for ruthenium-bidentate ligand complexes, such as [RuL₂B]X₂ and [(RuL₂)₂B]X₄, where L is 2,2′-bipyridine, B are tetradentate ligands of 2,2′-bis(2′-pyridyl)bibenzimidazole and 2,6-bis(2′-pyridyl)benzodiimidazole, bidentate ligand of 2-(2′-pyridyl)benzimidazole and related compounds and X is CIO₄^- or CI^-. ESI mass spectra showed a simple mass pattern for easy structural assignment and detecting impurities. The mass spectra for binuclear complexes provide a charge state distribution ranging from 4+ to 2+ for Ru(II)—Ru(II) compounds and 5+ to 2+ for Ru(II)—Rh(III) compounds. It was found that different multiply charged ions are generated by loss of counterions and by protonation/deprotonation at the proton site of ligands B. The abundances of these ions are qualitatively explained in terms of the acidity of metal complexes depending on the bridging ligand structures and the charge of the metal ions. Ions produced by removal of ligands were hardly observed.     

Observation of noncovalent complexes to the avidin tetramer by electrospray ionization mass spectrometry

Intact avidin-biotin and avidin-biotin maleimide noncovalent complexes have been observed by electrospray ionization mass spectrometry (ESI-MS) by using an extended mass range quadrupole mass spectrometer. By utilizing mild ES1 interface conditions, the expected solution behavior of four biotin or biotin maleimide molecules noncovalently binding to each avidin tetramer can be preserved in the gas phase. The ESI-MS results show the appropriate mass additions of 973 ± 60 Da for biotin and 1802 ± 40 Da for biotin maleimide to the avidin tetramer species. These results support the hypothesis that substantial retention of higher order structure is possible in the gas phase by using gentle ESI conditions.     

Determination of dissociation constants of cyclodextrin-ligand inclusion complexes by electrospray ionization mass spectrometry

The inclusion complexes of four ligands binding to cyclodextrins (CDs) were studied by electrospray ionization mass spectrometry (ESI-MS) and the dissociation constants of the complexes were obtained. The 1:1 stoichiometric inclusion complex was found in the system of CD and fenbufen or aspirin. The obtained KD values of the inclusion complexes of fenbufen binding to alpha-CD and to beta-CD are 4.38x10(-4) mol L(-1) and 2.12x10(-4) mol L(-1), respectively. The KD values of the inclusion complexes of alpha-CD-aspirin and beta-CD-aspirin are 3.33x10(-4) mol L(-1) and 1.83x10(-4) mol L(-1), respectively. A non-linear least squares regression method was applied to validate the results which were consistent with each other. For the system of tetracycline hydrochloride and CD, the 1:1 and 1:2 stoichiometric inclusion complexes were found in the mass spectra. The KD,1 and KD,2 values of the 1:1 and 1:2 stoichiometric inclusion complexes of alpha-CD and tetracycline hydrochloride are 4.47x10(-4) mol L(-1) and 6.51x10(-4) mol L(-1), respectively, and those of beta-CD and tetracycline hydrochloride are 2.26x10(-4) mol L(-1) and 8.57x10(-4) mol L(-1), respectively. For the system of norfloxacin and CD, besides the 1:1 and 1:2 inclusion complexes, the 1:3 stoichiometric inclusion complex was also found. The KD,1, KD,2 and KD,3 of alpha-CD and norfloxacin inclusion complexes are 4.61x10(-4) mol L(-1), 6.05x10(-4) mol L(-1) and 1.45x10(-3) mol L(-1), respectively. The three KD values of beta-CD and norfloxacin are 1.96x10(-4) mol L(-1), 4.93x10(-4) mol L(-1) and 1.15x10(-3) mol L(-1), respectively.     

Novel Mo(V)-dithiolene compounds: characterization of nonsymmetric dithiolene complexes by electrospray ionization mass spectrometry

Three new Mo(V) dithiolene compounds have been synthesized by addition of alkynes ((Me(3)Si)(2)C(2) (TMSA), (Me(3)Si)(2)C(4), and (Ph)(2)C(4) to MoO(2)S(2)(2-) in a MeOH/NH(3) mixture: [Mo(2)(O)(2)(mu-S)(2)(eta(2)-S(2))(eta(2)-S(2)C(2)H(2))](2)(-) 1, [Mo(2)(O)(X)(mu-S)(2)(eta(2)-S(2))(eta(2)-S(2)C(2)Ph(C(2)Ph))](2-) 2 (X = O or S), and [Mo(2)(O)(2)(mu-S)(2)(eta(2)-S(2))(eta(2)-S(2)C(2)H(C(2)H))](2-) 3. The structure of 1 as determined by single-crystal X-ray diffraction study (space group Pbca, a = 13.3148(1) A, b = 15.7467(4) A, c = 28.4108(7) A, V = 5956.7(2) A(3)) is discussed. 2 and 3 have been identified by ESMS (electrospray mass spectrometry), (1)H NMR, (13)C NMR, and infrared spectroscopies. This investigation completes our previous study devoted to the addition of DPA (C(2)Ph(2)) to MoO(2)S(2)(2-) which led to [Mo(2)(O)(X)(mu-S)(2)(eta(2)-S(2))(eta(2)-S(2)C(2)Ph(2))](2-) 4 (X = O or S). A reaction scheme is proposed to explain the formation of the different species present in solution. The reactivity of the remaining nucleophilic site of these complexes (eta(2)-S(2)) toward dicarbomethoxyacetylene (DMA) is also discussed.     

Sizing large proteins and protein complexes by electrospray ionization mass spectrometry and ion mobility

Mass spectrometry (MS) and ion mobility with electrospray ionization (ESI) have the capability to measure and detect large noncovalent protein-ligand and protein-protein complexes. Using an ion mobility method of gas-phase electrophoretic mobility molecular analysis (GEMMA), protein particles representing a range of sizes can be separated by their electrophoretic mobility in air. Highly charged particles produced from a protein complex solution using electrospray can be manipulated to produce singly charged ions, which can be separated and quantified by their electrophoretic mobility. Results from ESI-GEMMA analysis from our laboratory and others were compared with other experimental and theoretically determined parameters, such as molecular mass and cryoelectron microscopy and X-ray crystal structure dimensions. There is a strong correlation between the electrophoretic mobility diameter determined from GEMMA analysis and the molecular mass for protein complexes up to 12 MDa, including the 93 kDa enolase dimer, the 480 kDa ferritin 24-mer complex, the 4.6 MDa cowpea chlorotic mottle virus (CCMV), and the 9 MDa MVP-vault assembly. ESI-GEMMA is used to differentiate a number of similarly sized vault complexes that are composed of different N-terminal protein tags on the MVP subunit. The average effective density of the proteins and protein complexes studied was 0.6 g/cm(3). Moreover, there is evidence that proteins and protein complexes collapse or become more compact in the gas phase in the absence of water.     

Rapid classification of perfumes by extractive electrospray ionization mass spectrometry (EESI-MS)

Extractive electrospray ionization mass spectrometry (EESI-MS) was applied to rapid fingerprinting of various perfumes for quality classification. Unique EESI-MS fingerprints of ten famous brands were obtained. This technique was shown to be applicable to rapid forgery detection on the example of an authentic and a counterfeit 'Miss Dior' fragrance by Christian Dior. We believe that the high throughput and simplicity of this sample-preparation-free method can be advantageous in the perfume industry, for instance when applied to online quality control.     

Investigation of heptakis(2,6-di-O-methyl)-beta-cyclodextrin inclusion complexes with flavonoid glycosides by electrospray ionization mass spectrometry

The non-covalent complexes between three flavonoid glycosides (quercitrin, hyperoside and rutin) and heptakis(2,6-di-O-methyl)-beta-cyclodextrin (DM-beta-CD) were investigated by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR-MS). The 1:1 complexation of each flavonoid glycoside (guest) to the DM-beta-CD (host) was monitored in the negative ion mode by mixing each guest with an up to 30-fold molar excess of the host. The binding constants for all complexes were calculated by a linear equation in the order: DM-beta-CD:quercitrin > DM-beta-CD:rutin > DM-beta-CD:hyperoside. A binding model for the complexes has also been proposed based on the binding constants and tandem mass spectrometric data of these complexes.     

Factors that affect molecular weight distribution of Suwannee river fulvic acid as determined by electrospray ionization/mass spectrometry

Effects of methylation, molar response, multiple charging, solvents, and positive and negative ionization on molecular weight distributions of aquatic fulvic acid were investigated by electrospray ionization/mass spectrometry. After preliminary analysis by positive and negative modes, samples and mixtures of standards were derivatized by methylation to minimize ionization sites and reanalyzed.Positive ionization was less effective and produced more complex spectra than negative ionization. Ionization in methanol/water produced greater response than in acetonitrile/water. Molar response varied widely for the selected free acid standards when analyzed individually and in a mixture, but after methylation this range decreased. After methylation, the number average molecular weight of the Suwannee River fulvic acid remained the same while the weight average molecular weight decreased. These differences are probably indicative of disaggregation of large aggregated ions during methylation. Since the weight average molecular weight decreased, it is likely that aggregate formation in the fulvic acid was present prior to derivatization, rather than multiple charging in the mass spectra.     

Binding constant determination of high-affinity protein-ligand complexes by electrospray ionization mass spectrometry and ligand competition

We describe an approach for the determination of binding constants for protein-ligand complexes with electrospray ionization mass spectrometry, based on the observation of unbound ligands competing for binding to a protein target. For the first time, dissociation constants lower than picomolar could be determined with good accuracy by electrospray ionization mass spectrometry. The presented methodology relies only on the determination of signal intensity ratios for free ligands in the low mass region. Therefore, all the advantages of measuring low masses with mass spectrometry, such as high resolution are preserved. By using a reference ligand with known binding affinity, the affinity of a second ligand can be determined. Since no noncovalently bound species are observed, assumptions about response factors are not necessary. The method is validated with ligands binding to avidin and applied to ligands binding to p38 mitogen-activated protein kinase.     

Characterization of non-covalent complexes of rutin with cyclodextrins by electrospray ionization tandem mass spectrometry

Electrospray ionization tandem mass spectrometry (ESI-MS(n)) and the phase solubility method were used to characterize the gas-phase and solution-phase non-covalent complexes between rutin (R) and alpha-, beta- and gamma-cyclodextrins (CDs). The direct correlation between mass spectrometric results and solution-phase behavior is thus revealed. The order of the 1 : 1 association constants (K(c)) of the complexes between R and the three CDs in solution calculated from solubility diagrams is in good agreement with the order of their relative peak intensities and relative collision-induced dissociation (CID) energies of the complexes under the same ESI-MS(n) condition in both the positive and negative ion modes. Not only the binding stoichiometry but also the relative stabilities and even binding sites of the CD-R complexes can be elucidated by ESI-MS(n). The diagnostic fragmentation of CD-R complexes, with a significant contribution of covalent fragmentation of rutin leaving the quercetin (Q) moiety attached to the CDs, provides convincing evidence for the formation of inclusion complexes between R and CDs. The diagnostic fragment ions can be partly confirmed by the complexes between Q and CDs. The gas-phase stability order of the deprotonated CD-R complexes is beta-CD-R > alpha-CD-R > gamma-CD/R; beta-CD seems to bind R more strongly than the other CDs.