Structural characterization of alachlor complexes with transition metal ions by electrospray ionization tandem mass spectrometry

Electrospray ionization mass (ESI-MS) spectrometry was used to investigate the nature of metal complexes of alachlor and their dissociations on activation. Ions of the first row transition metal series were employed to react with alachlor and the products were subjected to collision-induced dissociation (CID) for further structural characterization. The formation of diverse complex ions including doubly charged metal/alachlor complexes; [3L + M]²⁺ and [4L + M]²⁺ (L: alachlor and M: transition metal ions) were observed depending on the experimental conditions including the tube lens offset voltage (TLOV) and relative concentrations of alachlor and transition metal ions. It is clear that complexation with transition metal ions alters the reactive site of alachlor, promoting the loss of chlorine over the loss of CH₃OH that is the major reaction pathway in uncomplexed system. Direct elimination of chlorine from alachlor molecule was confirmed by the use of MnBr₂ instead of MnCl₂. These evidences clearly illustrate the catalytic activities of the metal ions through insertion mechanism. The function of transition metal ions in complexation was emphasized comparing the fragmentation patterns with those of protonated molecule. A change in the oxidation state of copper from + 2 to + 1 during the dissociation of metal complex was observed in company with elimination of radicals which is specific for the copper complex ions.     

Electrospray ionization with aluminum foil: A versatile mass spectrometric technique

In this study, we developed a novel electrospray ionization (ESI) technique based on household aluminum foil (Al foil) and demonstated the desirable features and applications of this technique. Al foil can be readily cut and folded into desired configuration for effective ionization and for holding sample solution in bulk to allowing acquisition of durable ion signals. The present technique was demonstrated to be applicable in analysis of a wide variety of samples, ranging from pure chemical and biological compounds, e.g., organic compounds and proteins, to complex samples in liquid, semi-solid, and solid states, e.g., beverages, skincare cream, and herbal medicines. The inert, hydrophobic and impermeable surface of Al foil allows convenient and effective on-target extraction of solid samples and on-target sample clean-up, i.e., removal of salts and detergents from proteins and peptides, extending ESI device from usually only for sample loading and ionization to including sample processing. Moreover, Al foil is an excellent heat-conductor and highly heat-tolerant, permitting direct monitoring of thermal reactions, e.g., thermal denaturation of proteins. Overall, the present study showed that Al-foil ESI could be an economical and versatile method that allows a wide range of applications.     

A capillary electrophoresis system with dual capacitively coupled contactless conductivity detection and electrospray ionization tandem mass spectrometry

A commercial system that is comprised of a CE coupled to an ESI triple quadrupole mass spectrometer was equipped with two capacitively coupled contactless conductivity detectors (C⁴Ds). The first C⁴D was positioned inside the original cartridge, and the second C⁴D was positioned as close as possible to the ESI probe entrance by using a 3D‐printed support. The C⁴Ds electropherograms were matched to the ESI‐MS electropherogram by correcting their timescales by the factor L_T/L_D, where L_T and L_D are the total capillary length and the length until the C⁴D, respectively. A general approach for method development supporting the simultaneous conductivity and MS detection is discussed, while application examples are introduced. These examples include the use of C⁴D as a simple device that dismiss the use of an EOF marker, a low‐selectivity detector that continuously provide information about unexpected features of the sample, and even a detector that can be more sensitive than ESI‐MS. The C⁴D used in this setup proved to have a smaller contribution to the peak broadening than ESI‐MS, which allowed that a C⁴D, positioned at 12 cm from the inlet of an 80‐cm‐long capillary, could be used to foresee position and shape of the peaks being formed 6.8 times slower at the ESI‐MS electropherogram.     

Electrospray ionization mass spectrometry of ionic liquids and determination of their solubility in water

Electrospray ionization mass spectrometry is used to detect both the cations (C⁺) and the anions (A^–) of ionic liquids (CA). In this study, the ionic liquids are diluted with aqueous methanol before injection. In addition to the main peaks of the parent ions, fragmentation products are observed upon increasing the cone voltage, whereas aggregates of the parent ion with one or more ionic liquid molecules (e.g., C(CA)_n⁺, A(CA)_n^–) are observed upon decreasing the cone voltage. The ions of several ionic liquids in a mixture are also detected and the ratios of their concentrations estimated. A method is developed to determine quantitatively the concentration of an ionic liquid in solution by using the cation and anion of another ionic liquid as internal standards. By using this method, the solubilities in water at room temperature (22±1 °C) of three typical hydrophobic ionic liquids have been determined: 0.70±0.08 g L^–1 for methyltributylammonium bis(trifluoromethylsulfonyl)imide (MeBu₃NNTf₂), 6.0±0.5 g L^–1 for butylmethylpyrrolidinium bis(trifluoromethylsulfonyl)imide (BuMePyrNTf₂), and 18.6±0.7 g L^–1 for 1-butyl-3-methylimidazolium hexafluorophosphate (BMIPF₆).     

Electrospray ionization multistage tandem mass spectrometry of penta- and hexa-substituted aryloxycyclotriphosphazenes

Several penta- and hexa-substituted aryloxycyclotriphosphazenes were synthesized and investigated by electrospray ionization tandem mass spectrometry (ESI-MSⁿ). Their main fragmentation pathways are proposed based on the MSⁿ and accurate mass data. An apparent hydrolysis reaction is an important fragmentation process exhibited in the ESI-MS/MS spectra for all of them. Also interesting is the intramolecular electrocyclic ring closure observed in ESI-MS/MS spectra of them. These observations may have some potential applications in the distinction between the mass spectra of penta- and hexa-substituted hexachlorocyclotriphosphazene derivatives.     

Simple and selective determination of arsenite and arsenate by electrospray ionization mass spectrometry

Arsenic pollution of public water supplies has been reported in various regions of the world. Recently, some cancer patients are treated with arsenite (As^III); most Japanese people consume seafoods containing large amounts of negligibly toxic arsenic compounds. Some of these arsenic species are metabolized, but some remain intact. For the determination of toxic As^III, a simple, rapid and sensitive method has been developed using electrospray ionization mass spectrometry (ESI-MS). As^III was reacted with a chelating agent, pyrrolidinedithiocarbamate (PDC, C₄H₈NCSS^-) and tripyrrolidinedithiocarbamate-arsine, As(PDC)₃, extracted with methyl isobutyl ketone (MIBK). A 1 μL aliquot of MIBK layer was directly injected into ESI-MS instrument without chromatographic separation, and was detected within 1 min. Arsenate (As^V) was reduced to As^III with thiosulfate, and then the total inorganic As was quantified as As^III. This method was validated for the analysis of urine samples. The limit of detection of As was 0.22 μg L⁻¹ using 10 μL of sample solution, and it is far below the permissible limit of As in drinking water, 10 μg L⁻¹, recommended by the WHO. Results were obtained in < 10 min with a linear calibration range of 1-100 μg L⁻¹. Several organic arsenic compounds in urine did not interfere with As^III detection, and the inorganic As in the reference materials SRM 2670a and 1643e were quantified after the reduction of As^V to As^III.     

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.     

Investigation of the interaction between arsenic species and thiols via electrospray ionization tandem mass spectrometry

Arsenic species have been known to participate in a number of chemical and biological reactions, including oxidation-reduction reactions, acid-base reactions, covalent interactions, and methylation-demethylation reactions because of the element's multiple and interconvertible oxidation states. Little is known about the structure or bonding behavior between arsenic species and thiolcontaining biomolecules. Therefore, a better understanding of the bonding behavior and detailed information on the molecular structure for arsenic-thiol complexes is needed. As a result, we have investigated the interaction between arsenic species (arsenate (As^V), arsenite (As^III), monomethylarsonic acid (MMA^V), and dimethylarsinic acid (DMA^V)) with biomolecules containing thiol groups (glutathione and cysteine) by electrospray ionization mass spectrometry (ESI-MS). These compounds were dissolved in methanol/water solution and introduced into the MS instrument in order to elucidate the direct bonding behavior of thiol group of biomolecules with arsenic species. In addition, further detailed structural information on this complex was obtained by collision-induced dissociation (CID) measurements.In each mass spectrum for mixture solutions between arsenic species and thiol compounds, various peaks such as protonated arsenic-thiol complexes, protonated noncomplexed thiol compounds, sodium bound cluster ions, and proton bound cluster ions were observed. In these mass spectra, the arsenic complexes were formed by interaction with thiol groups on the cysteine residues. These arsenic-thiol complexes produced a variety of fragment ions by cleavage of chemical bonds, and by interaction of other binding site on thiol compounds in tandem mass spectrometry experiments.     

Electrospray ionization mass spectrometry of ginsenosides

Ginsenosides R(b1), R(b2), R(c), R(d), R(e), R(f), R(g1), R(g2) and F(11) were studied systematically by electrospray ionization mass spectrometry in positive- and negative-ion modes with a mobile-phase additive, ammonium acetate. In general, ion sensitivities for the ginsenosides were greater in the negative-ion mode, but more structural information on the ginsenosides was obtained in the positive-ion mode. [M + H](+), [M + NH(4)](+), [M + Na](+) and [M + K](+) ions were observed for all of the ginsenosides studied, with the exception of R(f) and F(11), for which [M + NH(4)](+) ions were not observed. The signal intensities of [M + H](+), [M + NH(4)](+), [M + Na](+) and [M + K](+) ions varied with the cone voltage. The highest signal intensities for [M + H](+) and [M + NH(4)](+) ions were obtained at low cone voltage (15-30 V), whereas those for [M + Na](+) and [M + K](+) ions were obtained at relatively high cone voltage (70-90 V). Collision-induced dissociation yielded characteristic positively charged fragment ions at m/z 407, 425 and 443 for (20S)-protopanaxadiol, m/z 405, 423 and 441 for (20S)-protopanaxatriol and m/z 421, 439, 457 and 475 for (24R)-pseudoginsenoside F(11). Ginsenoside types were identified by these characteristic ions and the charged saccharide groups. Glycosidic bond cleavage and elimination of H(2)O were the two major fragmentation pathways observed in the product ion mass spectra of [M + H](+) and [M + NH(4)](+). In the product ion mass spectra of [M - H](-), the major fragmentation route observed was glycosidic bond cleavage. Adduct ions [M + 2AcO + Na](-), [M + AcO](-), [M - CH(2)O + AcO](-), [M + 2AcO](2-), [M - H + AcO](2-) and [M - 2H](2-) were observed at low cone voltage (15-30 V) only.     

Direct analysis of herbal powders by pipette-tip electrospray ionization mass spectrometry

Conventional electrospray ionization mass spectrometry (ESI-MS) is widely used for analysis of solution samples. The development of solid-substrate ESI-MS allows direct ionization analysis of bulky solid samples. In this study, we developed pipette-tip ESI-MS, a technique that combines pipette tips with syringe and syringe pump, for direct analysis of herbal powders, another common form of samples. We demonstrated that various herbal powder samples, including herbal medicines and food samples, could be readily online extracted and analyzed using this technique. Various powder samples, such as Rhizoma coptidis, lotus plumule, great burdock achene, black pepper, Panax ginseng, roasted coffee beans, Fructus Schisandrae Chinensis and Fructus Schisandrae Sphenantherae, were analyzed using pipette-tip ESI-MS and quality mass spectra with stable and durable signals could be obtained. Both positive and negative ion modes were attempted and various compounds including amino acids, oligosaccharides, glycosides, alkaloids, organic acids, ginosensides, flavonoids and lignans could be detected. Principal component analysis (PCA) based on the acquired mass spectra allowed rapid differentiation of closely related herbal species.     

啤酒中单糖的衍生化HPLC-ESI-MS测定方法研究

单糖类样品在溶液中非常稳定,难于离子化,不适合于进行ESI-MS检测。采用1-苯基-3-甲基-5-吡唑啉酮(PMP)将糖类物质衍生化,HPLC-ESI-MS在线联用,选择性离子扫描方式对几种啤酒样品中的5种单糖进行了分离检测。检出限可达到80pg。     

Ambient desorption ionization mass spectrometry

Ambient desorption ionization mass spectrometry (MS) allows for the direct analysis of ordinary objects in the open atmosphere of the laboratory or in their natural environment. Analyte desorption usually accompanies the ionization step and these processes are often concerted, multi-step processes. Ambient desorption ionization methods typically require little or no sample preparation, offer a much simplified work flow and deliver unprecedented ease of use to MS analyses.

Since the introduction of desorption electrospray ionization (DESI [Z. Takats, J.M. Wiseman, B. Gologan, R.G. Cooks, Science (Washington, D. C.) 306 (2004) 471]) in 2004 and the direct analysis in real time (DART [R.B. Cody, J.A. Laramee, H.D. Durst, Anal. Chem. 77 (2005) 2297]) in 2005, this new field of MS has developed rapidly. Numerous permutations of the various options for analyte desorption and ionization have been demonstrated. Desorption steps, such as momentum transfer, dissolution into ricocheting droplets and thermal desorption, have been combined with ionization steps, including ESI, atmospheric pressure chemical ionization and photo-ionization. The large number of possible combinations of desorption and ionization components that have already been applied is creating a proliferation of techniques and acronyms that is becoming ever more complex.

Here, we provide a logical framework for the classification of these related experiments, based on the desorption and ionization processes involved in each.     

Electrospray ionization mass spectrometry of terrestrial humic substances and their size fractions

Electrospray ionization mass spectrometry (ESI-MS) was used to evaluate the average molecular mass of terrestrial humic substances, such as humic (HA) and fulvic (FA) acids from a soil, and humic acid from a lignite (NDL). Their ESI mass spectra, by direct infusion, gave average molecular masses comparable to those previously obtained for aquatic humic materials. The soil HA and FA were further separated in size-fractions by preparative high performance size exclusion chromatography (HPSEC) and analyzed with ESI-MS by both direct infusion and a further on-line analytical HPSEC. Unexpectedly, their average molecular mass was only slightly less than for the bulk sample and, despite different nominal molecular size, did not substantially vary among size-fractions. The values increased significantly (up to around 1200 Da) after on-line analytical HPSEC for the HA bulk sample, at both pH 8 and 4, and for the HA size-fractions when pH was reduced from 8 to 4. It was noticed that HA size-fractions at pH 8 were separated by on-line HPSEC in further peaks showing average masses which progressively increased with elution volume. Furthermore, when the HA and NDL bulk samples were sequentially ultracentrifuged at increasing rotational speed, their supernatants showed mass values which were larger than bulk samples and increased with rotational speed. These variations in mass values indicate that the electrospray ionization is dependent on the composition of the humic molecular mixtures and increases when their heterogeneity is progressively reduced. It is suggested that the dominance of hydrophobic compounds in humic supramolecular associations may inhibit the electrospray ionization of hydrophilic components. Our results show that ESI-MS is reasonably applicable to humic substances only after an extensive reduction of their chemical complexity.     

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.     

Electrospray ionization mass spectrometry of metal complexes. Gas phase formation of a binuclear copper(II)–5-Br-PADAP complex

The complexes formed from copper(II) and 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol (5-Br-PADAP or HL) in aqueous methanol solution was studied by electrospray ionization mass spectrometry. The solution of a 1:1 complex of Cu(II) with 5-Br-PADAP showed five peaks assignable to a binuclear complex [Cu₂L₂(AcO)]⁺ and mononuclear complexes [CuL]⁺, [CuL(H₂O)]⁺, [CuL(AcOH)]⁺ and [CuL(HL)]⁺ (AcO=acetate). Collision activated dissociation revealed the relative order of bonding strengths; Cu–L>Cu–HL>CuL–AcOH>CuL–H₂O. The peak intensities of the binuclear complex showed second-order dependency on those of the mono complex. As for the solution of Ni(II)–5-Br-PADAP, no binuclear complex was observed in the mass spectra. Thus, it was suggested that [Cu₂L₂(AcO)]⁺ was formed by the fast gas phase reaction: 2[CuL]⁺+AcO⁻[Cu₂L₂(AcO)]⁺.     

Improved design for high resolution electrospray ionization ion mobility spectrometry

An improved design for high resolution electrospray ionization ion mobility spectrometry (ESI-IMS) was developed by making some salient modifications to the IMS cell and its performance was investigated. To enhance desolvation of electrospray droplets at high sample flow rates in this new design, volume of the desolvation region was decreased by reducing its diameter and the entrance position of the desolvation gas was shifted to the end of the desolvation region (near the ion gate). In addition, the ESI source (both needle and counter electrode) was positioned outside of the heating oven of the IMS. This modification made it possible to use the instrument at higher temperatures, and preventing needle clogging in the electrospray process. The ion mobility spectra of different chemical compounds were obtained. The resolving power and resolution of the instrument were increased by about 15-30% relative to previous design. In this work, the baseline separation of the two adjacent ion peaks of morphine and those of codeine was achieved for the first time with resolutions of 1.5 and 1.3, respectively. These four ion peaks were well separated from each other using carbon dioxide (CO₂) rather than nitrogen as the drift gas. Finally, the analytical parameters obtained for ethion, metalaxyl, and tributylamine indicated the high performance of the instrument for quantitative analysis.     

Studies on the non-covalent complexes between oleanolic acid and cyclodextrins using electrospray ionization tandem mass spectrometry

Non-covalent inclusion complexes formed between an anti-inflammatory drug, oleanolic acid (OA), and alpha-, beta- and gamma-cyclodextrins (CDs) were investigated by means of solubility studies and electrospray ionization tandem mass spectrometry (ESI-MS(n)). The order of calculated association constants (K(1 : 1)) of complexes between OA and different CDs in solution is in good agreement with the order of their relative peak intensities and the relative CID energies of the complexes under the same ESI-MS(n) conditions. These results indicate a direct correlation between the behaviors of solution- and gas-phase complexes. ESI-MS can thus be used to evaluate solution-phase non-covalent complexes successfully. The experimental results show that the most stable 1 : 1 inclusion complexes between three CDs and OA can be formed, but 2 : 1 CD-OA complexes can be formed with beta- and gamma-CDs. Multi-component complexes of alpha-CD-OA-beta-CD (1 : 1 : 1), alpha-CD-OA-gamma-CD (1 : 1 : 1) and beta-CD-OA-gamma-CD (1 : 1 : 1) were found in equimolar CD mixtures with excess OA. The formation of 2 : 1 and multi-component 1 : 1 : 1 non-covalent CD-OA complexes indicates that beta- and gamma-CD are able to form sandwich-type inclusion non-covalent complexes with OA. The above results can be partly supported by the relative sizes of OA and CD cavities by molecular modeling calculations. All the complexes allow the detection of gaseous deprotonated CD-OA complexes in the negative ion mode at high abundances. The relative stabilities of the CDs-OA inclusion complexes in the gas phase can be evaluated from the relative CID energies in the ion trap (alpha-CD-OA < beta-CD-OA < gamma-CD-OA) in the negative ion mode.     

Accurate determination of allelic frequencies in mitochondrial DNA mixtures by electrospray ionization time-of-flight mass spectrometry

The mitochondrial locus 16519T/C was used as a model for the evaluation of the benefits of ion-pair reversed-phase high-performance liquid chromatography on-line hyphenated to electrospray ionization time-of-flight mass spectrometry (ICEMS assay) for the determination of allelic frequencies of single nucleotide polymorphisms. This marker has gained interest in forensic science owing to its ability to increase the discrimination power of mitochondrial DNA testing as a consequence of its high variability across various populations. In a first set of experiments, artificial mitochondrial DNA mixtures prepared from all four theoretically possible 16519 alleles served as samples. Any allele occurring at a frequency of as low as 1–5% was unequivocally detectable irrespective of the kind of allelic mixture. Measured and expected allelic frequencies correlated well following correction of observed experimental bias, which was most probably attributable to differential PCR amplification and/or preferential ionization. For thirteen different T/C mixtures with C contents in the range 1.0–99.0%, an average error of 1.2% and a maximum error of 2.2% were observed. Furthermore, ICEMS was applied to the quantitative genotyping of eight selected individuals of which four were heteroplasmic with C contents in the range 1.9–34.1%. To check the reliability of these results, allelic proportions were additionally determined by a cloning assay. The results of the two assays correlated well (R ²=0.9971). In all cases, deviations were obtained that were smaller than 5.4%. The overall observed assay performance suggests that the described mass spectrometric technique represents one of the most powerful assays for the determination of allelic frequencies available today.      

Electrospray mass spectrometry analysis of dendritic branches bearing peripheral fullerene subunits

The electrospray mass spectrometric characterization of neutral dendrons with a carboxylic acid function or a t-butyl ester moiety at the central point and up to eight peripheral C₆₀ subunits has been performed and is described in detail. Molecules bearing a carboxylic acid group at the center turned out to be preferentially ionized by deprotonation, whereas those with a t-butyl ester head group were ionized by reduction of the C₆₀ units in the infusion capillary of the electrospray source. Electronic supplementary material Supplementary material is available for this article at and is accessible for authorized users.     

Effects of an arginine-selective tagging procedure on the fragmentation behavior of peptides studied by electrospray ionization tandem mass spectrometry (ESI-MS/MS)

The presence of arginine as the naturally occurring amino acid with the highest gas-phase basicity strongly influences the fragmentation behavior of peptides undergoing collision-induced dissociation. Using a derivatization procedure recently developed in our group, based on a reversible reaction of the guanidino group with 2,3-butanedione and an arylboronic acid, we examined how this label affects the fragmentation patterns of labeled versus unlabeled peptides in MS/MS experiments. As part of this fundamental study, two groups of model peptides (angiotensins and bradykinins) as well as tryptic peptides were labeled according to our protocol and subjected to collision-induced dissociation (CID) in both a triple quadrupole and a quadrupole ion trap instrument. It was found that for angiotensins containing an AspArg sequence, C-terminal cleavage at Asp that occurs for native peptides was completely inhibited in Arg-labeled peptides. For bradykinins and peptides obtained from tryptic digests of standard proteins, some sample peptides were little affected by the tagging of arginine residues. Others, in contrast, exhibited an almost total loss of nonspecific backbone cleavage and their fragment ion spectra were dominated by loss of the arginine tag. These and other experimental results are discussed in view of the nature of the arginine tag and the concept of proton mobility.