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.     

Characterization of inositol phosphorylceramides from <Emphasis Type="Italic">Leishmania major</Emphasis> by tandem mass spectrometry with electrospray ionization

We describe tandem mass spectrometric approaches, including multiple stage ion-trap and source collisionally activated dissociation (CAD) tandem mass spectrometry with electrospray ionization (ESI) to characterize inositol phosphorylceramide (IPC) species seen as [M - H](-) and [M - 2H + Li](-) ions in the negative-ion mode as well as [M + H](+), [M + Li](+), and [M - H + 2Li](+) ions in the positive-ion mode. Following CAD in an ion-trap or a triple-stage quadrupole instrument, the [M - H](-) ions of IPC yielded fragment ions reflecting only the inositol and the fatty acyl substituent of the molecule. In contrast, the mass spectra from MS(3) of [M - H - Inositol](-) ions contained abundant ions that are readily applicable for assignment of the fatty acid and long-chain base (LCB) moieties. Both the product-ion spectra from MS(2) and MS(3) of the [M - 2H + Alk](-), [M + H](+), [M + Alk](+), and [M - H + 2Alk](+) ions also contained rich fragment ions informative for unambiguous assignment of the fatty acyl substituent and the LCB. However, the sensitivity of the ions observed in the forms of [M - 2H + Alk](-), [M + H](+), [M + Alk](+), and [M - H + 2Alk](+) (Alk = Li, Na) is nearly 10 times less than that observed in the [M - H](-) form. In addition to the major fragmentation pathways leading to elimination of the inositol or inositol monophosphate moiety, several structurally informative ions resulting from rearrangement processes were observed. The fragmentation processes are similar to those previously reported for ceramides. While the tandem mass spectrometric approach using MS(n) (n = 2, 3) permits the structures of the Leishmania major IPCs consisting of two isomeric structures to be unveiled in detail, tandem mass spectra from constant neutral loss scans may provide a simple method for detecting IPC in mixtures.     

N-(2-diethylamino)ethyl-4-aminobenzamide derivative for high sensitivity mass spectrometric detection and structure determination of N-linked carbohydrates

N-Linked glycans were derivatised by reductive amination using N-(2-diethylamino)ethyl-4-aminobenzamide (DEAEAB, procainamide) and examined by electrospray mass spectrometry. This derivative ionised primarily by protonation of the tertiary amine group and attachment of an alkali metal to give [M + H + X](2+) ions which were much more abundant that doubly charged ions from glycans derivatised with other aromatic amines. Fragmentation of these ions depended on the nature of the alkali metal (X). Lithium and sodium adducts fragmented to give prominent ions produced by cleavages within the DEAEAB derivative whereas the other adducts produced more abundant ions from fragmentation of the carbohydrate. Elimination of a sugar fragment, usually by cleavage adjacent to GlcNAc or sialic acid, together with a hydrogen atom, produced the most abundant singly charged fragment ions. These ions then formally fragmented by glycosidic cleavages. Potassium, rubidium and caesium adducts produced abundant losses of the alkali metal, but the resulting ions appeared not to undergo extensive fragmentation. Most fragment ions from all of the adducts were singly charged, the remainder being doubly charged. Although the spectra of the [M + X + H](2+) ions were not as informative as those from the singly charged ions from other derivatives, they, nevertheless, provided much valuable information on the structure of these glycans.     

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.     

Electrospray ionization mass spectrometric studies on the amphoteric surfactant cocamidopropylbetaine.

After liquid chromatographic (LC) separation, electrospray ionization mass spectrometry (ESI-MS) was investigated for the determination of the amphoteric surfactant cocamidopropylbetaine (CAPB). In the positive ion mode the molecule formed the adduct ions [M + H](+), [M + Na](+) and [M + K](+). Adducts of these cations were also detected with decreasing abundance as dimer and trimer clusters. Additionally, doubly charged molecular ions with different combinations of cations were identified. It was noticed that the relative abundances of individual cation adducts were not reproducible, apparently owing to varying contents of alkali metal ions originating from the solvent and the sample. Under negative ionization, the major molecular ion was [M - H](-). Higher clusters formed by two and three surfactant molecules, i.e. [2M - H](-) and [3M - H](-) were likewise registered. The tendency to form clusters in both positive and negative ion modes, even at 0.1 mg l(-1) levels, was attributed to strong electrostatic interactions between the zwitterionic head groups. Further evidence for this assumption was provided by the detection of a fragment formed from [2M - H](-) which contained the two charged head groups. Studies were undertaken in the negative ion mode on the concentration- and orifice voltage-dependent monomer, dimer and trimer formation of C(12)-CAPB in order to evaluate potential issues in using the ion [M - H](-) mode for quantitative analysis. Finally, the established (-)-LC/ESI-MS method was applied to follow up the primary degradation of CAPB in a laboratory-scale fixed-bed bioreactor (FBBR) spiked with a test concentration of 10 mg l(-1). Direct analysis without sample pretreatment revealed that higher alkyl homologues were more prone to adsorption. Primary biodegradation of all alkyl homologues was completed after a period of 4 days. Selected lyophilized FBBR samples were examined for the presence of transient or stable degradation intermediates, but no metabolite could be identified.     

Effects of transition metal ion coordination on the collision-induced dissociation of polyalanines

Transition metal-polyalanine complexes were analyzed in a high-capacity quadrupole ion trap after electrospray ionization. Polyalanines have no polar amino acid side chains to coordinate metal ions, thus allowing the effects metal ion interaction with the peptide backbone to be explored. Positive mode mass spectra produced from peptides mixed with salts of the first row transition metals Cr(III), Fe(II), Fe(III), Co(II), Ni(II), Cu(I), and Cu(II) yield singly and doubly charged metallated ions. These precursor ions undergo collision-induced dissociation (CID) to give almost exclusively metallated N-terminal product ions whose types and relative abundances depend on the identity of the transition metal. For example, Cr(III)-cationized peptides yield CID spectra that are complex and have several neutral losses, whereas Fe(III)-cationized peptides dissociate to give intense non-metallated products. The addition of Cu(II) shows the most promise for sequencing. Spectra obtained from the CID of singly and doubly charged Cu-heptaalanine ions, [M + Cu - H](+) and [M + Cu](2+) , are complimentary and together provide cleavage at every residue and no neutral losses. (This contrasts with [M + H](+) of heptaalanine, where CID does not provide backbone ions to sequence the first three residues.) Transition metal cationization produces abundant metallated a-ions by CID, unlike protonated peptides that produce primarily b- and y-ions. The prominence of metallated a-ions is interesting because they do not always form from b-ions. Tandem mass spectrometry on metallated (Met = metal) a- and b-ions indicate that [b(n) + Met - H](2+) lose CO to form [a(n) + Met - H](2+), mimicking protonated structures. In contrast, [a(n) + Met - H](2+) eliminate an amino acid residue to form [a(n-1) + Met - H](2+), which may be useful in sequencing.     

Disappearance of interfering alkali-metal adducted peaks from matrix-assisted laser desorption/ionization mass spectra of peptides with serine addition to alpha-cyano-4-hydroxycinnamic acid matrix

It has been described that ion yield in both positive- and negative-ion matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) of peptides is often inhibited by trace amounts of alkali metals and that the MALDI mass spectra are contaminated by the interfering peaks originating from traces of alkali metals, even when sample preparation is carefully performed. Addition of serine to the commonly used MALDI matrix alpha-cyano-4-hydroxycinnamic acid (CHCA) significantly improved and enhanced the signals of both protonated and deprotonated peptides, [M+H](+) and [M-H](-). The addition of serine to CHCA matrix eliminated the alkali-metal ion adducts, [M+Na](+) and [M+K](+), and the CHCA cluster ions from the mass spectra. Serine and serinephosphate as additives to CHCA enhanced and improved the formation of molecular-related ions of phosphopeptides in negative-ion MALDI mass spectra.     

Electrospray mass spectrometry and fragmentation of N-linked carbohydrates derivatized at the reducing terminus

Derivatives were prepared from N-linked glycans by reductive amination from 2-aminobenzamide, 2-aminopyridine, 3-aminoquinoline, 2-aminoacridone, 4-amino-N-(2-diethylaminoethyl)benzamide, and the methyl, ethyl, and butyl esters of 4-aminobenzoic acid. Their electrospray and collision-induced dissociation (CID) fragmentation spectra were examined with a Q-TOF mass spectrometer. The strongest signals were obtained from the [M + Na]+ ions for all derivatives except sugars derivatized with 4-amino-N-(2-diethylaminoethyl)benzamide which gave very strong doubly charged [M + H + Na]2+ ions. The strongest [M + Na]+ ion signals were obtained from the butyl ester of 4-aminobenzoic acid and the weakest from 2-aminopyridine. The most informative spectra were recorded from the [M + Li]+ or [M + Na]+ ions. These spectra were dominated by ions produced by sequence-revealing glycosidic cleavages and "internal" fragments. Linkage-revealing cross-ring cleavage ions were reasonably abundant, particularly from high-mannose glycans. Although the nature of the derivative was found to have little effect upon the fragmentation pattern, 3-aminoquinoline derivatives gave marginally more abundant cross-ring fragments than the other derivatives. [M + H]+ ions formed only glycosidic fragments with few, if any, cross-ring cleavage ions. Doubly charged molecular ions gave less informative spectra; singly charged fragments were weak, and molecular ions containing hydrogen ([M + 2H]2+ and [M + H + Na]2+) fragmented as the [M + H]+ singly charged ions with no significant cross-ring cleavages.     

Multiple lithium exchange under lithium cationization of cyclodextrins

Multiple lithium exchange is observed during electrospray ionization of alpha-, beta- and gamma-cyclodextrins from aqueous methanolic solution containing LiOH. Apart from [M + Li](+) and [M + nLi - (n - 1)H](+) ions, abundant multiply lithiated doubly charged ions corresponding to [M + nLi - (n - 2)H](2+) ions were observed. At least six lithium exchanges in alpha-cyclodextrin, seven in beta-cyclodextrin and eight in gamma-cyclodextrin were noted. The propensity of multiply lithiated doubly charged ions is much less in the open-ended maltoheptaose. It appears that during droplet or cluster formation and subsequent desolvation, LiOH trapped in the cavity of cyclodextrin reacts to form multiply lithiated ions. The singly charged [M + Li](+) and doubly charged [M + 2Li](2+) ions fragment by glycosidic cleavages, giving B series of ions, whereas the multiply lithiated ions fragment by cross ring cleavages ((2, 4)A or (O, 2)X) followed by glycosidic cleavage. From the tandem mass spectra, it appears that a maximum of two lithium exchanges occur in one sugar unit in these cyclodextrins.     

Evaluation of chiral recognition characteristics of metal and proton complexes of di-o-benzoyl-tartaric acid dibutyl ester and L-tryptophan in the gas phase

Chiral recognition of di-o-benzoyl-tartaric acid dibutyl ester (T) was achieved in the gas phase by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry. In this method two divalent transition metal cations, zinc(II) and copper(II), were used as binding metal ions, and L-tryptophan (A) was used as a chiral reference. Multimeric complexes were readily formed by electrospray ionization of a methanol:water (50:50) solution containing metal ion, L-tryptophan and T. These multimeric complexes included singly charged protonated dimeric [TAH](+), doubly charged copper(II) bound tetrameric [TACu-H](2)(2+) and doubly charged zinc(II) bound tetrameric [TAZn-H](2)(2+), together with other complexes. The mass-selected complex, i.e., [TAH](+), [TACu-H](2)(2+) and [TAZn-H](2)(2+), was used to acquire the second stage mass spectra. The chiral recognition capability of these three complexes was evaluated using the abundance ratios of daughter ion to parent ion. A high degree of chiral recognition ability was observed in [TACu-H](2)(2+) and [TAZn-H](2)(2+). It was found that the type of binding ion played an important role in the chiral recognition. Different binding ions exhibited distinctive dissociation pathways and unique chiral recognition characteristics. The present method is based not only on whole-molecule loss but also on fractional-molecule loss. In addition, the reproducibility of the chiral recognition method was confirmed by several determinations of the abundance ratios of daughter ion to parent ion with a fixed activation energy and with five different activation energies. It was also shown that this chiral recognition method can tolerate acid interference.     

Tandem mass spectra of divalent metal ion adducts of glycosyl sulfides, sulfoxides and sulfones; distinction among stereoisomers

The tandem mass spectra of the divalent metal ion (Mg2+, Ca2+, Sr2+, Ba2+, Mn2+, Ni2+, Co2+ and Zn2+) adducts of acetylated 1,2-trans-glycosyl sulfides, sulfoxides and sulfones were examined using low energy collision-induced dissociation on a Quattro II quadrupole tandem mass spectrometer. Abundant doubly charged ions, such as [3M + Met]2+ and [2M + Met]2+, were observed with alkaline earth metal chlorides. The other ions observed were [M + MetCl]+, [M + MetOAc]+, [M + MetO2SPh]+ and [2M + MetCl]+. The deprotonated metal adducts [M + Met-H]+ were seen only in the sulfones. The divalent metal ion adducts showed characteristic fragmentation pathways for the glycosyl sulfides, sulfoxides and sulfones, depending on the site of metal attachment. The doubly charged metal ion adducts dissociate to two singly charged ions, [M + MetOAc]+ and [M - OAc]+, in the sulfides and sulfoxides. In the sulfones, the adducts dissociate to [M + MetO2SPh]+ and [M - O2SPh]+. In contrast to the alkaline earth metals, which attach to the acetoxy functions, the transition metals attach to the sulfide and sulfoxide functions. The metal chloride adducts display characteristic fragmentation for the sulfides, sulfoxides and sulfones. The glucosyl, mannosyl and galactosyl sulfides, sulfoxides and sulfones could be differentiated on the basis of the stereochemically controlled MS/MS fragmentations of the metal chloride adducts.     

Observation of Na(+)-bound oligomers of quercetin in the gas phase

While developing a liquid chromatography/tandem mass spectrometry method for the analysis of the flavonoid quercitin, it was observed that quercetin (3,3',4',5,7-pentahydroxyflavone) exhibited clustering in both the positive and negative ion mode. Two series of positive ion clusters were observed; the first series corresponds to singly charged [2M + Na](+) at m/z 627.2 to [13M + Na](+) at m/z 3947.5, while the second series corresponds to doubly charged [7M + 2Na](2+) at m/z 1080.4 to [25M + 2Na](2+) at m/z 3798.5. In the negative ion mode, the behavior of quercetin parallels that of apigenin (4',5,7-trihydroxyflavone) in that [M + NO(3)](-), [2M + NO(3)](-), and [3M + NO(3)](-) were observed at m/z 364.1, 666.0, and 968.9, respectively; in addition, quercitin clusters with chloride ions ([2M + Cl](-) at m/z 638.9 and [3M + Cl](-) at m/z 940. 9) were observed. The results of tandem mass spectrometric examination of several cluster ions are reported.     

Structural determination of the novel fragmentation routes of zwitteronic morphine opiate antagonists naloxonazine and naloxone hydrochlorides using electrospray ionization tandem mass spectrometry

Electrospray ionization quadrupole time-of-flight (ESI-QqToF) mass spectra of the zwitteronic salts naloxonazine dihydrochloride 1 and naloxone hydrochloride 2, a common series of morphine opiate receptor antagonists, were recorded using different declustering potentials. The singly charged ion [M+H-2HCl](+) at m/z 651.3170 and the doubly charged ion [M+2H-2HCl](2+) at m/z 326.1700 were noted for naloxonazine dihydrochloride 1; and the singly charged ion [M+H-HCl](+) at m/z 328.1541 was observed for naloxone hydrochloride 2. Low-energy collision-induced dissociation tandem mass spectrometry (CID-MS/MS) experiments established the fragmentation routes of these compounds. In addition to the characteristic diagnostic product ions obtained, we noticed the formation of a series of radical product ions for the zwitteronic compounds 1 and 2, and also the formation of a distonic ion product formed from the singly charged ion [M+H-HCl](+) of naloxone hydrochloride 2. Confirmation of the various established fragmentation routes was effected by conducting a series of ESI-CID-QqTof-MS/MS product ion scans, which were initiated by CID in the atmospheric pressure/vacuum interface using a higher declustering potential. Deuterium labeling was also performed on the zwitteronic salts 1 and 2, in which the hydrogen atoms of the OH and NH groups were exchanged with deuterium atoms. Low-energy CID-QqTof-MS/MS product ion scans of the singly charged and doubly charged deuteriated molecules confirmed the initial fragmentation patterns proposed for the protonated molecules. Precursor ion scan analyses were also performed with a conventional quadrupole-hexapole-quadrupole tandem mass spectrometer and allowed the confirmation of the genesis of some diagnostic ions.     

Electrospray mass spectrometry and tandem mass spectrometry of bimetallic oxovanadium complexes

A series of six bimetallic oxovanadium complexes (1-6; only one was purified) were investigated by electrospray quadrupole time-of-flight tandem mass spectrometry (ESI-QTOF-MS/MS) in negative-ion mode. Radical molecular anions [M](.-) were observed in MS mode. Fragmentation patterns of [M](.-) were proposed, and elemental compositions of most of the product ions were confirmed on the basis of the high-resolution ESI-CID-MS/MS spectra. A complicated series of low-abundance product ions similar to electron impact (EI) ionization spectra indicated the radical character of the precursor ions. Fragment ions at m/z 214, 200, and 182 seem to be the characteristic ions of bimetallic oxovanadium complexes. These ions implied the presence of a V-O-V bridge bond, which might contribute to stabilization of the radical. To obtain more information for structural elucidation, three representative bimetallic oxovanadium complexes (1-3) were analyzed further by MS in positive-ion mode. Positive-ion ESI-MS produced adduct ions of [M + H](+), [M + Na](+), and [M + K](+). The fragmentation patterns of [M + Na](+) were different than those of radical molecular anions [M](.-). Relatively simple fragmentation occurred for [M + Na](+), possibly due to even-electron ion character. Negative-ion MS and MS/MS spectra of the hydrolysis product of Complex 1 supported these finding, in particular, the existence of a V-O-V bridge bond.     

Isomer differentiation by combining gas chromatography,selective self-ion/molecule reactions and tandem mass spectrometry in an ion trap mass spectrometer

This study presents a novel, simple and rapid procedure for isomer differentiation by combining gas chromatography (GC), a selective self-ion/molecule reaction (SSIMR) and tandem mass spectrometry (MS/MS) in an ion trap mass spectrometer (ITMS). SSIMR product ions were produced from four isomers. For aniline, SSIMR induces the formation of the molecular ion, [M+H](+), [M+CH](+), adduct ions of fragments ([M+F](+), where F represents fragment ions) and [2M-H](+). 2 and 3-Picoline produce [M+H](+), [2M-H](+) and [M+F](+), while 5-hexynenitrile produces [M+H](+), [M+F](+) and [2M+H](+) ions. The proposed method provides a relatively easy, rapid and efficient means of isomer differentiation via a SSIMR in the ITMS. Typically, isomer differentiation can be achieved within several minutes. The superiority of the SSIMR technique for isomer differentiation over electronic ionization (EI) is also demonstrated.     

Structural determination of hexadecanoic lysophosphatidylcholine regioisomers by fast atom bombardment tandem mass spectrometry

The structural determination of sn-1 and sn-2 hexadecanoic lysophosphatidylcholine (LPC) regioisomers was carried out using fast atom bombardment tandem mass spectrometry (FAB-MS/MS). The collision-induced dissociation (CID) of protonated and sodiated molecules produced diverse product ions due mainly to charge remote fragmentations. Based on the information obtained from the CID spectra of protonated and sodiated molecules, sn-1 and sn-2 hexadecanoic LPC isomers could be discriminated. Especially, the abundance ratio of the diagnostic ion pair [m/z 224/226] in the CID spectra of [M + H](+) ions was shown to be greatly different. Moreover, the CID-MS/MS spectra of sodium-adducted molecules for hexadecanoic LPC isomers showed characteristic product ions such as [M + Na - 103](+), [M + Na - 85](+), and [M + Na - 59](+), by which their regio-specificity can be differentiated.     

Characterization of peptides by liquid chromatography/electrospray ionization mass spectrometry using silver nitrate as a post-column complexant

Two model peptides, des-Arg1-bradykinin (DAB) and bradykinin (B), were cationized by Ag+ after their separation by reversed-phase liquid chromatography (RPLC) prior to mass spectrometry (MS). Silver nitrate solution was used as a post-column reagent. The RPLC and MS experimental conditions were optimized using flow injection in order to obtain sufficiently abundant silver adducts to permit MS/MS experiments. The use of water-methanol with 0.1% formic acid as mobile phase allowed a good chromatographic separation of the two peptides with a polymeric stationary phase and sufficiently abundant silver-containing adducts, [M + Ag + H]2+ and [M + 2Ag]2+. The gas-phase dissociation of [DAB + Ag + H]2+ and [DAB + 2Ag]2+ led to interpretable mass spectra during the on-line cationization experiment. Most of the ions obtained by dissociating [DAB + Ag + H]2+ and [DAB + 2Ag]2+ species are silver-containing ions but the ions produced depend on the parent. The ions coming from the dissociation of the doubly charged silver adducts [DAB + Ag + H]2+ or [DAB + 2Ag]2+ are of interest compared with those coming from the singly charged silver species or doubly charged protonated species. The fragmentation of the doubly charged silver adducts provides ions over the entire mass range. Although the presence of several prolines in des-Arg1-bradykinin prevents the formation of some expected ions, the observation of triplets [an-H + Ag]+, [bn-H + Ag]+ and [bn + OH + Ag]+ produced by the dissociation of on-line Ag(+)-cationized peptides could contribute to greater success of automatic sequencing of peptides.     

Structural differentiation of uronosyl substitution patterns in acidic heteroxylans by electrospray tandem mass spectrometry

The structures of two oligomers of acidic xylo-oligosaccharides (XOS) of the same molecular weight (634 Da), Xyl(2)MeGlcAHex and Xyl(2)GlcA(2) were differentiated by electrospray tandem mass spectrometry (ESI-MS/MS). These oligomers were present in a mixture of XOS obtained by acid hydrolysis of heteroxylans extracted from Eucalyptus globulus wood (Xyl(2)MeGlcAHex) and Olea europaea olive fruit (Xyl(2)GlcA(2)). In the ESI-MS spectra of the XOS, ions at m/z 657 and 652 were observed and assigned to [M + Na](+) and [M + NH(4)](+), respectively. The ESI-MS/MS spectrum of [M + Na](+) ion of Xyl(2)MeGlcAHex showed the loss of Hex residue from the reducing end followed by the loss of MeGlcA moiety. Simultaneously, the loss of a Xyl residue from either the reducing or the non-reducing ends was detected. On the other hand, the fragmentation of Xyl(2)GlcA(2) occurs mainly by the loss of one and two GlcA residues or by the loss of the GlcAXyl moiety, due to the glycosidic bond cleavage between the two Xyl residues. Loss of one and two CO(2) molecules was only observed for this oligomer, where the GlcA are in vicinal Xyl residues. The ESI-MS/MS spectra of [M + NH(4)](+) of both oligomers showed the loss of NH(3), resulting in the protonated molecule, where the presence of ions assigned as protonated molecules of aldobiuronic acid residues, [MeGlcA - Xyl + H](+) and [GlcA - Xyl + H](+), are diagnostic ions of the presence of MeGlcA and GlcA moieties in XOS. Since these structures occur in small amounts in complex acidic XOS mixtures and are very difficult, if possible, to isolate, tandem mass spectrometry revealed to be a powerful tool for the characterization of these types of substitution patterns present in heteroxylans.     

Electrospray ionization mass spectrometry of tribenzyltin substituted-phenoxyacetate compounds

Hydrolysis products of organotin compounds RC(6)H(4)OCH(2)COOSn(CH(2)ph)(3) (R = o-NO(2), 1; m-NO(2), 2; p-NO(2), 3; o-CH(3), 4; o-OCH(3), 5; o-Cl, 6; o-Br, 7) and RC(6)H(3)OCH(2)COOSn(CH(2)ph)(3) (R = o,o-2CH(3), 8, o-OCH(3), p-CHO, 9; o,p-2Cl, 10), produced in aqueous acetonitrile solution, have been investigated by electrospray mass spectrometry (MS) and MS(n) techniques. The complexes [Y(2)SnXR'](-), [Y(3)SnXR'](-), [Y(3)SnX(2)R'](-), [Y(2)SnX(3)R'](-), and fragment ions of [Y(3)SnR'](-), plus abundant RC(6)H(4)(or RC(6)H(3))OCH(2)COO(-) and RC(6)H(4)(or RC(6)H(3))O(-) ions are observed in negative mode, whereas the protonated molecular ion [M + H](+), complexes [Y(2)SnXR'](+), [Y(3)SnXR'](+), [Y(2)SnX(2)R'](+), [Y(3)SnX(2)R'](+), [Y(2)SnX(3)R'](+), [Y(3)SnX(3)R'](+), as well as [YSnXR'](+), [M - CH(2)ph](+), XSn(+), (phCH(2))(3)Sn(+), phCH(2)Sn(+) (Y = &bond;CH(2)ph, X = &bond;OOCCH(2)OC(6)H(4)R(or C(6)H(3)R)) are detected in the positive mode. Water adduct ions are seen in both modes. The assignments are facilitated by agreement between observed and calculated isotopic patterns and tandem mass spectrometry studies.     

Mass spectra of doubly charged ions

The mass spectra of biological molecules, whose molecular mass exceeds 10 kDa, invariably contain multiply charged ions. For example, a survey scan of a small protein will produce singly, doubly and triply protonated molecules, the intensity of the doubly charged species often being greater than that of the singly charged entity. Although the spectra resulting from doubly charged peptides have not previously been studied, collisional activation of such doubly charged species may result in significant additional information pertaining to molecular structure. The techniques employed to study ions originating from multiply charged species were linked scanning of constant B/E and tandem mass spectrometry, namely low collision energy spectra acquired on a BEQQ hybrid instrument. The methodology was applied to model compounds whose tandem mass spectrometry characteristics are well known, e.g. gramicidin S and angiotensin I. The results for the product ions of the [M + 2H]²⁺ species of the models were obtained which highlight the methodology required for high-mass materials.