Characterization of Diacylglycerylphosphocholine Molecular Species by FAB-CAD-MS/MS: A General Method Not Sensitive to the Nature of the Fatty Acyl Groups

The use of negative ion mode fast-atom bombardment-collision-activated dissociation-tandem mass spectrometry (FAB-CAD-MS/MS) for diacylglycerylphosphocholine molecular species determinations was investigated for 24 naturally occurring and synthetic compounds. The previously proposed method of selecting [M-15]^? as the parent ion and using the relative abundance of the carboxylate daughter ions to distinguish the positions of esterification was found to be unreliable in cases where the fatty acyl group at sn1 was much larger than that at sn2. The predicted greater abundance of the sn2 carboxylate daughter, relative to the sn1 carboxylate daughter, was also violated when polyunsaturated fatty acyl groups were esterifred at sn2. In addition, several marginal cases were found where the ratio of intensities of the sn2/sn1 carboxylate daughters followed the expected pattern (sn2 > sn1) initially, but reversed over extended scanning time. The use of an alternative FAB-CAD-MS/MS method is proposed where the [M-B6]^? ion is selected as the precursor and the relative intensities of the daughters resulting from loss of the free fatty acids at snl and sn2 are determined. In every case examined to date, the ion formed by loss of the free acid from the sn2 position was always more abundant. Because the parent ion is equivalent to the phosphatidic acid ion, this technique should be equally applicable to all other phospholipid classes where this fragment ion is present in the spectrum.     

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.     

UV photodissociation dynamics of deprotonated 2'-deoxyadenosine 5'-monophosphate [5'-dAMP-H]-

The UV photodissociation dynamics of deprotonated 2'-deoxyadenosine 5'-monophosphate ([5'-dAMP-H](-)) has been studied using a unique technique based on the coincident detection of the ion and the neutral fragments. The observed fragment ions are m/z 79 (PO(3)(-)), 97 (H(2)PO(4)(-)), 134 ([A-H](-)), 177 ([dAMP-H-A-H(2)O](-)), and 195 ([dAMP-H-A](-)), where "A" refers to a neutral adenine molecule. The relative abundances are comparable to that found in previous studies on [5'-dAMP-H](-) employing different excitation processes, i.e., collisions and UV photons. The fragmentation times of the major channels have been measured, and are all found to be on the microsecond time scale. The fragmentation mechanisms for all channels have been characterized using velocity correlation plots of the ion and neutral fragment(s). The findings show that none of the dissociation channels of [5'-dAMP-H](-) is UV specific and all proceed via statistical fragmentation on the ground state after internal conversion, a result similar to fragmentations induced by collisions.     

Electrospray ionization/tandem quadrupole mass spectrometric studies on phosphatidylcholines: the fragmentation processes

We applied low-energy collisionally activated dissociation (CAD) tandem quadrupole mass spectrometry to study the fragmentation pathways of the [M + H](+) and [M + Li](+) ions of phosphatidylcholine (PC), generated by electrospray ionization (ESI). It is revealed that the fragmentation pathways leading to loss of the polar head group and of the fatty acid substituents do not involve the hydrogens attached to the glycerol backbone as previously reported. The pathway for formation of the major ion of m/z 184 by loss of the polar head group from the [M + H](+) precursor of a diacyl PC involves the participation of the alpha-hydrogen of the fatty acyl substituents, whereas the H(+) participates in the loss of fatty acid moieties. The alpha-hydrogens of the fatty acid substituents also participate in the major fragmentation processes, including formation of [M + Li-R(x)CO(2)H](+) and [M + Li-59-R(x)CO(2)H](+) ions for the [M + Li](+) ions of diacyl PCs, when subjected to low-energy CAD. These fragmentation processes are deterred by substitution of the fatty acyl moieties with alkyl, alkenyl, or hydroxyl groups and consequentially, result in a distinct product-ion spectrum for various PC, including diacyl-, plasmanyl- plasmenyl-, and lyso-PC isomers. The alpha-hydrogens of the fatty acyl substituents at sn-2 are more labile than those at sn-1. This is reflected by the preferential loss of the R(1)CO(2)H over the R(2)CO(2)H observed for the [M + Li](+) ions of diacyl PCs. The spectrum features resulting from the preferential losses permit identification and assignment of the fatty acid moieties in the glycerol backbone. The new fragmentation pathways established by tandem and source CAD tandem mass spectra of various PC molecules, including deuterium-labeling analogs, were proposed. These pathways would clarify the mechanisms underlying the ion formations that lead to the structural characterization of PC molecules.     

Liquid chromatography/tandem mass spectrometry analysis of long-chain oxidation products of cardiolipin induced by the hydroxyl radical

The anionic phospholipid cardiolipin (CL) is found almost exclusively in the inner membrane of mitochondria, playing an important role in energy metabolism. Oxidation of CL has been associated with apoptotic events and various pathologies. In this study, electrospray ionization mass spectrometry coupled with liquid chromatography (LC/ESI-MS) was used to identify tetralinoleoyl-cardiolipin (TLCL) modifications induced by the OH(·) radical generated under Fenton reaction conditions (H(2)O(2) and Fe(2+)). The identified oxidation products of TLCL contained 2, 4, 6 and 8 additional oxygen atoms. These long-chain oxidation products were characterized by LC/ESI-MS/MS as doubly [M-2H](2-) and singly charged [M-H](-) ions. A detailed analysis of the fragmentation pathways of these precursor ions allowed the identification of hydroperoxy derivatives of CL. MS/MS analysis indicated that CL oxidation products with 4, 6 and 8 oxygen atoms have one fatty acyl chain bearing 4 oxygen atoms ([RCOO+4O](-)). Even when the TLCL molecule was oxidized by the addition of eight oxygen atoms, one of the acyl chains remained non-modified and one fatty acyl chain contained three or four oxygen atoms. This led us to conclude that under oxidative conditions by the OH(·) radical, the distribution of oxygens/peroxy groups in the CL molecule is not random, even when CL has the same fatty acyl chains in all the positions. Using mass spectrometry, the oxidation products have been unequivocally assigned, which may be useful for their detection in biological samples.     

Characterization of cardiolipin as the sodiated ions by positive-ion electrospray ionization with multiple stage quadrupole ion-trap mass spectrometry

The application of multiple-stage ion-trap (IT) mass spectrometric methods for the structural characterization of cardiolipin (CL), a 1,3-bisphosphatidyl-sn-glycerol that consists of four fatty acyl chains and three glycerol backbones (designated as A, B, and central glycerol, respectively), as the sodiated adduct ions in the positive-ion mode was evaluated. Following collisionally activated dissociation (CAD), the [M - 2H + 3Na]+ ions of CL yield two prominent fragment ion pairs that consist of the phosphatidyl moieties attached to the 1'- and 3'-position of the central glycerol, respectively, resulting from the differential losses of the diacylglycerol moieties containing A and B glycerol, respectively. The results are consistent with those previously described for the [M - H]- and [M - 2H + Na]- ions in the negative-ion mode, thus permitting assignment of the two phosphatidyl moieties attached to the 1'- or 3'-position of the central glycerol. The identities of the fatty acyl substituents and their positions on the glycerol backbones (glycerol A and B) are deduced from further degradation of the above ion pairs that give the fragment ions reflecting the fatty acid substituents at the sn-1 (or sn-1') and sn-2 (or sn-2') positions. The ions that arise from losses of the fatty acid substituents at sn-1 and sn-1', respectively, are prominent, but the analogous ions from losses of the fatty acid substituents at sn-2 and sn-2', respectively, are of low abundance in the MS2 product-ion spectra. This feature further confirms the assignment of the positions of the fatty acid substituents. The similar IT multiple-stage mass spectrometric approaches including MS2 and MS3 for structural characterization of CL using its [M + Na]+ and the [M - H + 2Na]+ ions are also readily applicable. However, their uses for structural characterization are less desirable because formation of the [M + Na]+ and the [M - H + 2Na]+ ions for CL is not predictable.     

Amide bond cleavage in deprotonated tripeptides: a newly discovered pathway to "b2 ions

The fragmentation reactions of the [M-H](-) ions of the tripeptides H-Gly-Leu-Sar-OH, H-Leu-Gly-Pro-OH and H-Gly-Leu-Gly-OH have been investigated in detail using energy-resolved mass spectrometry, isotopic labelling and MS(3) experiments. It is shown that the major route to the "b(2) ions involves loss of a neutral amine from the a(3) ([M-H-CO(2)](-)) ion rather than being formed directly by fragmentation of the [M-H](-) ion. When there is no C-terminal amidic hydrogen (Sar, Pro), loss of a neutral amine is the dominant primary fragmentation reaction of the a(3) ion. However, when there is a C-terminal amidic hydrogen (Gly), elimination of the N-terminal amino acid residue is the major fragmentation reaction of the a(3) ion and formation of the "b(2) ion is greatly reduced in importance. It is proposed that the "b(2) ions are deprotonated oxazolones.     

Synthesis, reactivity, electrochemical behavior, and crystal structure of a family of multivalent metal carbido-carbonyl clusters based on the rh(10)(c)(2)au(4-6) framework

Six metal carbido-carbonyl clusters have been isolated and recognized as members of a multivalent family based on the dioctahedral Rh(10)(C)(2) frame, with variable numbers of CO ligands, AuPPh(3) moieties, and anionic charge: [Rh(10)(C)(2)(CO)(x)(AuPPh(3))(y)](n-) (x = 18, 20; y = 4, 5, 6; n = 0, 1, 2). Anions [Rh(10)(C)(2)(CO)(18)(AuPPh(3))(4)](-) ([2](-)) and [Rh(10)(C)(2)(CO)(18)(AuPPh(3))(4)](2-) ([2](2-)) have been obtained by the reduction of [Rh(10)(C)(2)(CO)(18)(AuPPh(3))(4)] (2) under N(2), while [Rh(10)(C)(2)(CO)(18)(AuPPh(3))(5)](-) ([3](-)) was obtained from [Rh(10)(C)(2)(CO)(20)(AuPPh(3))(4)] (1) by reduction under a CO atmosphere. [3](-) can be better obtained by the addition of AuPPh(3)Cl to [2](2-). [Rh(10)(C)(2)(CO)(18)(AuPPh(3))(6)] (4) is obtained from [3](-) and 2 as well by the reduction and subsequent addition of AuPPh(3)Cl. The molecular structures of [2](2-) ([NBu(4)](+) salt), [3](-) ([NMe(4)](+) salt), and 4 have been determined by single-crystal X-ray diffraction. The redox activities of complexes 1, 2 and [3](-) have been investigated by electrochemical and electron paramagnetic resonance (EPR) techniques. The data from EPR spectroscopy have been accounted for by theoretical calculations.     

Dinuclear amine-thiophenolate complexes coligated by ferrocenemonocarboxylate and 1,1'-ferrocenedicarboxylate anions: preparation, characterization and structures of trinuclear [LMII 2(O2CC5H4FeCp)]+ and Pentanuclear [(LMII 2)2(O2CC5H4)2Fe]2+ complexes (M=Co, Ni, Zn)

A series of novel tri- and pentanuclear complexes composed of dinuclear LM(2) units (M=Co, Ni, Zn; L=24-membered macrocyclic hexaazadithiophenolate ligand) and ferrocenecarboxylate ([CpFeC(5)H(4)CO(2)](-)) or 1,1'-ferrocenedicarboxylate ([Fe(C(5)H(4)CO(2))(2)](2-)) groups is reported. The complexes [LM(II) (2)(O(2)CC(5)H(4)FeCp)](+) (M=Co (6), Ni (7), Zn (8)) and [(LM(II) (2))(2)(O(2)CC(5)H(4))(2)Fe](2+) (M=Co (9), Ni (10)) have been prepared by substitution reactions from labile [LM(II) (2)L'](+) precursors (L'=Cl, OAc) and the respective ferrocenecarboxylate anions in methanol. Mixed-valent [(LCo(II)Co(III))(2)(O(2)CC(5)H(4))(2)Fe](4+) (11) was prepared by oxidation of 9 with bromine. Complexes 7[BPh(4)], 8[BPh(4)], 9[BPh(4)](2), 10[BPh(4)](2), and 11[ClO(4)](4) have been characterized by X-ray crystallography; showing that the ferrocenyl carboxylates act as bidentate (7, 8) or bis-bidentate (9-11) bridging ligands towards one or two bioctahedral LM(2) subunits, respectively. The structures are retained in solution as indicated by NMR spectroscopic studies on the diamagnetic Zn(2)Fe complex 8[ClO(4)]. Electrochemical studies reveal significant anodic potential shifts for the oxidation potential of the ferrocenyl moieties upon complexation and the magnitude of the potential shift appears to correlate with the charge of the LM(2) subunits. This is qualitatively explained in terms of destabilizing electrostatic (Coulomb) interactions between the M(2+) ions of the LM(2) unit and the proximate ferrocenium fragment. An analysis of the temperature-dependent magnetic susceptibility data for 10[BPh(4)](2) shows the presence of weak ferromagnetic magnetic exchange interactions between the Ni(II) ions in the LNi(2) units. The exchange coupling across the ferrocenedicarboxylate bridge is negligible.     

Hydrido phosphanido bridged polynuclear complexes obtained by protonation of a phosphinito bridged Pt(I) complex with HBF4 and HF

The protonation of the phosphinito-bridged Pt(I) complex [(PHCy(2))Pt(μ-PCy(2)){κ(2)P,O-μ-P(O)Cy(2)}Pt(PHCy(2))](Pt-Pt) (1) by aqueous HBF(4) or hydrofluoric acid leads selectively to the hydrido-bridged solvento species syn-[(PHCy(2))(H(2)O)Pt(μ-PCy(2))(μ-H)Pt(PHCy(2)){κP-P(OH)Cy(2)}](Y)(2)(Pt-Pt) ([2-H(2)O]Y(2)) {Y = BF(4), F(HF)(n)} when an excess of acid was used. On standing in halogenated solvents, complex [2-H(2)O](BF(4))(2) undergoes a slow but complete isomerization to [(PHCy(2))(2)Pt(μ-PCy(2))(μ-H)Pt{κP-P(OH)Cy(2)}(H(2)O)](BF(4))(2)(Pt-Pt) ([4-H(2)O][BF(4)](2)) having the P(OH)Cy(2) ligand trans to the hydride. The water molecule coordinated to platinum in [2-H(2)O][BF(4)](2) is readily replaced by halides, nitriles, and triphenylphosphane, and the acetonitrile complex [2-CH(3)CN][BF(4)](2) was characterized by XRD analysis. Solvento species other than aqua complexes, such as [2-acetone-d(6)](2+) or [2-CD(2)Cl(2)](2+) were obtained in solution by the reaction of excess etherate HBF(4) with 1 in the relevant solvent. The complex [2-H(2)O](Y)(2) [Y = F(HF)(n)] spontaneously isomerizes into the terminal hydrido complexes [(PHCy(2))Pt(μ-PCy(2)){κ(2)P,O-μ-P(O)Cy(2)}Pt(H)(PHCy(2))](Y)(Pt-Pt) ([6](Y)). In the presence of HF, complex [6](Y) transforms into the bis-phosphanido-bridged Pt(II) dinuclear complex [(PHCy(2))(H)Pt(μ-PCy(2))(2)Pt{κP-P(OH)Cy(2)}](Y)(Pt-Pt) ([7](Y)). When the reaction of 1 with HF was carried out with diluted hydrofluoric acid by allowing the HF to slowly diffuse into the dichloromethane solution, the main product was the linear 60e tetranuclear complex [(PHCy(2)){κP-P(O)Cy(2)}Pt(1)(μ-PCy(2))(μ-H)Pt(2)(μ-PCy(2))](2)(Pt(1)-Pt(2)) (8). Insoluble compound 8 is readily protonated by HBF(4) in dichloromethane, forming the more soluble species [(PHCy(2)){κP-P(OH)Cy(2)}Pt(1)(μ-PCy(2))(μ-H)Pt(2)(μ-PCy(2))](2)(BF(4))(2)(Pt(1)-Pt(2)) {[9][BF(4)](2)}. XRD analysis of [9][BF(4)](2)·2CH(2)Cl(2) shows that [9](2+) is comprised of four coplanar Pt atoms held together by four phosphanido and two hydrido bridges. Both XRD and NMR analyses indicate alternate intermetal distances with peripheral Pt-Pt bonds and a longer central Pt···Pt separation. DFT calculations allow tracing of the mechanistic pathways for the protonation of 1 by HBF(4) and HF and evaluation of their energetic aspects. Our results indicate that in both cases the protonation occurs through an initial proton transfer from the acid to the phosphinito oxygen, which then shuttles the incoming proton to the Pt-Pt bond. The different evolution of the reaction with HF, leading also to [6](Y) or 8, has been explained in terms of the peculiar behavior of the F(HF)(n)(-) anions and their strong basicity for n = 0 or 1.     

Direct qualitative analysis of triacylglycerols by electrospray mass spectrometry using a linear ion trap

Triacylglycerols (TAGs) isolated from a biological sample provide a challenge for mass spectrometric analysis because of the complexity of naturally occurring TAGs, which may contain different fatty acyl substituents resulting in a large number of molecular species having the identical elemental composition. We have investigated the use of mass spectrometry to obtain unambiguous information as to the individual TAG molecular species present in a complex mixture of triacylglycerols using a linear ion trap mass spectrometer. Ammonium adducts of TAGs, [M+NH4]+, were generated by electrospray ionization, which permitted the molecular weight of each TAG molecular species to be determined. The mechanisms involved in the decomposition of the [M+NH4]+ and subsequent fragment ions were investigated using deuterium labeling, MS/MS, and MS3 experiments. Collision induced decomposition of [M+NH4]+ ions resulted in the neutral loss of NH3 and an acyl side-chain (as a carboxylic acid) to generate a diacyl product ion. MS/MS data were used to identify each acyl group present for a given [M+NH4]+ ion, and this information could be combined with molecular weight data to identify possible TAG molecular species present in a biological extract. Subsequent MS3 experiments on the resultant diacyl product ions, which gave rise to acylium (RCO+) and related ions, enabled unambiguous TAG molecular assignments. These strategies of MS, MS/MS, and MS3 experiments were applied to identify components within a complex mixture of neutral lipids extracted from RAW 264.7 cells.     

Characterization of cardiolipin from Escherichia coli by electrospray ionization with multiple stage quadrupole ion-trap mass spectrometric analysis of [M−2H+Na]− ions

We report a multiple-stage ion-trap (IT) mass spectrometric approach with electrospray ionization (ESI) for structural characterization of the [M - 2H + Na]- ion of cardiolipin (CL), a 1,3-bisphosphatidyl-sn-glycerol that consists of four fatty acyl chains and three glycerol backbones designated as A, B, and central glycerol, respectively (see Scheme 1). Following collisionally activated dissociation (CAD), the [M - 2H + Na]- ions of CL yield two prominent fragment ions that arise from the differential losses of the diacylglycerol moieties containing A or B glycerol, respectively. The tentative assignment of the two phosphatidyl moieties attached to the 1'- or 3'-position of the central glycerol is based on the observation that the ions arising from loss of the diacylglycerol moiety containing glycerol B is more abundant than that containing glycerol A. The structures of the above two ions, including the identities of the fatty acyl substituents and the position of fatty acyl substituents on the glycerol backbones (glycerol A and B) are determined by MS3 experiments that give spectra comprising several sets of prominent ions informative for the structural assignment of the fatty acyl substituents on the glycerol A and glycerol B. This method permits the structures of CL in a mixture isolated from Escherichia coli, including species that consist of various isomers, to be unveiled in detail.     

An experimental and theoretical investigation of gas-phase reactions of Ca2+ with glycine

The gas-phase reactions between Ca(2+) and glycine ([Ca(gly)](2+)) have been investigated through the use of mass spectrometry techniques and B3-LYP/cc-pWCVTZ density functional theory computations. The major peaks observed in the electrospray MS/MS spectrum of [Ca(gly)](2+) correspond to the formation of the [Ca,C,O(2),H](+), NH(2)CH(2) (+), CaOH(+), and NH(2)CH(2)CO(+) fragment ions, which are produced in Coulomb explosion processes. The computed potential energy surface (PES) shows that not only are these species the most stable product ions from a thermodynamic point of view, but they may be produced with barriers lower than for competing processes. Carbon monoxide is a secondary product, derived from the unimolecular decomposition of some of the primary ions formed in the Coulomb explosions. In contrast to what is found for the reactions of Ca(2+) with urea ([Ca(urea)](2+)), minimal unimolecular losses of neutral fragments are observed for the gas-phase fragmentation processes of [Ca(gly)](2+), which is readily explained in terms of the topological differences between their respective PESs.     

Factors affecting the solid-state structure and dimensionality of mercury cyanide/chloride double salts, and NMR characterization of coordination geometries

In the reaction of organic monocationic chlorides or coordinatively saturated metal-ligand complex chlorides with linear, neutral Hg(CN)(2) building blocks, the Lewis-acidic Hg(CN)(2) moieties accept the chloride ligands to form mercury cyanide/chloride double salt anions that in several cases form infinite 1-D and 2-D arrays. Thus, [PPN][Hg(CN)(2)Cl].H(2)O (1), [(n)Bu(4)N][Hg(CN)(2)Cl].0.5 H(2)O (2), and [Ni(terpy)(2)][Hg(CN)(2)Cl](2) (4) contain [Hg(CN)(2)Cl](2)(2-) anionic dimers ([PPN]Cl = bis(triphenylphosphoranylidene)ammonium chloride, [(n)Bu(4)N]Cl = tetrabutylammonium chloride, terpy = 2,2':6',6' '-terpyridine). [Cu(en)(2)][Hg(CN)(2)Cl](2) (5) is composed of alternating 1-D chloride-bridged [Hg(CN)(2)Cl](n)(n-) ladders and cationic columns of [Cu(en)(2)](2+) (en = ethylenediamine). When [Co(en)(3)]Cl(3) is reacted with 3 equiv of Hg(CN)(2), 1-D [[Hg(CN)(2)](2)Cl](n)(n-) ribbons and [Hg(CN)(2)Cl(2)](2-) moieties are formed; both form hydrogen bonds to [Co(en)(3)](3+) cations, yielding [Co(en)(3)][Hg(CN)(2)Cl(2)][[Hg(CN)(2)](2)Cl] (6). In [Co(NH(3))(6)](2)[Hg(CN)(2)](5)Cl(6).2H(2)O (7), [Co(NH(3))(6)](3+) cations and water molecules are sandwiched between chloride-bridged 2-D anionic [[Hg(CN)(2)](5)Cl(6)](n)(6n-) layers, which contain square cavities. The presence (or absence), number, and profile of hydrogen bond donor sites of the transition metal amine ligands were observed to strongly influence the structural motif and dimensionality adopted by the anionic double salt complex anions, while cation shape and cation charge had little effect. (199)Hg chemical shift tensors and (1)J((13)C,(199)Hg) values measured in selected compounds reveal that the NMR properties are dominated by the Hg(CN)(2) moiety, with little influence from the chloride bonding characteristics. delta(iso)((13)CN) values in the isolated dimers are remarkably sensitive to the local geometry.     

Structural elucidation of the ceramide moiety of starfish gangliosides by collision-induced dissociation of the sodium ion complex.

Collision-induced dissociation (CID) spectra of sodium ion complexes ([M+Na]+ ions), produced by FAB-MS of methyl ester derivatives of ganglioside, indicate the length of the fatty acyl chain of the ceramide moieties without chemical degradation. In the case of a genuine ganglioside, only the fission of the glycosyl linkage of sialic acid was prominently observed.     

Structure characterization of lipocyclopeptide antibiotics,aspartocins A,B & C,by ESI‐MSMS and ESI‐nozzle‐skimmer‐MSMS

Three lipocyclopeptide antibiotics, aspartocins A (1), B (2), and C (3), were obtained from the aspartocin complex by HPLC separation methodology. Their structures were elucidated using previously published chemical degradation results coupled with spectroscopic studies including ESI‐MS, ESI‐Nozzle Skimmer‐MSMS and NMR. All three aspartocin compounds share the same cyclic decapeptide core of cyclo [Dab2 (Asp1‐FA)‐Pip3‐MeAsp4‐Asp5‐Gly6‐Asp7‐Gly8‐Dab9‐Val10‐Pro11]. They differ only in the fatty acid side chain moiety (FA) corresponding to (Z)‐13‐methyltetradec‐3‐ene‐carbonyl, (+,Z)‐12‐methyltetradec‐3‐ene‐carbonyl and (Z)‐12‐methyltridec‐3‐ene‐carbonyl for aspartocins A (1), B (2), and C (3), respectively. All of the sequence ions were observed by ESI‐MSMS of the doubly charged parent ions. However, a number of the sequence ions observed were of low abundance. To fully sequence the lipocyclopeptide antibiotic structures, these low abundance sequence ions together with complementary sequence ions were confirmed by ESI‐Nozzle‐Skimmer‐MSMS of the singly charged linear peptide parent fragment ions H‐Asp5‐Gly6‐Asp7‐Gly8‐Dab9‐Val10‐Pro11‐Dab2¹⁺‐Asp1‐FA. Cyclization of the aspartocins was demonstrated to occur via the β‐amino group of Dab2 from ions of moderate intensity in the ESI‐MSMS spectra. As the fatty acid moieties do not undergo internal fragmentations under the experimental ESI mass spectral conditions used, the 14 Da mass difference between the fatty acid moieties of aspartocins A (1) and B (2) versus aspartocin C (3) was used as an internal mass tag to differentiate fragment ions containing fatty acid moieties and those not containing the fatty acid moieties. The most numerous and abundant fragment ions observed in the tandem mass spectra are due to the cleavage of the tertiary nitrogen amide of the pipecolic acid residue‐3 (16 fragment ions) and the proline residue‐11 (7 fragment ions). In addition, the neutral loss of ethanimine from α,β‐diaminobutyric acid residue 9 was observed for the parent molecular ion and for 7 fragment ions. Copyright © 2009 John Wiley & Sons, Ltd.     

Structural characterization of cardiolipin by tandem quadrupole and multiple-stage quadrupole ion-trap mass spectrometry with electrospray ionization

We report negative-ion electrospray tandem mass spectrometric methods for structural characterization of cardiolipin (CL), a four-acyl-chain phospholipid containing two distinct phosphatidyl moieties, of which structural assignment of the fatty acid residues attached to the glycerol backbones performed by low-energy CAD tandem mass spectrometry has not been previously described. The low-energy MS2-spectra of the [M - H]- and [M - 2H]2- ions obtained with ion-trap or with tandem quadrupole instrument combined with ion-trap MS3-spectra or with source CAD product-ion spectra provide complete structural information for CL characterization. The MS2-spectra of the [M - H]- ions contain two sets of prominent fragment ions that comprise a phosphatidic acid, a dehydrated phosphatidylglycerol, and a (phosphatidic acid + 136) anion. The substantial differences in the abundances of the two distinct phosphatidic anions observed in the MS2-spectra of the [M -H]- ions lead to the assignment of the phosphatidyl moieties attached to the 1' or 3' position of central glycerol. Upon further collisional dissociation, the MS3-spectra of the phosphatidic anions provide information to identify the fatty acyl substituents and their position in the glycerol backbone. The MS2-spectra of the [M - 2H]2- ions obtained with TSQ or ITMS contain complementary information to confirm structural assignment. The applications of the above methods in the differentiation of cardiolipin isomers and in the identification of complex cardiolipin species consisting of multiple molecular structures are also demonstrated.     

Mass spectral behavior of the hydrolysis products of sesqui- and oxy-mustard type chemical warfare agents in atmospheric pressure chemical ionization

Bis(2-hydroxyethylthio)alkanes and bis(2-hydroxyethylthioalkyl)ethers are important biological and environmental degradation products of sulfur mustard analogs known as sesqui- and oxy-mustards. We used atmospheric pressure chemical ionization mass spectrometry (APCI MS) to acquire characteristic spectra of these compounds in positive and negative ionization modes. Positive APCI mass spectra exhibited [M + H](+); negative APCI MS generated [M + O(2)](-), [M - H](-), and [M - 3H](-); and both positive and negative APCI mass spectra contained fragment ions due to in-source collision-induced dissociation. Product ion scans confirmed the origin of fragment ions observed in single-stage MS. Although the spectra of these compounds were very similar, positive and negative APCI mass spectra of the oxy-mustard hydrolysis product, bis(2-hydroxyethylthiomethyl)ether, differed from the spectra of the other compounds in a manner that suggested a rearrangement to the sesqui-mustard hydrolysis product, bis(2-hydroxyethylthio)methane. We evaluated the [M + O(2)](-) adduct ion for quantification via liquid chromatography-MS/MS in the multiple-reaction monitoring (MRM) mode by constructing calibration curves from three precursor/product ion transitions for all the analytes. Analytical figures of merit generated from the calibration curves indicated the stability and suitability of these transitions for quantification at concentrations in the low ng/mL range. Thus, we are the first to propose a quantitative method predicated on the measurement of product ions generated from the superoxide adduct anion of the sesqui-and oxy-mustard hydrolysis products.     

Lipid composition of membranes of Escherichia coli by liquid chromatography/tandem mass spectrometry using negative electrospray ionization

A liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) method using reversed-phase chromatography was developed for the analysis of phospholipids from bacterial extracts of a wild-type strain of Escherichia coli. Product ion mass spectra from [M--H](-) precursor ions allowed an identification of individual phospholipid species that includes both fatty acid composition and fatty acyl location on the glycerol backbone using diagnostic product ions. Thus, complete assignment, including sn-1/sn-2 fatty acyl position, was achieved for this strain of E. coli. In addition, the phospholipids were quantified relative to one another using an internal standard method.     

Cation-cation complexes of pentavalent uranyl: from disproportionation intermediates to stable clusters

Three new cation-cation complexes of pentavalent uranyl, stable with respect to the disproportionation reaction, have been prepared from the reaction of the precursor [(UO(2)py(5))(KI(2)py(2))](n) (1) with the Schiff base ligands salen(2-), acacen(2-), and salophen(2-) (H(2)salen = N,N'-ethylene-bis(salicylideneimine), H(2)acacen = N,N'-ethylenebis(acetylacetoneimine), H(2)salophen = N,N'-phenylene-bis(salicylideneimine)). The preparation of stable complexes requires a careful choice of counter ions and reaction conditions. Notably the reaction of 1 with salophen(2-) in pyridine leads to immediate disproportionation, but in the presence of [18]crown-6 ([18]C-6) a stable complex forms. The solid-state structure of the four tetranuclear complexes, {[UO(2)(acacen)](4)[μ(8)-](2)[K([18]C-6)(py)](2)} (3) and {[UO(2)(acacen)](4)[μ(8)-]}?2?[K([222])(py)] (4), {[UO(2)(salophen)](4)[μ(8)-K](2)[μ(5)-KI](2)[(K([18]C-6)]}?2?[K([18]C-6)(thf)(2)]?2?I (5), and {[UO(2)(salen)(4)][μ(8)-Rb](2)[Rb([18]C-6)](2)} (9) ([222] = [222]cryptand, py = pyridine), presenting a T-shaped cation-cation interaction has been determined by X-ray crystallographic studies. NMR spectroscopic and UV/Vis studies show that the tetranuclear structure is maintained in pyridine solution for the salen and acacen complexes. Stable mononuclear complexes of pentavalent uranyl are also obtained by reduction of the hexavalent uranyl Schiff base complexes with cobaltocene in pyridine in the absence of coordinating cations. The reactivity of the complex [U(V)O(2)(salen)(py)][Cp*(2)Co] with different alkali ions demonstrates the crucial effect of coordinating cations on the stability of cation-cation complexes. The nature of the cation plays a key role in the preparation of stable cation-cation complexes. Stable tetranuclear complexes form in the presence of K(+) and Rb(+), whereas Li(+) leads to disproportionation. A new uranyl-oxo cluster was isolated from this reaction. The reaction of [U(V)O(2)(salen)(py)][Cp*(2)Co] (Cp* = pentamethylcyclopentadienyl) with its U(VI) analogue yields the oxo-functionalized dimer [UO(2)(salen)(py)](2)[Cp*(2)Co] (8). The reaction of the {[UO(2)(salen)(4)][μ(8)-K](2)[K([18]C-6)](2)} tetramer with protons leads to disproportionation to U(IV) and U(VI) species and H(2)O confirming the crucial role of the proton in the U(V) disproportionation.