Differentiation of aminomethyl corrole isomers by mass spectrometry.

Two new isomeric aminomethyl corrole derivatives of [5,10,15-tris(pentafluorophenyl)corrolato]gallium(III) were synthesized with pyridine (py) molecules as axial ligands. When investigated by electrospray ionization mass spectrometry, in the positive and the negative ion modes, these compounds showed an unusual gas-phase behavior that could be used for their differentiation. In the positive ion mode, the differentiation was achieved through the formation of diagnostic fragment ions formed from [M-py + H](+) precursors, by (CH(3) )(2) NH and HF losses. An unusual addition of water to the main fragment ions provides an alternative route for isomer identification. Semi-empirical calculations were performed to elucidate the structures and stabilities of the main ionic species formed in the positive ion mode. In the negative ion mode isomer discrimination is accomplished via the fragmentation of the methoxide adduct ions [M-py + CH(3) O](-) through (CH(3) )(2) N(.) and HF losses.

Reduction and adduct formation from electrosprayed solutions of porphyrin salts.

The solutions of four meso-tetrakis(N-alkylpyridinium-4-yl)porphyrin salts and of the p-toluenesulfonate salt of meso-tetrakis(4-trimethylammoniumphenyl)porphyrin, in methanol, were studied by electrospray mass spectrometry, in order to investigate the influence of the type of counter ion, the length of the substituent N-alkyl groups of the four (N-alkylpyridinium-4-yl)porphyrins and the presence of an aromatic (alkylpyridinium) or aliphatic (trimethylammonium) nitrogen, in ion formation. In our experimental conditions, adducts with the counter ions were formed only for the meso-tetrakis(4-trimethylammoniumphenyl)porphyrin and were not observed for the other porphyrins, even when the counter ion was the same. In contrast, formation of reduced species, such as the [M(4+) + e(-)]3+, [M(4+) + 2e(-)]2+, [M(4+) + 4e(-) + 2H(+)]2+, and [M(4+) + 5e(-) + 2H(+)]+ ions was observed only for the (N-alkylpyridinium-4-yl)porphyrins and the appearance of these species is apparently solvent related and may occur via counter ion/solvent adducts.

Radical peroxidation of palmitoyl-lineloyl-glycerophosphocholine liposomes: Identification of long-chain oxidised products by liquid chromatography-tandem mass spectrometry.

Liquid chromatography coupled with electrospray tandem mass spectrometry (LC-MS/MS) was used to identify palmitoyl-lineloyl-glycerophosphatidylcholine oxidation products (PL(O(1-6))PC). Structural and positional isomers of keto, hydroxy and/or epoxy, and hydroperoxide derivatives of PLPC were identified based on MS/MS data, namely product ions attributed to lyso-phosphatidylcholines, product ions formed by loss of nH(2)O and H(2)O(2) from [MH](+) ions groups, and product ions involving the hydroxy groups, providing information about the position of these groups and of the double bonds along the carbon chain of lineloyl moiety.

Identification of free radicals of glycerophosphatidylcholines containing omega-6 fatty acids using spin trapping coupled with tandem mass spectrometry.

Metal-catalysed radical oxidation of diacyl-glycerophosphatidylcholines (GPC) with omega-6 acyl polyunsaturated fatty acids (PAPC, palmitoyl-arachidonoyl-glycerophosphatidylcholine and PLPC, palmitoyl-lineloyl-glycerophosphatidylcholine) was studied. Free radical oxidation products were trapped by spin trapping with 5,5-dimethyl-1-pyrrolidine-N-oxide (DMPO) and identified by electrospray mass spectrometry (ES-MS). The spin adducts of oxidised GPC containing one and two oxygen atoms and one and two DMPO molecules were observed as doubly charged ions. Structural characterisation by tandem mass spectrometry (MS/MS) of these ions revealed product ions corresponding to loss of the acyl chains (sn-1-palmitoyl and sn-2-oxidised spin adduct of lineloyl or arachidonoyl), loss of the spin trap (DMPO) and product ions attributed to oxidised sn-2 fatty acid spin adduct (lineloyl and arachidonoyl). Product ions formed by homolytic cleavages near the spin trap and also from 1,4 hydrogen elimination cleavages involving the hydroxy group in the sn-2 fatty acid spin adduct allowed to infer the nature of the radical. Altogether, the presence of GPC hydroxy-alkyl/DMPO and hydroxy-alkoxyl/DMPO spin adducts was proposed.

Study by liquid secondary ion and electrospray mass spectrometry of synthesized and formed-in-source metallocorroles.

Liquid secondary ion and electrospray mass spectrometry were used to study the complexation in-source of 5,10,15-tris(pentafluorophenyl)corrole with several divalent transition-metal ions. The metallocorrole ions formed in-source were identified by comparing their product ion mass spectra with the spectra of the same ions formed from metallocorroles obtained from classical procedures. Positive metallocorrole ion formation is accompanied by oxidation of the metal centre. Mechanisms were proposed for the oxidation processes, and data from negative-ion spectra reinforced these mechanisms.

Characterization of isomeric cationic porphyrins with beta-pyrrolic substituents by electrospray mass spectrometry: the singular behavior of a potential virus photoinactivator.

Electrospray ionization mass spectrometry (ESI-MS) and tandem mass spectrometry (ESI-MS/MS) have been used to differentiate the 2- and 4-methylpyridyl isomers of free-base and metallated cationic beta-vinylpyridylporphyrins. The analysis by ESI-MS/MS of the deuterated analogs and semiempirical calculations of structural and electronic parameters were also undertaken. The two free-base isomers are easily differentiated by ESI-MS/MS but the presence of a metallic center renders differentiation of the metallated isomers less effective. The data acquired show that of all the studied compounds, the free-base 2-methylpyridyl isomer, which was operative in the in vitro photoinactivation of Herpes simples virus, has a different gas-phase behavior. Local distortion of the macrocycle due to the presence of the beta-vinylpyridyl substituent occurs for all the compounds, but a different electron density distribution can account for the observed gas-phase behavior of this potential virus photoinactivator.

Identification of linoleic acid free radicals and other breakdown products using spin trapping with liquid chromatography-electrospray tandem mass spectrometry.

Linoleic acid radical products formed by radical reaction (Fenton conditions) were trapped using 5,5-dimethyl-1-pyrrolidine-N-oxide (DMPO) and analysed by reversed-phase liquid chromatography coupled to electrospray mass spectrometry (LC-MS). The linoleic acid radical species detected as DMPO spin adducts comprised oxidized linoleic acid and short-chain radical species that resulted from the breakdown of carbon and oxygen centred radicals. Based on the m/z values, the short-chain products were identified as alkyl and carboxylic acid DMPO radical adducts that exhibited different elution times. The ions identified as DMPO radical adducts were studied by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The LC-MS/MS spectra of linoleic acid DMPO radical adducts exhibited the fragment ion at m/z 114 and/or the loss of neutral molecule of 113 Da (DMPO) or 131 Da (DMPO + H2O), indicated to be DMPO adducts. The short-chain products identified allowed inference of the radical oxidation along the linoleic acid chain by abstraction of hydrogen atoms in carbon atoms ranging from C-8 to C-14. Other ions containing the fragment ion at m/z 114 in the LC-MS/MS spectra were attributed to DMPO adducts of unsaturated aldehydes, hydroxy-aldehydes and oxocarboxylic acids. The identification of aldehydic products formed by radical oxidation of linoleic acid peroxidation products, as short-chain product DMPO adducts, is a means of identifying lipid peroxidation products.

Electrospray tandem mass spectrometry of underivatised acetylated xylo-oligosaccharides.

Acetylated neutral (Xyl(n)Ac(m)) and acidic xylo-oligosaccharides (Xyl(n)Ac(m)MeGlcA, and Xyl(n)Ac(m)MeGlcAHex) obtained by partial acid hydrolysis of Eucalyptus globulus wood glucuronoxylans and fractionated by preparative ligand exchange/size-exclusion chromatography were identified by electrospray ionisation mass spectrometry (ESI-MS). Low molecular weight acetylated xylo-oligosaccharides were studied by ESI-tandem mass spectrometry (MS/MS). All the acetylated xylo-oligosaccharides showed an abundant ion due to the neutral loss of 60 Da (CH(3)CO(2)H) in the MS/MS spectra. The presence of diacetylated xylo-oligosaccharides was confirmed by the ions formed by loss of two molecules of acetic acid. Furthermore, characteristic [Xyl(res)Ac(2)+Na](+) and [XylAc(2)+Na](+) ions, and ions due to loss of XylAc(2), indicate that both acetyl groups are located in the same Xyl residue. On the other hand, losses of Xyl(res)Ac and XylAc are also observed as well as [Xyl(res)Ac+Na](+) and [XylAc+Na](+) , indicating the location of both acetyl groups in different Xyl residues, in some cases even in adjacent xyloses. The MS/MS spectra of triacetylated xylo-oligosaccharides were complex due to the presence of different isobaric xylo-oligosaccharides containing the acetyl groups at different locations in the xylo-oligosaccharide backbone. In the MS/MS spectra of acidic xylo-oligosaccharides, the ion at m/z 387, [Xyl(res)AcMeGlcA+Na](+), indicates that the acetyl groups are preferentially linked to Xyl substituted with MeGlcA. However, acidic xylo-oligosaccharides with the acetyl and 4-O-methylglucuronic acid groups in different Xyl residues were also identified. In neutral and in acidic xylo-oligosaccharides several possible locations of the acetyl groups were identified, namely at terminal positions. In summary, ESI-MS/MS is shown to be a powerful tool for the characterisation of acetylated patterns in complex mixtures of oligosaccharides.

Electrospray tandem mass spectrometry of new porphyrin amino acid conjugates.

Porphyrin amino acid conjugates with one or two porphyrin units were analyzed by electrospray ionization tandem mass spectrometry (ESI-MS/MS). The ESI-MS spectra of all the porphyrins studied, obtained in positive ion mode, show the presence of the corresponding protonated molecule [M+H]+; ESI-MS spectra of diporphyrinyl compounds also show the doubly charged ions [M+2H]2+. The fragmentations of these ions induced by collision with argon were studied (ESI-MS/MS). ESI-MS/MS gives detailed structural information about the amino acids associated with the porphyrin. Cleavage of the bonds in the vicinity of the porphyrin moiety and those involving the side chain of amino acid residues gives structural information about this type of association. A fragmentation common to all derivatives corresponds to the cleavage of the phenyl-CO bond. The expected cleavage of the amide bond, that links the porphyrin to the amino acid moiety, is a minor fragmentation, which in some cases is even absent. The MS/MS spectra of the monoporphyrinyl derivatives show product ions characteristic of the amino acid linked to the porphyrin; the fragmentation also indicates when the amino acids has a terminal carboxylic group or a terminal ester group. The fragmentations of the diporphyrinyl compounds occur mainly by the cleavage of the spacer, leading, in the case of the doubly charged ions, to predominantly mono-charged ions, indicating a preferential location of the two protons in separated porphyrinic units.

Characterization of dinitroporphyrin zinc complexes by electrospray ionization tandem mass spectrometry. Unusual fragmentations of beta-(1,3-dinitroalkyl) porphyrins.

The zinc complexes of diaryl bis(p-nitrophenyl)porphyrins and beta-(1,3-dinitroalkyl)tetraphenylporphyrins were studied by electrospray ionization (ESI) tandem mass spectrometry (MS/MS). All porphyrins showed the protonated molecule under ESI conditions. The protonated molecules were induced to fragment and the corresponding ESI tandem mass spectra were analysed. Porphyrins with two p-nitrophenyl groups showed, as expected, characteristic fragmentations including either loss of one nitro group, as the major fragment of the tandem mass spectra, and loss of both nitro groups. In contrast, MS/MS of the beta-(1,3-dinitroalkyl)porphyrins provided interesting and unexpected results such as the absence (or in insignificant abundance) of the ions formed by loss of one nitro group. However, these porphyrins show an abundant fragment due to combined loss of the two nitro groups. Also, the typical beta-cleavage of the alkyl chain is not observed per se, only when combined with loss of HNO2 or *NO2. Instead, alpha-cleavage, with loss of the beta-pyrrolic substituent, is the most favourable process.

Separation of peroxidation products of diacyl-phosphatidylcholines by reversed-phase liquid chromatography-mass spectrometry.

Lipid peroxidation process has attracted much attention due to the growing evidence of its involvement in the pathogenesis of age-related diseases. The monitoring of the lipid peroxidation products in phospholipids, formed under oxidative stress conditions, may provide new markers for oxidative stress signaling and for disease states, giving new insights in the pathogenesis process. Reversed-phase liquid chromatographic method coupled to mass spectrometry was developed for the separation of oxidized glycero-phosphatidylcholine (GPC) peroxidation products formed by the Fenton reaction that mimic in vivo oxidative stress conditions. The LC-MS conditions were applied for the separation of peroxidation products of oleoyl- (POPC), lineloyl- (PLPC) and arachidonoyl-palmitoyl phosphatidylcholine (PAPC). The peroxidation products separated included products resulting from the insertion of oxygen atoms in the sn-2 chain (long-chain), and products with the sn-2 chain shortened resulting from cleavage of oxygen-centered radicals (short-chain). Among long-chain products were the keto, hydroxy, hydroperoxide and poly-hydroxy derivatives, while short-chain products included dicarboxylic acids, aldehydes and hydroxy-aldehydes. Separation of long-chain products formed in each phosphatidylcholine was observed, and the reconstructed ion chromatogram of each ion showed an increase in the number of peaks with the increase in the number of oxygen atoms inserted into the phospholipid. Separation of short-chain products took place according to the functional group present at the sn-2 moiety that allowed the elution of dicarboxylic acids distinct from aldehydes. Separation between isomeric structures that were present in short- and long-chain products was also achieved.

Tandem mass spectrometry of intact oxidation products of diacylphosphatidylcholines: evidence for the occurrence of the oxidation of the phosphocholine head and differentiation of isomers.

Three glycerophosphatidylcholine (GPC) phospholipids (oleoyl-, linoleoyl- and arachidonoylpalmitoylphosphatidylcholine) were oxidized under Fenton reaction conditions (H(2)O(2) and Fe(2+)), and the long-chain oxidation products were detected by electrospray mass spectrometry (ES-MS) and characterized by ES-MS/MS. The intact oxidation products resulted from the insertion of oxygen atoms into the phospholipid structure. The tandem mass spectra of the [MNa](+) molecular ion showed, apart from the characteristic fragments of GPC, fragment ions resulting from neutral losses from [MNa](+), and combined with loss of 59 and 183 Da from [MNa](+). These ions resulted from cleavage of the bond near the hydroxy group by a charge-remote fragmentation mechanism, allowing its location to be pinpointed. The fragments thus formed reflected the positions of the double bonds and of the derivatives along the unsaturated fatty acid chain, giving very useful information, as they allowed the presence of structural isomers and positional isomers to be established. The identification of the fragment ion at m/z 163, which is 16 Da higher than the five-membered cyclophosphane ion (m/z 147), in some tandem mass spectra, is consistent with the oxidation of the phosphocholine head. Some ions were found to occur with the same m/z value; in two of the phospholipids and based on the MS/MS data, structural and positional isomers were differentiated. Our findings indicate that MS/MS is a valuable tool for the identification of the wide complexity of structural features occurring in oxidized phosphatidylcholines during lipid peroxidation in cellular membranes.

Fragmentation study of short-chain products derived from oxidation of diacylphosphatidylcholines by electrospray tandem mass spectrometry: identification of novel short-chain products.

Lineloyl-palmitoyl (PLPC) and arachidonoyl-palmitoyl (PAPC) phosphatidylcholine were oxidized under Fenton reaction conditions (H2O2 and Fe2+), and the short-chain products formed were identified by electrospray ionization mass spectrometry (ESI-MS). The short-chain products resulted from beta-cleavage of oxygen-centered radicals and comprised aldehydes, hydroxyaldehydes and dicarboxylic acids that yielded both [MH]+ and [MNa]+ ions. The fragmentation of the [MH]+ and [MNa]+ ions of the peroxidation products was studied by tandem mass spectrometry (MS/MS). The MS/MS spectra of both ions showed ions resulting from characteristic losses of glycerophosphatidylcholine. Other product ions, resulting from C-C cleavages occurring in the vicinity of the functional group, and fragmentations involving the hydroxy groups, were the most informative since they allowed us to obtain structural information relating to the sn-2 acyl residue. Both fragmentation pathways are due to charge-remote fragmentation occurring by a 1,4-hydrogen elimination mechanism and/or by homolytic cleavage. Furthermore, the fragmentation pathway of some ions observed in the ESI-MS spectrum was not consistent with the fragmentation behavior expected for some of the short-chain species identified in the literature and allowed the reassignment of the ions as different structures. Isobaric ions were observed in the ESI-MS spectra of both oxidized phospholipids, and were differentiated based on distinct fragmentation. The detailed knowledge of lipid peroxidation degradation products is of major importance and should be very valuable in providing new markers for oxidative stress signaling and for disease states monitoring.

Analysis of salivary peptides using HPLC-electrospray mass spectrometry.

Salivary peptides are involved in a wide range of functions constituting the first line of defence of oral cavity and precursors of dental pellicle formation. The presence of mucins in saliva makes difficult the analysis of the proteic content. This is due mainly to aggregation phenomenon between mucins and other high molecular weight glycoproteins and salivary proteins. Considering the importance of salivary peptides in biological functions, we have evaluated the influence of four different extraction methodologies on the separation and identification of these proteins by HPLC-MS. Based on their molecular weight, we identified a total of 22 peptides when extraction was performed using a solution of guanidine (6 m), compared with 14 peptides identified when saliva is acidified with TFA, which is an often used procedure. Our results also show the presence of mucin bind peptides, which include statherin, PRP1, PRP3, Histatin 1 and Histatin 5.

Identification by electrospray tandem mass spectrometry of spin-trapped free radicals from oxidized 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine.

GPC radical species formed during oxidation of a glycerophosphocholine (16:0/18:1) under the Fenton reaction conditions were detected using a spin trap, 5,5-dimethyl-1-pyrrolidine N-oxide (DMPO). The stable spin-trapped radical adducts were identified by mass spectrometry (MS) using electrospray (ES) as ionization method and characterized by tandem mass spectrometry (MS/MS). Radical adducts of oxidized free sn-2 fatty acid and of oxidized intact GPC, containing one, two and three additional oxygen atoms, were assigned. DMPO adducts of oxidized intact GPC were observed as singly and doubly charged ions in ES-MS, while adducts of oxidized free fatty acids were observed as singly charged ions. Oxidized free sn-2 fatty acids and intact GPC-DMPO adducts correspond to carbon- and oxygen-centered radicals that were identified by MS/MS as alkyl, hydroxy-alkyl, alkoxyl, hydroxy-alkoxyl, peroxyl and hydroperoxide-alkoxyl spin adducts. The DMPO molecule was attached predominantly at C(9) of the oleic chain. The fragmentation pathway of spin adducts with two DMPO molecules strongly suggests the presence of species that were simultaneously carbon- and oxygen-centered radicals. Several fragments identified are consistent with the presence of isomeric structures contributing to the same ions.

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.

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.

Detection and characterization by mass spectrometry of radical adducts produced by linoleic acid oxidation.

The formation of linoleic acid radical species under the oxidative conditions of the Fenton reaction (using hydrogen peroxide and Fe (II)) was monitored by FAB-MS and ES-MS using the spin trap 5,5-dimethyl-1-pyrrolidine-N-oxide, DMPO. Both the FAB and ES mass spectra were very similar and showed the presence of ions corresponding to carbon- and oxygen centered spin adducts (DMPO/L*, DMPO/LO*, and DMPO/LOO*). Cyclic structures, formed between the DMPO oxygen and the neighboring carbon of the fatty acid, were also observed. Electrospray tandem mass spectrometry of these ions was performed to confirm the proposed structure of these adducts. All MS/MS spectra showed an ion at m/z 114, correspondent to the [DMPO + H]+, and a fragment ion due to loss of DMPO (loss of 113 Da), confirming that they are DMPO adducts. ES-MS/MS spectra of alkoxyl radical adducts (DMPO/LO*) showed an additional ion at m/z 130 [DMPO - O + H]+, while ES MS/MS of peroxyl radical adducts (DMPO/LOO*) showed a fragment ion at m/z 146 [DMPO - OO + H]+, confirming both structures. Other fragment ions were observed, such as alkyl acylium radical ions, formed by cleavage of the alkyl chain after loss of water and the DMPO molecule. The identification of fragment ions observed in the MS/MS spectra of the different DMPO adducts suggests the occurrence of structural isomers containing the DMPO moiety both at C9 and C13. The use of ES tandem mass spectrometry, associated with spin trapping experiments, has been shown to be a valuable tool for the structural characterization of carbon and oxygen-centered spin adducts of lipid radicals.

Positive and negative electrospray ionisation tandem mass spectrometry as a tool for structural characterisation of acid released oligosaccharides from olive pulp glucuronoxylans.

Xylo-oligosaccharides with degrees of polymerisation 5-13, formed by partial acid hydrolysis from an extract representative of olive pulp glucuronoxylans (GX), were analysed by electrospray ionisation mass spectrometry (ESI-MS), both in positive and negative modes. The positive spectrum showed the presence of xylo-oligosaccharides in the mass range between m/z 500 and 1500 corresponding to singly [M+Na](+) charged ions of neutral (Xyl(7-9)) and acidic xylo-oligosaccharides (Xyl(5-9)MeGlcA), and doubly [M+2Na](2+) charged ions of Xyl(9-13) and Xyl(7-11)MeGlcA. Ammonium adducts [M+NH(4)](+) were also observed for Xyl(5-9)MeGlcA. The negative spectra showed the contribution of ions in the mass range between m/z 600 and 1400, ascribed to the deprotonated molecules [M-H](-) of Xyl(3-9)MeGlcA. Tandem mass spectrometry (MS/MS) of the major ions observed in the MS spectra was performed. The MS/MS spectra of the [M+Na](+) adducts showed the loss of MeGlcA residues as the major fragmentation pathway and glycosidic fragment ions of Xyl(n) and Xyl(n)MeGlcA structures. The MS/MS spectra of the [M+NH(4)](+) adducts suggests the occurrence of isomers of Xyl(5-9)MeGlcA oligosaccharides with the MeGlcA residue at the reducing end and at the non-reducing end of the molecules, although other structural isomers can also occur. Both glycosidic bond and cross-ring cleavages in the MS/MS spectra of the [M-H](-) ion suggest the occurrence of Xyl(3-9)MeGlcA with the substituting group at the reducing end position of the xylose backbone, as the main fragmentation ions. The results obtained by ESI-MS/MS, both in positive and negative modes, of Xyl(7-13)- and Xyl(5-11)MeGlcA, allow to identify fragmentation patterns of the structural isomers with MeGlcA linked to the terminal xylosyl residues of the oligosaccharides. The occurrence of these higher molecular weight oligosaccharides with a low substitution pattern allows to infer a scatter and random distribution of MeGlcA along the xylan backbone of olive pulp.

Synthesis and analysis of aminochromes by HPLC-photodiode array. Adrenochrome evaluation in rat blood.

The catecholamine oxidation process induces cardiotoxicity and neurotoxicity. Catecholamines can oxidize to aminochromes through autoxidation or by enzymatic or non-enzymatic catalysis. Although some toxic effects seem to be related to the formation of aminochromes there is still scarce information concerning the identification and evaluation of these compounds in in vivo models. In this study five catecholamines were oxidized to their respective aminochromes: adrenaline/adrenochrome; noradrenaline/noradrenochrome; dopa/dopachrome; dopamine/dopaminochrome; and isoproterenol/isoprenochrome. The evaluation of the catecholamines oxidation profile was performed by HPLC with photodiode array detection and using either enzymatic (tyrosinase) or non-enzymatic [Ag(2)O, CuSO(4), NaIO(4) and K(3)Fe(CN)(6)] catalytic systems. The NaIO(4) was found to be the most efficient oxidant of catecholamines. An isocratic reverse-phase HPLC method was developed to analyse each pair of catecholamine-aminochrome. The analytical system was then applied to the detection of adrenochrome in rat blood at 490 nm. Thus, adrenochrome was administered i.p. to rats and its concentration in whole blood was monitored after 5, 15 and 25 min. Blood treatment for adrenochrome evaluation consists of an acidification for protein precipitation followed by a rapid neutralization. The results showed a rapid decrease of adrenochrome concentration in blood after its administration. The adrenochrome present in blood was characterized by UV and tandem mass spectrometry.