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Spirulina microalga (Arthrospira platensis) is an interesting phototrophic organism because of its high content of nutrients including proteins, lipids, essential amino acids, antioxidants, vitamins, polysaccharides, and minerals. Hydrophilic interaction liquid chromatography (HILIC) coupled to linear ion trap (LIT) and Orbitrap Fourier transform mass spectrometry (FTMS) via ESI was employed for the separation and characterization of lipid species in A. platensis. Inositolphosphoceramides (IPC) are minor but important constituents of spirulina; their investigation was accomplished by HILIC-ESI-MS including collision-induced dissociation (MS2 , MS3 ) of deprotonated molecules in the LIT analyzer and a schematic fragmentation pattern is described. All four commercial spirulina samples revealed the occurrence of the same IPC species at m/z 796.6 (d18:0/16:0;1), 810.6 (d18:0/17:0;1), 824.6 (d18:0/18:0;1), and 826.6 (d18:0/17:0;2) but in diverse relative abundance. This study sets the stage for future investigations on IPC in other algae and microalgae.

RATIONALE Several bioactive compounds, including phenolic acids and secoiridoids, are transferred from olive drupes to olive oil during the first stage of production. Here, the characterization of these low molecular weight (LMW) compounds in olive oil and in closely related processing materials, like olive leaves (OL) and olive mill wastewaters (OMW), was faced up, for the first time, by matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry (TOF MS). METHODS A novel binary matrix composed of 1,8-bis(tetramethylguanidino)naphthalene (TMGN) and 9-aminoacridine (9AA) (1:1 molar ratio), displaying excellent ionization properties at low levels of laser energy, was employed in reflectron negative ion mode by a MALDI TOF/TOF system equipped with a neodymium-doped yttrium lithium fluoride (Nd:YLF) laser (345 nm). MS/MS experiments were performed by using ambient air as the collision gas. RESULTS Four major secoiridoids typically present in olive oil, i.e., the aglycones of oleuropein and ligstroside, and oleacein and olecanthal at m/z 377.1, 361.1, 319.1 and 303.1, respectively, were detected in virgin olive oil (VOO) extracts, along with some of their chemical/enzymatic hydrolysis by-products, such as elenolic (m/z 241.1), decarboxymethyl-elenolic acids (m/z 183.1) and hydroxytyrosol (m/z 153.1). Besides oleuropein aglycone and oleacein, hydroxylated derivatives of decarboxymethyl-elenolic acid and hydroxytyrosol were evidenced in OMW. CONCLUSIONS While oleuropein was confirmed in OL extracts, several interesting phenolic compounds, including hydroxytyrosol, were recognized in OMW. The proposed approach based on the use of a novel binary matrix for MALDI MS/MS analyses of LMW bioactive compounds can be considered a promising analytical tool for a rapid screening of the phenolic fraction in olive oils and related processing wastes.

The reactivity of the sulfur-containing nucleoside 4-thio-(2 '-deoxy)-thymidine usually abbreviated as 4-thio-thymidine, (S-4-TdR) under Fenton conditions, ie, in the presence of H2O2 and catalytic amounts of Fe(II), was investigated by UV-vis spectroscopy and electrospray ionization single and tandem mass spectrometry (ESI-MS and MS/MS). S-4-TdR hydroxylated on the S atom was found to be a key reaction intermediate, ultimately leading to (2 '-deoxy)-thymidine usually abbreviated as thymidine, (TdR) as the main reaction product. This finding was in accordance with the outcome of the reaction between S-4-TdR and H2O2, previously investigated in our laboratory. On the other hand, the additional presence of center dot OH radicals, induced by the Fe(II)/H2O2 combination, led to the increased generation of another interesting S-4-TdR product, already observed after its reaction with H2O2 alone, ie, the covalent dimer including a S-S bridge between two S-4-TdR molecules. More importantly, multihydroxylated derivatives of S-4-TdR and TdR were detected as peculiar products obtained under Fenton conditions. Among them, a product bearing an OH group both on the methyl group linked to the thymine ring and on the C5 atom of the ring was found to prevail. The results obtained during this study, integrated by those found previously in our laboratory, indicate 4-thiothymidine as a promising molecular probe for the recognition, through a careful characterization of its reaction products, of the prevailing species among reactive oxygen species (ROS) corresponding to singlet-state oxygen, hydrogen peroxide, and hydroxylic radical.

Several mobile phase additives (i.e., organic acids and their ammonium salts) were used to modulate the chromatographic retention of cyanocobalamin and its cis-diaminemonochloroplatinum(II) conjugate, depending on the specific nature of the stationary phase. Regardless of the mobile phase additive, the positively charged cyanocobalamin-cis-diaminemonochloroplatinum(II) conjugate was systematically less retained than cyanocobalamin on a conventional octadecyl-silica column. In contrast, the amide-embedded C18 column exhibited a progressive increase in the conjugate retention time upon changing the mobile phase additive from organic (acetic, formic and trifluoroacetic) acids to ammonium salts, ultimately leading to an inversion of the elution order. This change of retention was interpreted by invoking the interplay between hydrophobic interactions, hydrogen bonding between the conjugate and the polar amide groups and the ion-pairing ability of the lyophilic counterions, whereby the acetate anion was found to be the most suitable to control the solute retention.

Rationale Cisplatin (CP) is a widely used anticancer drug characterized by toxic side effects that could be alleviated using novel delivery systems including CP prodrugs. The in vitro incubation of a putative prodrug, obtained from cyanocobalamin (CNCbl) and cis-diamminemonochloroplatinum(II) (mCP), with nucleoside monophosphates (NMPs) was investigated. Methods The in vitro reactions between the putative prodrug CNCbl-mCP and the NMPs of adenosine (AMP), guanosine (GMP), cytidine (CMP) and uridine (UMP) were carried out in slightly acidic water-methanol solutions at 37°C for 24 h. Each sample was examined using reversed-phase liquid chromatography coupled with electrospray ionization in positive ion mode and tandem mass spectrometry (RPLC/ESI-MS/MS) by collision-induced dissociation in a linear ion-trap mass spectrometer. Results Seven adducts were recognized as formed by substitution reactions of the chloride ligand in planar CP. Comparison between observed and theoretical isotopic patterns together with MS/MS fragmentation pathways revealed the presence of single or multiple binding sites depending on the NMP involved. The CNCbl-mCP conjugate was found to interact with N7 or O4 atoms of GMP and UMP, respectively, generating single adducts, while two isomeric adducts were observed for CMP. Finally, AMP gave rise to three isomeric adducts. Conclusions In agreement with literature data relevant to the interaction between CP and NMPs, the most reactive nucleotides were AMP and GMP. The present RPLC/ESI-MS/MS approach is very promising for investigation of the reactions of CP conjugates with ribonucleotides not only in vitro but also in vivo.

Hydrophilic interaction liquid chromatography coupled to negative-ion electrospray linear ion-trap multiple-stage MS (HILIC-ESI-MSn , n = 2,3) was used to characterize polar lipids occurring in donkey milk. Besides the detection of abundant phospholipids, the structural characterization and content evaluation of minor glycosphingolipids (GSLs) were assessed. We report an unprecedented characterization of 11 hexosyl-ceramides (HexCer), 10 Hex2 Cer, and 4 Hex3 Cer. CID-MS/MS spectra in negative ion mode mainly afford information on the monosaccharide number and ceramide constitution (i.e., N-acyl residue and long-chain base), whereas MS/MS/MS spectra on the ceramide anions allow to recognize for each GSL the sphingoid base. The occurrence of sphingosine (S), sphinganine (DS), and phytosphingosine (P) was inferred from the fragmentation patterns. The milk samples exhibit a relatively high number of phytosphingosine substitutes, perhaps because of the feeding of donkeys, mainly based on pasture grass. However, the incidence of hydroxylated species on the α-carbon of the acyl chain was also revealed. The fatty acid composition of N-acyl chains showed high values of long-chain saturated fatty acids such as 20:0, 22:0, 23:0, and 24:0. An average content of GSL is also provided and three representative mono-, di-, and tri-HexCer in donkey milk are the following: HexCer 18:0/24:1 phytosphingosine nonhydroxylated [PN] at m/z 862.6 as chloride adduct [M+Cl]- , and content 225.9 ± 2.8 μg 100 mL-1 ; Hex2 Cer 18:0/16:0 sphinganine nonhydroxylated [DSN] at m/z 862.7 as deprotonated adduct [M-H]- , and content 70.8 ± 1.4 μg 100 mL-1 ; and Hex3 Cer 18:1/24:1 [SN] at m/z 1132.8 as [M-H]- , and content 38.5 ± 0.7 μg 100 mL-1 .

Rationale Sulfoquinovosylmonoglycerides (SQMG) and sulfoquinovosyldiglycerides (SQDG) in the lipid extracts of parsley (Petroselinum crispum) and spinach (Spinacia oleracea) leaves were investigated. The aim of this work was to assess and establish the chemical characterization of fatty acyl chains in sulfolipids (SQMG and SQDG) and their regiochemistry. Methods A key component of this approach is a combination of hydrolysis reactions catalyzed by Lecitase® Ultra, which is a sn1-regioselective hydrolase enzyme, and reversed-phase liquid chromatography with electrospray ionization and sequential mass spectrometry (RPLC-ESI-MS) by collision induced dissociation (CID)-MSn (n=2,3). Results The occurrence of SQMG bearing 16:0 or 18:3 acyl chain was established for the first time. A regiochemistry-dependent fragmentation pattern of SQMG was attained whereby the sulfoquinovosyl anion ([C6H11O8S]¯ at m/z 243.0) provides a diagnostic product ion. Regioselective enzymatic treatment also provided a posteriori confirmation of a widely-accepted fragmentation rule for SQDG. The sulfoquinovosyl anion was found to play a role also in the fragmentation pattern of SQDG, whose regiochemical assignment could be ultimately confirmed by MS3 experiments. Conclusions The predominant sulfolipid in leaf extracts of raw parsley (Petroselinum crispum) and spinach (Spinacia oleracea) was identified as SQDG 18:3/16:0, along with of SQMG 18:3/0:0 and SQMG 16:0/0:0. The present CID MS-based method can be considered a successful approach to validate the regiochemical characterization of sulfolipids paving the way for their unambiguous characterization.

Hydrophilic interaction liquid chromatography coupled to electrospray ionization-Fourier transform mass spectrometry was employed to study ceramide lipids occurring in mussels of sp. Mytilus galloprovincialis. Lipid extracts from alive mussels and mussels deliberately subjected to specific thermal treatments were analyzed. In particular, single and tandem MS measurements were performed on a hybrid quadrupole-Orbitrap mass spectrometer and then complemented by MS(n) analyses (n = 2, 3) achieved by a linear ion trap mass spectrometer. This approach enabled the characterization of 66 ceramide lipids, encompassing ceramide phosphoethanolamines (CPE), ceramide aminoethylphosphonates (CAEP) and N-monomethylated CAEP. The sphingoid and acyl chains of each ceramide lipid could be distinctly recognized in terms of numbers of carbon atoms and C=C bonds, and indications on the possible location of the latter on the sphingoid chain could be often inferred from fragmentation patterns. The occurrence of several species hydroxylated on the α carbon of the acyl chain was also discovered. On the other hand, the sphingoid chain of ceramide lipids was never found to be involved in oxidation processes, unless forced exposure of the mussel lipid extracts to atmospheric oxygen was performed. CPE(d19:3/16:0) and its hydroxylated form, CPE(d19:3/2-OH-16:0), were found to be the prevailing species among CPE, whereas CAEP(d18:2/16:0), CAEP(d19:3/16:0) and CAEP(d19:3/2-OH-16:0) were the most abundant CAEP. Finally, ceramide lipids showed a remarkably higher stability, compared with glycerophospholipids, in mussels subjected to different thermal treatments. This finding opens interesting perspectives on the role of ceramide-based lipids in the adaptation of aquatic organisms to thermal stresses. Copyright © 2016 John Wiley & Sons, Ltd.

4-Chloro-α-cyanocinnamic acid (ClCCA) is a very useful matrix able to give the protonated adduct [M+H](+) of intact cyanocobalamin (CNCbl) as the base peak (m/z 1355.58) in matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS). The only fragment observed is [M-CN + H](+•) formed through the facile (•) CN neutral loss reflecting the fairly low Co-C bond energy. All other investigated proton transfer matrices, including α-cyano-4-hydroxycinnamic acid, para-nitroaniline and 2,5-dihydroxybenzoic acid, give rise to a complete decyanation of CNCbl with concomitant formation of [M-CN + H](+•) , [M-CN + Na](+•) and [M-CN + K](+•) adducts at m/z 1329.57, 1351.55 and 1367.51, respectively. Depending on the matrix used, a variable degree of fragmentation involving the α-side axial ligand was observed. A plausible explanation of the specific behaviour of 4-chloro-α-cyanocinnamic acid as a soft matrix is discussed. Tandem mass spectra of both [M + H](+) and [M-CN + H](+•) ions were obtained and product ions successfully assigned. The possibility of detecting the protonated adduct of intact CNCbl was exploited in foodstuff samples such as cow milk and hen egg yolk by MALDI tandem MS upon sample extraction. We believe that our data provide strong basis for the application of MALDI tandem MS in the qualitative analysis of natural CNCbl, including fish, liver and meat samples. Copyright © 2016 John Wiley & Sons, Ltd.

Rationale Here hardly ionizable and low molecular weight compounds are detected in negative ion mode by using novel superbasic proton sponges based on 1,8-bisphosphazenylnaphthalene (PN) as MALDI matrices. Among the selected proton sponges, 1,8-bis(trispyrrolidinophosphazenyl)naphthalene (TPPN) has shown the best behaviour as matrix since it allows the direct detection of intact cholesterol without derivatization also in real challenging samples. Methods Very weakly acidic compounds such as sterols, steroids, fatty alcohols and saccharides were detected in reflectron negative ion mode by a MALDI TOF/TOF system equipped with a neodymium-doped yttrium lithium fluoride (Nd:YLF) laser (345 nm) with typical mass accuracy of 10 ppm. MS/MS experiments were performed by using ambient air as the collision gas. Results Contrary to traditional MALDI matrices, superbasic proton sponges allowed the easy deprotonation of an alcohol functional group without a previous chemical derivatization step. Experimental evidence indicates that analyte deprotonation is achieved in the condensed phase, i.e. PN superbasic proton sponges operate according to a recently proposed model named matrix assisted ionization/laser desorption (MAILD). A detection limit of 3 pmol/spot of cholesterol (model compound) with a signal-to-noise ratio ≥ 10 was typically obtained. Conclusions For the first time, the usefulness of novel superbasic proton sponges is demonstrated for MALDI detection of hardly ionizable compounds such as sterols, steroids, fatty alcohols and saccharides. The leading candidate TPPN has been successfully applied for negative ion MAILD-MS analysis of cholesterol, fatty acids and phospholipids in egg yolk and brain tissue extracts. Copyright © 2016 John Wiley & Sons, Ltd.

The separation efficiency of hydrophilic interaction liquid chromatography and the high resolution/accuracy of electrospray ionization-Fourier transform MS were successfully applied to the detailed characterization of lyso-phosphatidylcholines (LPCs) and lyso-phosphatidylethanolamines (LPEs) contained in the lipid extracts of Mytilus galloprovincialis (Mediterranean mussel). As a result, 57 LPCs, including regio- and positional isomers, and 45 LPEs, including acyl and plasma(e)nyl species, were identified. Four lyso-phosphonocholines were also identified among mussel Lyso-Phospholipids. To the best of our knowledge this represents the first characterization, at a molecular level, ever reported for LPEs in mussels. No significant variation was observed in the composition of both LPCs and LPEs when mussels were refrigerated at +4°C for up to 48 h, i.e. under conditions usually employed for seafood transportation and storage. Treatments mimicking more severe thermal stresses, namely eight day-refrigeration at + 4°C, two week-freezing at -15°C and 6 h-storage at 25°C, resulted in a significant increase in the molar abundance of LPCs and LPEs (expressed with respect to that of their precursors, PCs and PEs, respectively) and was accompanied by the death of all or part of the molluscs. These results were interpreted invoking the generation of lyso-phospholipids, mediated by endogenous phospholipases, as an intermediate process toward the partial replacement of side chains in phospholipids, perhaps functional to a better adaptation of mussels to adverse temperature conditions. Interestingly, the relative abundances of specific compounds belonging to the LPC and LPE classes were found to follow the seasonal variations of sea temperature.

The purpose of this work is to explore the mass fragment characterization of commonly used drugs through a novel approach, which involves isotope-selective tandem mass spectrometry (MS/MS). Collision-induced dissociation (CID) was performed with a low-resolution linear ion trap mass spectrometer in positive electrospray ionization. Three pharmacologically active ingredients, i.e. omeprazole, meloxicam and brinzolamide, selected as model compounds in their own formulation, were investigated as a sodiated adduct [C17H19N3O3S + Na]+ (omeprazole) and as protonated adducts, [C14H13N3O4S2 + H]+ and [C12H21N3O5S3 + H]+, meloxicam and brinzolamide, respectively. Selecting a narrow window of ±0.5 m/z units, precursor ion fragmentation by CID-MS/MS of isotopologues A + 0, A + 1 and A + 2 was found very useful to confirm the chemical formula of product ions, thus aiding the establishment of characteristic fragmentation pathways of all three examined compounds. The correctness of putative molecular formula of product ions was easily demonstrated by exploiting the isotope peak abundance ratios (i.e. IF+0/IF+1 and IF+0/IF+2) as simple constraints in low-resolution MS instrumentations. Copyright © 2014 John Wiley & Sons, Ltd.

A full characterization of sulfoquinovosyldiacylglycerols (SQDGs) in the lipid extract of spinach leaves has been achieved using liquid chromatography/electrospray ionization-linear quadrupole ion trap mass spectrometry (MS). Low-energy collision-induced dissociation tandem MS (MS/MS) of the deprotonated species [M − H]− was exploited for a detailed study of sulfolipid fragmentation. Losses of neutral fatty acids from the acyl side chains (i.e. [M − H − RCOOH]−) were found to prevail over ketene losses ([M − H − R'CHCO]−) or carboxylates of long-chain fatty acids ([RCOO]−), as expected for gas-phase acidity of SQDG ions. A new concerted mechanism for RCOOH elimination, based on a charge-remote fragmentation, is proposed. The preferential loss of a fatty acids molecule from the sn-1 position (i.e. [M − H − R1COOH]−) of the glycerol backbone, most likely due to kinetic control of the gas-phase fragmentation process, was exploited for the regiochemical assignment of the investigated sulfolipids. As a result, 24 SQDGs were detected and identified in the lipid extract of spinach leaves, their number and variety being unprecedented in the field of plant sulfolipids. Moreover, the prevailing presence of a palmitic acyl chain (16:0) on the glycerol sn-2 position of spinach SQDGs suggests a prokaryotic or chloroplastic path as the main route for their biosynthesis. Copyright © 2013 John Wiley & Sons, Ltd.

An liquid chromatography–mass spectrometry method using electrospray ionization in negative ion mode coupled with a hybrid quadrupole linear ion trap and Fourier transform ion cyclotron resonance (FTICR) mass spectrometer was applied to characterize of intact glucosinolates (GLSs) in crude sample extracts of wild bud flowers of Capparis spinosa (Capparis species, family Capparaceae). Structural information of GLSs was obtained upon precursor ions' isolation within the FTICR trapping cell and subsequent fragmentation induced by infrared multiphoton dissociation (IRMPD). Such a fragmentation was found very useful in terms of chemical identification of all precursor ions [M–H]− including sulfur-rich GLSs reported here for the first time. Along with most common GLSs already found in capers such as glucocapparin, isopropyl/n-propyl-GLS, mercapto-glucocapparin, and two indolic GLS, i.e., 4-hydroxyglucobrassicin and glucobrassicin, the occurrence of the uncommon glycinyl-glucocapparin as well as two sulfur-rich GLSs is reported. IRMPD showed an increased selectivity towards disulfide bond cleavages with thiol migration, suggesting the side chain structure of non-targeted compounds, i.e., disulfanyl-glucocapparin and trisulfanyl-glucocapparin. Glucocapparin [2.05 ± 0.25 mg/g, dry weight (dw)] was the most abundant GLS, followed by glucobrassicin (232 ± 18 µg/g, dw) and 4-hydroxyglucobrassicin (89 ± 12 µg/g, dw). All other compounds were present at very low content ranging from 0.5 to 1.5 µg/g dw. Copyright © 2012 John Wiley & Sons, Ltd.

Samples of Mytilus galloprovincialis were examined to investigate the levels of polybrominated diphenyl ethers (PBDEs) in the Apulia's marine environment, a region in the South of Italy. The levels of nine PBDE congeners were measured in 40 mussel samples taken from the aquaculture farms in the South of Adriatic Sea and North Ionian Sea along the Apulia coast. While accelerated solvent extraction (ASE) was the extraction technique adopted using acetone/n-hexane (1:1, v/v), the content of PBDEs was evaluated by using gas chromatography coupled with high-resolution mass spectrometry (GC-HRMS) via electron ionization (EI) in the multiple ion monitoring (MIM) mode (two ions for each compound). The 13 C mass-labeled compounds were used to establish the levels of PBDEs in M. galloprovincialis samples. The total concentration of PBDEs (ΣPBDEs) ranged from 0.2 to 6.9 ng/g dry mass, with the highest concentrations found around coastal areas of Salento. With regard to the composition of PBDE congeners, BDE-47, BDE-99 and BDE-1 00 were the predominant congeners in most of the samples. The present study is the first to report levels of PBDEs occurring in aquatic organisms living in aquaculture farms of Apulia region.

A comprehensive and highly selective method for detecting in bacterial supernatants a modified sulfur nucleoside, S-adenosyl-L-methionine (SAM), and its metabolites, i.e., S-adenosylhomocysteine (SAH), adenosine (Ado), 5'-deoxy-5'-methylthioadenosine (MTA), adenine (Ade), S-adenosyl-methioninamine (dcSAM), homocysteine (Hcy) and methionine (Met), was developed. The method is based on reversed-phase liquid chromatography with positive electrospray ionization (ESI+) coupled to a hybrid linear quadrupole ion trap (LTQ) and 7-T Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS). A gradient elution was employed with a binary solvent of 0.05 M ammonium formate at pH 4 and acetonitrile. The assay involves a simultaneous cleanup of cell-free bacterial broths by solid-phase extraction and trace enrichment of metabolites with a 50-fold concentration factor by using immobilized phenylboronic and anion-exchange cartridges. While the quantitative determination of SAM was performed using stable-isotope-labeled SAM-d3 as an internal standard, in the case of Met and Ade, Met-13C and Ade-15N2 were employed as isotope-labeled internal standards, respectively. This method enabled the identification of SAM and its metabolites in cell-free culture of Pseudomonas aeruginosa grown in Davis minimal broth (formulation without sulphur organic compounds), with routine sub-ppm mass accuracies (-0.27 +/- 0.68 ppm). The resulting contents of S(C)S(S)-SAM, S(S)-dcSAM, MTA, Ado and Met in the free-cell supernatant of P. aeruginosa was 56.4 +/- 2.1 nM, 32.2 +/- 2.2 nM, 0.91 +/- 0.10 nM, 19.6 +/- 1.2 nM and 1.93 +/- 0.02 microM (mean +/- SD, n = 4 extractions), respectively. We report also the baseline separation (Rs > or = 1.5) of both diastereoisomeric forms of SAM (S(C)S(S) and S(C)R(S)) and dcSAM (S(S) and R(S)), which can be very useful to establish the relationship between the biologically active versus the inactive species, S(C)S(S)/S(C)R(S) and S(S)/R(S) of SAM and dcSAM, respectively. An additional confirmation of SAM-related metabolites was accomplished by a systematic study of their MS/MS spectra.

Some newly synthesized 10B nido-carborane derivatives, i.e., 7,8-dicarba-nido-undecaborane monoanions ([7-Me-8-R-C2B9H10]-K+, R = H, butyl, hexyl, octyl and decyl), have been fully characterised and examined by electrospray ionization and Fourier transform ion cyclotron resonance mass spectrometry with liquid chromatographic separation (LC/ESI-FTICR-MS). These boron-containing compounds exhibit abundant molecular ions ([M]−) at m/z 140.22631 [CB9H14]−, m/z 196.28883 [CB9H22]−, m/z 224.32032 [CB9H26]−, m/z 252.35133 [CB9H30]− and m/z 280.38354 [CB9H34]− at the normal tube lens voltage setting of −90 V, which was an instrumental parameter value selected in the tuning operation. Additional [M–nH2]− (n = 1−4) ions were observed in the mass spectra when higher tube lens voltages were applied, i.e., −140 V. High-resolution FTICR-MS data revealed the accurate masses of fragment ions, bearing either an even or an odd number of electrons. Collision-induced dissociation of the [M–nH2]− ions (n = 0–4) in the quadrupole linear ion trap (LTQ) analyzer confirmed the loss of hydrogen molecules from the molecular ions. It is suggested that the loss of H2 molecules from the alkyl chain is a consequence of the stabilization effect of the nido-carborane charged polyhedral skeleton. Copyright © 2009 John Wiley & Sons, Ltd.

N-acyl-homoserine-lactones (AHSLs) are widely conserved signal molecules present in quorum sensing systems of Gram-negative bacteria such as Pseudomonas aeruginosa. We present here the results obtained with a hybrid linear trap/Fourier transform ion cyclotron resonance (LTQ-FTICR) mass spectrometer used to investigate the occurrence of AHSLs and cognate N-acyl-homoserines (AHSs) in bacterial isolates of P. aeruginosa (strain PAO1). Two hydrolysed AHSs were found in significant amounts, most likely formed through the lactone opening of N-3-oxo-decanoyl-L-homoserine-lactone (3OC10-HSL) and N-3-oxo-dodecanoyl-L-homoserine-lactone (3OC12-HSL). Structure elucidation of these ring-opened molecules, i.e. N-3-oxo-decanoyl-L-homoserine (3OC10-HS), and N-3-oxo-dodecanoyl-L-homoserine (3OC12-HS), which are not detected by bacterial biosensors, was performed by high-resolution and accurate mass measurements upon liquid chromatography (LC) and confirmed by tandem MS in the LTQ analyser. Assignment of chemical formula, with mass spectra in the form of [M + H]+, was significantly expedited by extracted ion chromatograms (XICs) because the number of potentially plausible formulae for each protonated signalling molecule was considerably reduced a priori by the LC behaviour, the high mass measurement accuracy available in FTICR mass spectra and the isotopic patterns. At least two concentration levels were observed in spent culture supernatants of P. aeruginosa: compounds at a relatively high content (5–15 µM) that is C4-HSL, 3OC10-HS, and 3OC12-HS and those occurring at a lower content (

A method for the comprehensive profiling of the N-acyl-homoserine lactone (AHL) family of bacterial quorum-sensing molecules is presented using liquid chromatography (LC) coupled to a hybrid quadrupole linear ion trap (LTQ) and Fourier-transform ion-cyclotron-resonance mass spectrometer (FTICR). We demonstrate an increase in signal intensity in MS with electrospray ionization (ESI) of the protonated molecules, [M + H](+), by using acetonitrile (ACN) instead of methanol (MeOH) as the organic solvent under the conditions in which the samples were supplied to the probe by direct infusion at constant flow rates. The presence of ACN prevents the formation of methanol adducts such as [M + MeOH + H](+) and [M + MeOH + Na](+), while also lowering the signal intensity of sodiated [M + Na](+) ions. Sensitivity of these signaling molecules in terms of signal-to-noise ratio (S/N) using low-resolution LTQ-MS and high-resolution FTICR-MS were compared under reversed-phase (RP) LC separations with ESI interface. Special emphasis was paid to the choice of the separation column, its elution conditions and detection of the major AHL compounds produced by the Serratia liquefaciens strain ATCC 27592. The most promising results were obtained using a RP C16-amide column eluted with a linear mobile phase gradient ACN/H(2)O containing 0.1% formic acid. The whole set of AHL homologs in bacterial extracts was detected in the extracted-ion chromatographic (XIC) mode, and the calculations of molecular formulae were performed by including the isotopic pattern. This mode of displaying data, with a very narrow mass-to-charge ratio window (i.e. +/- 0.0010 as m/z unit) around each selected ion, has allowed the identification of all the eight known homoserine lactones, viz. C(4)-HSL, 3-oxo-C(6)-HSL, C(6)-HSL, 3-oxo-C(8)-HSL, C(8)-HSL, C(10)-HSL, C(12)-HSL and C(14)-HSL. In addition, at least four uncommon signaling mediators previously unreported, namely, 3-oxo-C(10:1)-HSL, 3-oxo-C(11:2)-HSL, 3-oxo-C(13:2)-HSL and 3-OH-C(16)-HSL, were identified and characterized; their roles in cell-to-cell communication has to be elucidated.

A rapid, simple and selective method involving direct separation by gas chromatography (GC) with electron ionization mass spectrometry (EI-MS) was employed to determine some N-acylhomoserine lactones (AHLs). Using GC/EI-MS, simultaneous separation and characterization of AHLs were possible without prior derivatization. Informative fragmentation patterns were obtained to identify the structures of N-acyl chains of AHLs. Electron ionization resulted in a common fragmentation pattern with the most abundant ion at m/z 143 and other minor peaks at m/z 71, 57, and 43. The presence of AHLs in extracts of Burkholderia cepacia strains was achieved in selected ion monitoring mode by using the prominent fragment at m/z 143.

The photochemical behavior of oxyfluorfen [2-chloro-1-(3-etoxy-4-nitrophenoxy)-4-(trifluoromethyl) benzene] on two Greek soils was investigated. Soils were sampled from Nea Malgara and Preveza regions, characterized by a different organic matter content. Soils were spiked with the diphenyl-ether herbicide and irradiation experiments were performed either in the laboratory with a solar simulator (xenon lamp) or outside, under natural sunlight irradiation; other soil samples were kept in the dark to control the retention reaction. Kinetic parameters of both retention and photochemical reactions were calculated using zero-, first- and second- (Langmuir-Hinshelwood) order equations, and best fit was checked through statistical analysis. The soil behaviors were qualitatively similar but quantitatively different, with the soil sampled from the Nea Malgara region much more sorbent as compared with Preveza soil. All studied reactions followed second-order kinetics and photochemical reactions were influenced by retaining capability of the soils. The contributions of the photochemical processes to the global dissipation rates were also calculated. Two main metabolites were identified as 2-chloro-1-(3-ethoxy-4-hydroxyphenoxy)-4-(trifluoromethyl)benzene and 2-chloro-1- (3-hydroxy-4-nitrophenoxy)-4-(trifluoromethyl)benzene.

A simple method is described for the determination of the ionic content of vegetable samples by ion chromatography with suppressed conductivity detection. Extracts of leaves of cucumber (Cucumis sativus), leaves and cotyledons of watermelon (Citrullus lanantus), cotyledons of zucchini (Cucurbita pepo), and leaves and roots of olive (Olea europaea) obtained at room temperature yielded chromatographic profiles with substantial differences in the relative contents of Cl−, NO3−, HPO42− and SO42− as well as of Na+, NH4+, K+, Mg2+ and Ca2+. Although NO3−, Cl− and K+ were common to each extracted sample and accounted for most of the ions present, two additional anion peaks (i.e. malate and oxalate) were detected. Among the vegetable tissues investigated, olive roots contained a considerable amount of oxalate (37 mg/g dry weight), while Na+, which is present in very low amount in extracted samples of leaves and cotyledons, represented ca. 30% of the cationic content of olive roots. In all the examined tissue extracts, K+ was the main cation (16–55 mg/g dry weight) and NO3−, Cl− and HPO42− were the main inorganic anions. Copyright © 2003 John Wiley & Sons, Ltd.

The electrooxidation of 2-naphthol in phosphate buffer at pH 7 leads to the formation of a nonconducting polymer of poly(2-naphthol) on a platinum electrode. Such a resulting thin film displays an interesting permselective behavior, which proved useful in minimizing the interference of ascorbate, acetaminophen, cysteine, and urate sample molecules. Electrochemical detection in flowing streams was used to investigate the relevance of permselectivity for sensor development. Nonconducting poly(2-naphthol) film demonstrated useful also as a novel permselective membrane for glucose oxidase immobilization. The glucose response time, t0.95, evaluated in batch addition experiments, was lower than 4 s. The calibration plot was linear up to 15 mM of glucose with a sensitivity of 2.2 nA/mM.

The electrocatalytic activity of a glassy carbon electrode modified with electrochemically deposited indium(III)-hexacyanoferrate (InHCF) film, which has been subsequently cycled in a solution of ruthenium(III), is described. Although the resulting ruthenium-modified InHCF thin film electrode behaves in a manner analogous to the parent InHCF film, its stability was found to be greatly improved. Such a modified electrode was characterized, in acidic (pH 2) 0.5 M KCl solutions, by cyclic voltammetry and flow injection analysis. It exhibits a high catalytic activity for the electrooxidation of some some inorganic and organic compounds of analytical interest, such as As(III), S2O32-, cysteine, and 2-furaldehyde. The analytical applicability of this novel ruthenium-modified InHCF film electrode for the amperometric detection in flowing streams is discussed. Repetitive sample injections with amperometric detection of thiosulfate at +0.8 V (vs. Ag/AgCl) as applied potential produced a stable response over 8 h of operation, making it very attractive for practical applications.

The electrochemical treatment of an electropolymerized (NiIIL)n thin film (where L represents 5,7,12,14-tetramethyl-dibenzo[b,i]-1,4,8,11-tetraaza [14] annulene) gives rise to a nickel-based catalytic deposit on conducting substrates [glassy carbon (GC), platinum, or gold electrodes], which shows strong similarities to the nickel hydroxide electrode. The effect of potential cycling on surface composition, chemical status, and morphology of this nickel-based chemically modified electrode (Ni-CME) has been investigated by cyclic voltammetry (CV) [1], X-ray photoelectron spectroscopy (XPS), and scanning electron microscopy. XPS analysis was very useful for characterizing the chemical changes in the nickel coordination caused by electrochemical treatments in alkaline solution. The featureless surface of the as-prepared GC/(NiII)Ln, film compared with the fractal nature of the GC/Ni-CME was revealed by electron micrographs. The Ni- CME exhibits high electrocatalytic activity towards the oxidation of carbohydrates in alkaline solution. The electrooxidation capability of the Ni-CME is highlighted by the amperometric detection of sugars after anion-exchange liquid chromatographic separations with alkaline mobile phases.

Sugars and alditols undergo an electrocatalytic oxidation in alkaline solution at cobalt-oxide-based glassy carbon (Co/GC) electrodes. The catalytic deposit was investigated by cyclic voltammetry and scanning electron microscopy (SEM). It is pointed out that, at relatively low coverage of cobalt, two main redox processes, CoII CoIII and CoIII CoIV, are observed in 0.2 M NaOH. The redox behavior of the Co/GC electrode is strongly dependent on hydroxide ion concentration, and no electrocatalytic activity is observed at pH lower than 11.5. Under voltammetric conditions the modified electrode exhibits a non-Nernstian potential/pH shift (−88 ± 3 mV per pH unit). A reasonable explanation of the E/pH behavior in alkaline solution is given and an oxidation mechanism of CoII to CoIII is proposed in terms of hydrous cobalt oxides. No detrimental effects on the electrocatalytic oxidation of polyhydric compounds are observed in the presence of high acetonitrile concentrations (up to 40%). Inspection by SEM of the cobalt-based deposit evidenced the presence of grain-like microcrystallites on the glassy carbon surface.

The voltammetric behavior in alkaline solution of a nickel-based chemically modified electrode (Ni-CME) prepared by oxidative electropolymerization of a nickel–tetraazaannulene complex (NiIIL) was investigated. After electrochemical deposition of the (NiIIL)n film in dichloromethane on a conducting substrate, the modified electrode was continuously cycled in alkaline solution until a steady voltammetric profile was obtained. Although upon electrochemical treatment in basic solutions the nickel–nitrogen tetracoordination of the (NiIIL)n film is probably lost, the nickel species remain entrapped inside the polymeric skeleton and exhibit stable redox behavior resembling that of the nickel hydroxide electrode. This makes it possible to prepare very thin films of nickel-based modified electrodes. In cyclic voltammetry experiments the Ni-CME exhibits a pH dependence of the peak potentials typical of the α-Ni(OH)2/γ-NiOOH redox process, and a separation between anodic and cathodic peak potentials of 64mV at pH 13.2 and 10 mVs−1. The redox catalysis at Ni-CMEs of representative compounds that are not easily electrooxidized, such as choline, cadaverine, γ-amino butyric acid (GABA), benzyl alcohol, and glycine, is demonstrated. The good stability of these nickel-based modified electrodes makes them attractive in analytical applications.

A non-linear least squares iterative refinement has been implemented which shows high performance on a multiple-peak spectrum including baseline or background. Constraints as well as links within a range are introduced to drive the mathematical optimization: each peak parameter (i.e. height, position, Gaussian/Lorentzian mixing ratio and HWHM on both left and right sides) has assigned to it an allowed range of variation and can be strained to be correlated with other parameters belonging either to the same peak (symmetrical peaks) or to other peaks (doublets, triplets, etc.). Peak shapes typical of XP spectra are used and applications in the field of XPS are discussed. Through emulated curves with Poisson distributed noise, the accuracy and precision of back-calculated (refined) parameters have been estimated. Moreover, a confidence level calculated from X2 and degrees of freedom has been suggested to check the overall fitting of experimental curves where the signal-to-noise ratio is a priori unknown. An application to real C ls XP spectra is described as an example and a list of suggestions is given to match operator requirements. Finally, features of NLLSRC are discussed with respect to other approaches.

A novel procedure that yields extremely stable ruthenium-modified inorganic film electrodes is presented. The following prussian blue (PB) analogues were investigated: indium(III)-hexacyanoferrate (InHCF), nickel(II)-hexacyanoferrate (NiHCF), cobalt(II)-hexacyanoferrate (CoHCF), and iron(III)-hexacyanorutenate also known as ruthenium purple (RP). The process includes electrochemical film deposition and subsequent voltammetric conditioning in a Ru(III)-containing solution at pH 2. Electrochemical and spectroscopic properties of each resulting modified film are better defined, redox and switching rates improved, and stability strongly enhanced. XPS supports the conclusions that the incorporation of ruthenium imparts an extensive crosslinking between hydrate microparticles through dinuclear [Fe, Ru] oxo ( O ) and cyanide ( CN ) bridges.

The characterization of carbon fibres by x-ray photoelectron spectroscopy depends on several experimental and theoretical aspects that noticeably improve the accuracy of the analysis when properly taken into account. These aspects are briefly discussed in the light of the most representative literature available. Attention is focused on points that need a deeper understanding and suggestions are given about possible analysis strategies.