Rapid monitoring of sulfur mustard degradation in solution by headspace solid-phase microextraction sampling and gas chromatography mass spectrometry.

A method using headspace solid-phase microextraction (HS-SPME) followed by gas chromatography/mass spectrometry (GC/MS) analysis has been developed to gain insight into the degradation of the chemical warfare agent sulfur mustard in solution. Specifically, the described approach simplifies the sample preparation for GC/MS analysis to provide a rapid determination of changes in sulfur mustard abundance. These results were found to be consistent with those obtained using liquid-liquid extraction (LLE) GC/MS. The utility of the described approach was further demonstrated by the investigation of the degradation process in a complex matrix with surfactant added to assist solvation of sulfur mustard. A more rapid reduction in sulfur mustard abundance was observed using the HS-SPME approach with surfactant present and was similar to results from LLE experiments. Significantly, this study demonstrates that HS-SPME can simplify the sample preparation for GC/MS analysis to monitor changes in sulfur mustard abundance in solution more rapidly, and with less solvent and reagent usage than LLE.

Ion-molecule reactions of O,S-dimethyl methylphosphonothioate: evidence for intramolecular sulfur oxidation during VX perhydrolysis.

The alkaline perhydrolysis of the nerve agent O-ethyl S-[2-(diisopropylamino)ethyl] methylphosphonothioate (VX) was investigated by studying the ion-molecule reactions of HOO(-) with O,S-dimethyl methylphosphonothioate in a modified linear ion-trap mass spectrometer. In addition to simple proton transfer, two other abundant product ions are observed at m/z 125 and 109 corresponding to the S-methyl methylphosphonothioate and methyl methylphosphonate anions, respectively. The structure of these product ions is demonstrated by a combination of collision-induced dissociation and isotope-labeling experiments that also provide evidence for their formation by nucleophilic reaction pathways, namely, (i) S(N)2 at carbon to yield the S-methyl methylphosphonothioate anion and (ii) nucleophilic addition at phosphorus affording a reactive pentavalent intermediate that readily undergoes internal sulfur oxidation and concomitant elimination of CH(3)SOH to yield the methyl methylphosphonate anion. Consistent with previous solution phase observations of VX perhydrolysis, the toxic P-O cleavage product is not observed in this VX model system and theoretical calculations identify P-O cleavage to be energetically uncompetitive. Conversely, intramolecular sulfur oxidation is calculated to be extremely exothermic and kinetically accessible explaining its competitiveness with the facile gas phase proton transfer process. Elimination of a sulfur moiety deactivates the nerve agent VX and thus the intramolecular sulfur oxidation process reported here is also able to explain the selective perhydrolysis of the nerve agent to relatively nontoxic products.

Detection of the abundance of diacylglycerol and triacylglycerol molecular species in cells using neutral loss mass spectrometry.

Triacylglycerols (TAGs) are neutral lipids present in all mammalian cells as energy reserves, and diacylglycerols (DAGs) are present as intermediates in phospholipid biosynthesis and as signaling molecules. The molecular species of TAGs and DAGs present in mammalian cells are quite complex, and previous investigations revealed multiple isobaric species having molecular weights at virtually every even mass between 600 and 900 Da, making it difficult to assess changes of individual molecular species after cell activation. A method has been developed, using tandem MS and neutral loss scanning, to quantitatively analyze changes in those glyceryl ester molecular species containing identical fatty acyl groups. This was carried out by neutral loss scanning of 18 common fatty acyl groups where the neutral loss corresponded to the free carboxylic acid plus NH(3). Deuterium-labeled internal standards were used to normalize the signal for each nominal [M+NH(4)](+) ion undergoing this neutral loss reaction. This method was applied in studies of TAGs in RAW 264.7 cells treated with the toll-like receptor 4 ligand Kdo(2)-lipid A. A 50:1-TAG containing 18:1 was found to increase significantly over a 24-h time course after Kdo(2)-lipid A exposure, whereas an isobaric 50:1-TAG containing 16:1 did not change relative to controls.

Is the hypothiocyanite anion (OSCN)- the major product in the peroxidase catalyzed oxidation of the thiocyanate anion (SCN)-? A joint experimental and theoretical study.

The hypothiocyanate anion (OSCN)(-) is reported to be a major product of the lactoperoxidase/H(2)O(2)/(SCN)(-) system, and this anion is proposed to have significant antimicrobial properties. The collision induced (CID) negative ion mass spectrum of "(OSCN)(-)" has been reported: there is a pronounced parent anion at m/z 74, together with fragment anions at m/z 58 (SCN)(-) and 26 (CN)(-). These fragment anions are consistent with structure (OSCN)(-). However there is also a lesser peak at m/z 42 (OCN(-) or CNO(-)) in this spectrum which is either formed by rearrangement of (OSCN)(-) or from an isomer of this anion. The current theoretical investigation of (OSCN)(-) and related isomers, together with the study of possible rearrangements of these anions, indicates that ground-state singlet (OSCN)(-) is a stable species and that isomerization is unlikely. The three anions (OSCN)(-), (SCNO)(-), and (SNCO)(-) have been synthesized (in the ion source of a mass spectrometer) by unequivocal routes, and their structures have been confirmed by a consideration of their collision induced (negative ion) and charge reversal (positive ion) mass spectra. The CID mass spectrum of (SCNO)(-) shows formation of m/z 42 (CNO(-)), but the corresponding spectra of (OSCN)(-) or (SNCO)(-) lack peaks at m/z 42. Combined theoretical and experimental data support earlier evidence that the hypothiocyanite anion is a major oxidation product of the H(2)O(2)/(SCN)(-) system. However, the formation of m/z 42 in the reported CID spectrum of "(OSCN)(-)" does not originate from (OSCN)(-) but from another isomer, possibly (SCNO)(-).

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.

Neutral cumulene oxide CCCCO is accessible by one-electron oxidation of [CCCCO]-* in the gas phase.

Calculations at the CCSD(T)/aug-cc-pVDZ//B3LYP/6-31G* level of theory indicate that doublet [CCCCO]-* is a stable species which should undergo collision-induced Franck-Condon vertical oxidation under neutralisation-reionisation conditions (-NR+) to produce both triplet CCCCO (ground state) and singlet CCCCO. Some of the neutral CCCCO species formed (particularly the triplet) should be stable for the microsecond duration of the NR experiment, whereas others will be energised (particularly the singlet) and should decompose to C3 and CO. The [CCCCO]-* radical anion has been formed in the ion source of the mass spectrometer by the reaction CH3OCH2C[triple bond]C-CO-CH(CH3)2 + O-* --> [CCCCO]-* + CH3O* + H2O + (CH3)2CH*. The -NR+ spectrum of [CCCCO]-* shows a recovery signal at m/z 64 corresponding to ionised CCCCO, together with a pronounced peak at m/z 36 (CCC+*) produced by ionisation of CCC (formed by the reaction CCCCO --> CCC + CO). The experimental observations are in agreement with the predictions of the theoretical study.

Rearrangements of transient neutral molecules in the gas phase. Does the conversion of CCCHO to HCCCO involve oxygen or hydrogen migration?

Stable (CC13CHO)- may be formed in the chemical ionisation ion source of a VG ZAB 2HF mass spectrometer by the SN2(Si) reaction between Me3SiC[triple bond]C13CHO and F-. Vertical (Franck-Condon) one-electron oxidation of (CC13CHO)- in the first of the tandem collision cells of a VG ZAB 2HF mass spectrometer gives CC13CHO. Some of these neutrals have sufficient excess energy to effect rearrangement to HCC13CO, some of which are energised and to decompose to HCC. and 13CO. Thus the neutral rearrangement exclusively involves H migration: no products from O migration are detected. The corresponding two-electron oxidation of (CC13CHO)- gives mainly unrearranged (CC13CHO)+. A minority of these cations are energised and rearrange by H and O migration to yield (HCC13CHO)+ and (OCC13CHO)+ respectively. All experimental observations are backed up by molecular modelling at the CCSD(T)/aug-cc-pVDZ//B3LYP/6-31G(d) level of theory.

Generation of transient neutrals in the gas phase from anionic precursors. Does energised CNCCO rearrange to NCCCO?

The stability and reactivity of the neutral species CNCCO generated by one electron oxidation of the anion [CNCCO](-) have been investigated by a combination of theoretical calculations (carried out at the CCSD(T)/aug-cc-pVDZ//B3LYP/6-31G(d) level of theory) and tandem mass spectrometric experiments. Some of the neutrals formed in this way are stable for the microsecond duration of the experiment, but others are energised. The neutrals which are energised may either (i) dissociate [CNCCO --> CNC + CO (+92 kJ mol(-1))], and/or (ii), undergo the isonitrile to nitrile rearrangement to yield NCCCO energised neutrals (barrier 133 kJ mol(-1), reaction exothermic by 105 kJ mol(-1)). Some of these rearranged neutrals NCCCO have excess energies as high as 238 kJ mol(-1) and will dissociate [NCCCO --> NCC + CO (+203 kJ mol(-1))].

The formation of RCCCO and CCC(O)R (R = Me, Ph) neutral radicals from ionic precursors in the gas phase: the rearrangement of CCC(O)Ph.

Neutrals MeCCCO, CCC(O)Me, PhCCCO and CCC(O)Ph have been made by neutralisation of [MeCCCO](+), [CCC(O)Me](-), [PhCCCO](+) and [CC(CO)Ph](-). Neutrals MeCCCO, CCC(O)Me and PhCCCO are stable for the microsecond duration of the neutralisation experiment. A joint experimental and theoretical study (energies calculated at the B3LYP/aug-cc-pVDZ//B3LYP/6-31G(d) level of theory) suggests that the neutral radical CCC(O)Ph rearranges via a four-centred ipso radical cyclisation/ring opening to form the isomer PhCCCO in an exothermic reaction. (13)C labelling confirms that the rearrangement does not involve O migration. Some of the PhCCCO radicals formed in this reaction are sufficiently energised to effect decomposition to give PhCC and CO.

Do the interstellar molecules CCCO and CCCS rearrange when energised?

Neutrals CCCO, CC(13)CO, CCCS and CC(13)CS have been prepared by one-electron vertical (Franck-Condon) oxidation of the precursor anion radicals (CCCO)(-*), (CC(13)CO)(-*), (CCCS)(-*) and (CC(13)CS)(-*)respectively in collision cells of a reverse sector mass spectrometer. Ionisation of the neutrals to decomposing cations shows the neutrals to be stable for the microsecond duration of the neutralisation-ionisation ((-)NR(+)) experiment. No rearrangement of the label in energised CC(13)CO or CC(13)CS occurs during these experiments. In contrast, minor rearrangement of (CC(13)CO)(+*) is observed [(CC(13)CO)(+*)-->(OCC(13)C)(+*), while significant rearrangement occurs for (CC(13)CS)(+*) [(CC(13)CS)(+*)-->(SCC(13)C)(+*)]. Theoretical calculations at the CCSD(T)/aug-cc-pVDZ//B3LYP/6-31G(d) level of theory show that the cationic rearrangements occur by stepwise processes via key rhombic structures. Overall, the degenerate processes result in O and S migration from C-3 to C-1. The cations (CCCO)(+*) and (CCCS)(+*) require excess energies of > or = 516 and > or = 226 kJ mol(-1) respectively to effect rearrangement.

The fragmentations of [M-H]- anions derived from underivatised peptides. The side-chain loss of H2S from Cys. A joint experimental and theoretical study.

Loss of H2S is the characteristic Cys side-chain fragmentation of the [M-H]- anions of Cys-containing peptides. A combination of experiment and theory suggests that this reaction is initiated from the Cys enolate anion as follows: RNH-(-)C(CH2SH)CONHR' Ø [RNHC(=CH2)CONHR' (HS-)] Ø [RNHC(=CH2)CO-HNR'-H]-+H2S. This process is facile. Calculations at the HF/6-31G(d)//AM1 level of theory indicate that the initial anion needs only > or =20.1 kcal mol(-1) of excess energy to effect loss of H2S. Loss of CH2S is a minor process, RNHCH(CH2SH)CON(-)-R' Ø RNHCH(CH2S-)CONHR' Ø RNH -CHCONHR+CH2S, requiring an excess energy of > or =50.2 kcal mol(-1). When Cys occupies the C-terminal end of a peptide, the major fragmentation from the [M-H]- species involves loss of (H2S+CO2). A deuterium-labelling study suggests that this could either be a charge-remote reaction (a process which occurs remote from and uninfluenced by the charged centre in the molecule), or an anionic reaction initiated from the C-terminal CO2- group. These processes have barriers requiring the starting material to have an excess energy of > or =79.6 (charge-remote) or > or =67.1 (anion-directed) kcal mol(-1), respectively, at the HF/6-31G(d)//AM1 level of theory. The corresponding losses of CH2O and H2O from the [M-H]- anions of Ser-containing peptides require > or =35.6 and > or =44.4 kcal mol(-1) of excess energy (calculated at the AM1 level of theory), explaining why loss of CH2O is the characteristic side-chain loss of Ser in the negative ion mode.