p-Tolyl isocyanate derivatization for analysis of CWC-related polar degradation products by mass spectrometry.

Most of the precursors and/or degradation products related to the Chemical Weapons Convention (CWC) are polar. Identification of these molecules in environmental samples provides clues regarding the alleged usage and/or synthesis of the parent toxic chemicals. Such polar compounds need to be derivatized in order to analyze them by gas chromatography-mass spectrometry (GC-MS). In this study, we developed a new derivatizing reagent, para-tolyl isocyanate (PTI), for derivatization of polar CWC-related compounds. The PTI reagent selectively derivatizes the -OH and/or-SH functional groups with high efficiency, but does not react with carboxylic acid (-COOH) or phosphonic acid (-(O)P(OH)2) groups. The PTI derivatives of dialkyl aminoethanols, dialkyl aminoethanol-N-oxides, and 3-quinuclidinol were successfully eluted through GC, and their electron ionization (EI) mass spectra were distinct and provided the structure information by which the isomeric compounds can be easily distinguished. We also calculated the GC-retention index values that can be used for further confirmation of the target compounds. All the studied PTI derivatives can be analyzed by EI-MS with direct insertion probe and/or by direct electrospray ionization mass spectrometry (ESI-MS) together with the MS-MS data; both sets of data provide full structure information. The PTI reagent was found to be better in some respects than the conventional bistrimethylsilyl trifluoroacetamide (BSTFA), a trimethyl silylating reagent. The PTI reagent is commercially available, and the PTI derivatives are highly stable for months and are not sensitive to moisture. The applicability of the PTI derivatization for trace-level determination of the target CWC-related polar compounds in environmental matrices and in human plasma samples is also evaluated.

Mass spectral characterization of the CWC-related isomeric dialkyl alkylphosphonothiolates/alkylphosphonothionates under gas chromatography/mass spectrometry conditions.

RATIONALE: The isomeric dialkyl alkylphosphonothiolates and dialkyl alkylphosphonothionates are listed as scheduled chemicals of the Chemical Weapons Convention (CWC) implemented by the OPCW. The P-S and P-R bond connectivity has to be correctly identified for the verification of the CWC. The present study demonstrates successful identification of the target isomers by selective fragmentation under electron ionization (EI) or chemical ionization (CI) conditions. METHODS: All the studied isomeric compounds (27 in total) were synthesized in our laboratory using established methods, then analyzed by EI and CI gas chromatography/mass spectrometry (GC/MS) using an Agilent 6890 gas chromatograph equipped with a HP-5MS capillary column and interfaced to a 5973 N mass-selective detector. The retention index (RI) values of all the compounds were calculated using Van den Dool's formula. GC/MS/MS and GC/HRMS experiments were also performed using a VG-Autospec (magnetic sector) and JEOL-AccuToF (time-of-flight) mass spectrometer, respectively. RESULTS: The EI mass spectra of all the compounds had an abundant molecular ion at m/z 182, except in the case of a few selected butyl-substituted compounds, where this ion was of low abundance. The EI fragmentation pathways include α-cleavage, McLafferty rearrangement, McLafferty + 1 rearrangement, O/S-alkyl radical loss, and an alkene loss with a hydrogen shift. The characteristic fragment ions and their relative abundances are significant in elucidating the alkyl group attached to the P/S/O-atoms as well as the P-S/P = S bond connectivity. The EI and CI mass spectra together with RI values enable unambiguous identification of all the studied isomeric compounds. CONCLUSIONS: The present study highlights the structural characterization of the isomeric phosphonothiolates and phosphonothionates based on their selective EI fragmentation. The assigned fragmentation pathway helps in the assignment of P-S and P-alkyl connectivity in phosphonothiolates and phosphonothionates, consequently the structure of the unknown compounds. The EI mass spectra (27 compounds) of isomeric compounds are immensely useful in the OPCW official proficiency tests and for off-site analysis.

Mass spectral analysis of N-oxides of Chemical Weapons Convention related aminoethanols under electrospray ionization conditions.

N,N'-Dialkylaminoethanols are the hydrolyzed products or precursors of chemical warfare agents such as V-agents and nitrogen mustards, and they are prone to undergo oxidation in environmental matrices or during decontamination processes. Consequently, screening of the oxidized products of aminoethanols in aqueous samples is an important task in the verification of chemical weapons convention-related chemicals. Here we report the successful characterization of the N-oxides of N,N'-dialkylaminoethanols, alkyl diethanolamines, and triethanolamine using positive ion electrospray ionization mass spectrometry. The collision-induced dissociation (CID) spectra of the [M+H](+) and [M+Na](+) ions show diagnostic product ions that enable the unambiguous identification of the studied N-oxides, including those of isomeric compounds. The proposed fragmentation pathways are supported by high-resolution mass spectrometry data and product/precursor ion spectra. The CID spectra of [M+H](+) ions included [MH-CH(4)O(2)](+) as the key product ion, in addition to a distinctive alkene loss that allowed us to recognize the alkyl group attached to the nitrogen. The [M+Na](+) ions show characteristic product ions due to the loss of groups (R) attached to nitrogen either as a radical (R) or as a molecule [R+H or (R-H)] after hydrogen migration.

Electrospray ionization mass spectral studies of N,N-dialkylaminoethane-2-sulphonic acids.

Oxidative reactions of VX type compounds and N,N-dialkylaminoethane-2-thiols that are precursors for VX compounds produce N,N-dialkylaminoethane-2-sulphonic acids, N(R(1))(R(2))-CH(2)-CH(2)SO(3)H (where R(1) and R(2) = methyl, ethyl, n-propyl and isopropyl, 1-10), as the degradation products, and these degradation products are considered as markers for the detection of chemicals listed in the schedules of Chemical Weapons Convention (CWC) chemicals. Off-site detection of such degradation products in aqueous samples is an important task in the verification of CWC-related chemicals. Here we report a simple method involving the direct analysis of aqueous samples using positive and/or negative ion electrospray ionization (ESI) for the screening, detection and identification of N,N-dialkylaminoethane-2-sulphonic acids, avoiding sample preparation and chromatographic steps. The positive ion ESI mass spectra of all the compounds result in abundant [M+Na](+) ions, and the negative ion spectra show abundant [M-H](-) ions to confirm their molecular weight. The collision-induced dissociation spectra of [M+Na](+) and [M-H](-) give characteristic product ions by which it is easy to detect and identify all the studied N,N-dialkylaminoethane-2-sulphonic acids including those of isomeric compounds. The method is successfully applied to detect the spiked chemical, N,N-diisopropylaminoethane-2-sulphonic acid, present in a water sample received in a proficiency test.

Differential effects of cholesterol and 7-dehydrocholesterol on ligand binding of solubilized hippocampal serotonin1A receptors: implications in SLOS.

The serotonin1A receptor is an important member of the G-protein coupled receptor family, and is involved in the generation and modulation of a variety of cognitive, behavioral, and developmental functions. Solubilization of the hippocampal serotonin1A receptor by 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate (CHAPS) is accompanied by loss of membrane cholesterol which results in a reduction in specific agonist binding activity. Replenishment of cholesterol to solubilized membranes restores the cholesterol content of the membrane and significantly enhances specific agonist binding activity. In order to test the stringency of the requirement of cholesterol in this process, we solubilized native hippocampal membranes followed by replenishment with 7-dehydrocholesterol (7-DHC). 7-DHC is an immediate biosynthetic precursor of cholesterol differing only in a double bond at the 7th position in its sterol ring. Our results show, for the first time, that replenishment of solubilized hippocampal membranes with 7-DHC does not restore ligand binding activity of the serotonin1A receptor, in spite of recovery of the overall membrane order. This observation shows that the requirement for restoration of ligand binding activity is more stringent than the requirement for the recovery of overall membrane order. These novel results have potential implications in understanding the interaction of membrane sterols with this important neuronal receptor under pathogenic conditions such as the Smith-Lemli-Opitz syndrome.

Gas chromatographic-mass spectrometric determination of alkylphosphonic acids from aqueous samples by ion-pair solid-phase extraction on activated charcoal and methylation.

In the present paper, we report an improved ion-pair solid-phase extraction (IP-SPE) method for the analysis of alkylphosphonic acids, namely, methyl, ethyl and propylphosphonic acids, present in the aqueous sample. The aqueous sample was mixed with an ion-pair reagent, phenyltrimethylammonium hydroxide (PTMAH) and passed through activated charcoal SPE cartridge. The retained chemicals in the cartridge were extracted with methanol and analysed by gas chromatography-mass spectrometry (GC-MS) under the electron impact ionization (EI) mode. The analytes were converted to their methyl esters by pyrolytic methylation in the hot GC injection port. The recoveries of alkylphosphonic acids were above 95% and the minimum detection limits were as low as 10 ng/mL. The recovery of the test chemicals was tested with solvents, dichloromethane, n-hexane, ethyl acetate, acetone, acetonitrile and methanol. The chemicals could be efficiently extracted by the hydrophilic solvents. The method did not work at the highly acidic pH (when acidified with dilute HCl) but worked well from pH 4.0 to 14.0. The present method was also tested with other tetra-(methyl, ethyl, propyl and n-butyl)ammonium hydroxides. The test chemicals were not converted to their methyl and ethyl esters with tetramethyl and tetraethylammonium hydroxides, whereas they were converted to their corresponding propyl and n-butyl esters with tetrapropyl and tetra(n-butyl)ammonium hydroxides. The method was also applied to two highly cross-linked polymeric sorbents DSC-6S and Oasis HLB. The recovery of the chemicals on these sorbents was observed to be poor. Methylation using phenyltrimethylammonium hydroxide is non-hazardous and advantageous over methylation using diazomethane. The method was applied to the analysis of aqueous samples given in one of the official proficiency tests conducted by the Organization for the Prohibition of Chemical Weapons and all the spiked chemicals were identified as methyl esters.

In situ nucleophilic substitution reaction of N,N-dialkylaminoethyl-2-chlorides monitored by gas chromatography/mass spectrometry.

The detection and identification of degradation products of scheduled chemicals, which are characteristic markers of Chemical Warfare agents (CWAs), plays a key role in verification analysis. Identification of such non-scheduled but specific markers of CWAs helps in deciphering the kind of agent that was present in the sample submitted for off-site analysis. This paper describes the stability of N,N-dialkylaminoethyl-2-chlorides, which are precursors for highly toxic chemicals like VX, in different solvents. These compounds are stable in chloroform, acetonitrile, hexane and dichloromethane but tend to undergo in situ nucleophilic substitution reaction in the presence of alcohols giving the corresponding alkyl ether. The study shows that N,N-dialkylaminoethyl alkyl ethers can be used as markers of N,N-dialkylaminoethyl-2-chlorides. A detailed degradation study of these compounds in the presence of alcohols was carried out and it was found that the reaction follows pseudo-first order kinetics. Electron ionization mass spectral data for the methyl ethers of all the compounds are briefly discussed.

Electrospray ionisation mass spectral studies on hydrolysed products of sulfur mustards.

Off-site detection of the hydrolysed products of sulfur mustards in aqueous samples is an important task in the verification of Chemical Weapons Convention (CWC)-related chemicals. The hydrolysed products of sulfur mustards are studied under positive and negative electrospray ionisation (ESI) conditions using an additive with a view to detecting them at trace levels. In the presence of cations (Li(+), Na(+), K(+) and NH(4) (+)), the positive ion ESI mass spectra of all the compounds include the corresponding cationised species; however, only the [M+NH(4)](+) ions form [M+H](+) ions upon decomposition. The tandem mass (MS/MS) spectra of [M+H](+) ions from all the hydrolysed products of the sulfur mustard homologues were distinct and allowed these compounds to be characterised unambiguously. Similarly, the negative ion ESI mass spectra of all the compounds show prominent adducts with added anions (F(-), Cl(-), Br(-), and I(-)), but the [M-H](-) ion can only be generated by decomposition of an [M+F](-) ion. The MS/MS spectra of the [M-H](-) ions from all the compounds result in a common product ion at m/z 77. A precursor ion scan of m/z 77 is shown to be useful in the rapid screening of these compounds in aqueous samples at trace levels, even in the presence of complex masking agents, without the use of time-consuming sample preparation and chromatography steps. An MS/MS method developed to measure the detection limits of the hydrolysed products of sulfur mustards found these to be in the range of 10-500 ppb.

Mass spectral analysis of chloropicrin under negative ion chemical ionization conditions.

A chemical ionization (CI) method is developed for the first time to obtain molecular weight information for chloropicrin (CP), which is used as a chemical warfare agent and as an insecticide. The study includes a detailed investigation on the behavior of CP under electron impact (EI) and CI. Reagent gases of different nature, i.e., methane, isobutane, ammonia, hydrogen, and carbon dioxide, were used for CI analysis. Negative ion mode is found more sensitive than positive ion mode for the EI/CI mass spectrometric analysis of CP, but none of the methods provided molecular weight information, except negative ion CI using ammonia as the reagent gas (NICI (NH3)). The NICI (NH3) showed formation of the quasi-molecular ion, [M + H]-, in addition to other adduct ions. The [M + H]- abundance critically depends on the source temperature, reagent gas pressure, and concentration of the analyte, and it can be 13% under optimized conditions by which CP can be confirmed unambiguously. This method meets the criteria used in official proficiency tests conducted by OPCW for confirming the molecular weight of the unknowns.