Alkyl Alkylphosphonic Acids Research Articles

Single vial sample preparation of markers of nerve agents by dispersive solid-phase extraction using magnetic strong anion exchange resins

A sample preparation method involving extraction, enrichment and derivatization of acidic degradation products of nerve agents was developed using magnetic strong anion exchange resins (MSAX). The method was performed in a single vial involving magnetic dispersive solid phase extraction (MDSPE). Analytes were derivatized with N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) in the presence of resins. MSAX were custom synthesized using Fe3O4 nanoparticles as core, 4-vinylpyridine-co-divinylbenzene as polymer shell and quaternary pyridinium function as anion-exchanger. Hydroxide ions were the counter-anions of MSAX to effectively capture the acidic alkyl alkylphosphonic acids (AAPAs) and alkylphosphonic acids (APAs). Quantitative measurements of analytes were performed in the selected ion monitoring mode of GC–MS. Full scan mode of analysis was followed for identifications. Under the optimized conditions analytes were recovered in the range of 39.7–98.8% (n=3, relative standard deviations (RSD) from 0.3 to 6.5%). Limits of detection (LODs) were in the range of 0.1–1.1ngmL−1; and the linear dynamic range was 5–1000ngmL−1 with r2 of 0.9977–0.9769. Applicability of the method was tested with rain-, tap-, muddy-water and Organization for Prohibition of Chemical Weapons (OPCW) Proficiency Test samples.

Solid supported in situ derivatization extraction of acidic degradation products of nerve agents from aqueous samples

This study deals with the solid supported in situ derivatization extraction of acidic degradation products of nerve agents present in aqueous samples. Target analytes were alkyl alkylphosphonic acids and alkylphosphonic acids, which are important environmental signatures of nerve agents. The method involved tert-butyldimethylchlorosilane mediated in situ silylation of analytes on commercially available diatomaceous solid phase extraction cartridges. Various parameters such as derivatizing reagent, its concentration, reaction time, temperature and eluting solvent were optimized. Recoveries of the analytes were determined by GC–MS which ranged from 60% to 86%. The limits of detection (LOD) and limit of quantification (LOQ) with selected analytes were achieved down to 78 and 213ngmL−1 respectively, in selected ion monitoring mode. The successful applicability of method was also demonstrated on samples of biological origin such as plasma and to the samples received in 34th official proficiency test conducted by the Organization for Prohibition the of Chemical Weapons.

Determination of alkyl alkylphosphonic acids by liquid chromatography coupled with electrospray ionization tandem mass spectrometry using a dicationic reagent

A new analytical method based on liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) is proposed and validated for the identification and quantification of alkyl alkylphosphonic acids (AAPAs) in aqueous matrices. Retrospective detection and identification of degradation products of chemical warfare agents is important as an indicator of possible use of chemical warfare agents or of environmental contamination. A commercially available solution of 1,9-nonanediyl-bis-(3-methylimidazolium)bisfluoride (NBMI) allowed detection of AAPAs by positive mode electrospray ionization mass spectrometry by forming an adduct with AAPAs. MS/MS experiments using an ion trap analyzer were carried out for unambiguous identification of AAPAs. Different parameters were optimized in order to obtain both an adequate chromatographic separation and a high sensitivity using experimental design methodology. Quantification was done with matrix-matched calibration standards of AAPAs. The method was validated in terms of linearity (r(2) >0.982), intra- and inter-day precisions (RSD below 15%), and robustness. The method is sensitive enough for the determination of AAPAs in aqueous matrices, with limits of detection in the 1-5 ng mL(-1) range and limits of quantification in the 5-20 ng mL(-1) range. Finally, the method was successfully applied to determine these AAPAs in aqueous samples provided by the Organization for the Prohibition of Chemical Weapons during 26(th) and 29(th) official proficiency tests. The added advantage of this method is identification of low mass range analyte at high mass range, which obviates the background noise at low mass range.

Zirconia coated stir bar sorptive extraction combined with large volume sample stacking capillary electrophoresis-indirect ultraviolet detection for the determination of chemical warfare agent degradation products in water samples

In this study, a sensitive, selective and reliable analytical method by combining zirconia (ZrO2) coated stir bar sorptive extraction (SBSE) with large volume sample stacking capillary electrophoresis-indirect ultraviolet (LVSS-CE/indirect UV) was developed for the direct analysis of chemical warfare agent degradation products of alkyl alkylphosphonic acids (AAPAs) (including ethyl methylphosphonic acid (EMPA) and pinacolyl methylphosphonate (PMPA)) and methylphosphonic acid (MPA) in environmental waters. ZrO2 coated stir bar was prepared by adhering nanometer-sized ZrO2 particles onto the surface of stir bar with commercial PDMS sol as adhesion agent. Due to the high affinity of ZrO2 to the electronegative phosphonate group, ZrO2 coated stir bars could selectively extract the strongly polar AAPAs and MPA. After systematically optimizing the extraction conditions of ZrO2-SBSE, the analytical performance of ZrO2-SBSE-CE/indirect UV and ZrO2-SBSE-LVSS-CE/indirect UV was assessed. The limits of detection (LODs, at a signal-to-noise ratio of 3) obtained by ZrO2-SBSE-CE/indirect UV were 13.4–15.9μg/L for PMPA, EMPA and MPA. The relative standard deviations (RSDs, n=7, c=200μg/L) of the corrected peak area for the target analytes were in the range of 6.4–8.8%. Enhancement factors (EFs) in terms of LODs were found to be from 112- to 145-fold. By combining ZrO2 coating SBSE with LVSS as a dual preconcentration strategy, the EFs were magnified up to 1583-fold, and the LODs of ZrO2-SBSE-LVSS-CE/indirect UV were 1.4, 1.2 and 3.1μg/L for PMPA, EMPA, and MPA, respectively. The RSDs (n=7, c=20μg/L) were found to be in the range of 9.0–11.8%. The developed ZrO2-SBSE-LVSS-CE/indirect UV method has been successfully applied to the analysis of PMPA, EMPA, and MPA in different environmental water samples, and the recoveries for the spiked water samples were found to be in the range of 93.8–105.3%.

Polymer supported N-benzyl-N-nitroso-4-toluenesulfonamide: An improved reagent for the derivatization of the acidic markers of nerve agents

We report herein the synthesis, characterization, stability study of a new solid supported derivatizing agent, N-benzyl-N-nitroso-4-poly(styrene-co-divinylbenzene)sulfonamide beads, for the preparation of phenyldiazomethane (PDM). The active component loading was found to be 2.0 ± 0.1 mmol g−1. This derivatizing agent is easy to synthesize and purify. The impurities and byproducts generated during the formation of PDM remain bound to the polymer backbone. These byproducts can be easily removed by filtration. Thus, pure PDM is obtained without the need for further purification. Due to the formation of a dilute solution of PDM, the chances of explosion and intoxication are minimized. The PDM from this derivatizing agent was used for the benzylation of a variety of acids. We have evaluated this derivatizing agent for the benzylation of alkylphosphonic acids (APAs) and alkyl alkylphosphonic acids (AAPAs). Subsequently, this reagent was applied for analysis of a real sample containing pinacolyl methylphosphonic acid by onflow LC-UV-1H NMR analysis.

Thermally assisted methylation and subsequent silylation of scheduled acids of chemical weapon convention for on-site analysis and its comparison with the other methods of methylation

On-site verification of the chemical weapon convention (CWC) requires provision for the detection and identification of alkyl phosphonic acids as well as some organic acids that are amenable to GC–MS only after derivatisation. Various derivatisation methods have been used for the identification of these acids and for many cases the methyl derivatives are less prone to artifacts possibly leading to false positive identification. Methylation with diazomethane is widely used but, especially for on-site analysis it has limitation due to the potential explosive and health hazards. Other methylation procedures like trimethylsilyldiazomethane (TMSD), thermally assisted methylation (TAM) by trimethylphenylammonium hydroxide (TMPAH) and trimethylsulfonium hydroxide (TMSH) are evaluated. Data for methylation for the alkyl alkylphosphonic acids, alkylphosphonic acids and benzilic acid are reported. In addition, TAM followed by the silylation in the same sample without any additional sample preparation is also reported. Several parameters such as solvent, temperature, amount of reagents, time, etc. were studied. The two commercially available reagents namely, TMPAH and TMSH for TAM and subsequent silylation were evaluated. The LOD with TMPAH was below 0.5ng per injection since all of the acids were detected by GC–MS with the S/N of >3 in full scan mode by AMDIS and their inter day relative standard deviation was from 4.7% to 10.8%.

On-line high-performance liquid chromatography–ultraviolet–nuclear magnetic resonance method of the markers of nerve agents for verification of the Chemical Weapons Convention

This paper details an on-flow liquid chromatography–ultraviolet–nuclear magnetic resonance (LC–UV–NMR) method for the retrospective detection and identification of alkyl alkylphosphonic acids (AAPAs) and alkylphosphonic acids (APAs), the markers of the toxic nerve agents for verification of the Chemical Weapons Convention (CWC). Initially, the LC–UV–NMR parameters were optimized for benzyl derivatives of the APAs and AAPAs. The optimized parameters include stationary phase C 18, mobile phase methanol:water 78:22 (v/v), UV detection at 268 nm and 1H NMR acquisition conditions. The protocol described herein allowed the detection of analytes through acquisition of high quality NMR spectra from the aqueous solution of the APAs and AAPAs with high concentrations of interfering background chemicals which have been removed by preceding sample preparation. The reported standard deviation for the quantification is related to the UV detector which showed relative standard deviations (RSDs) for quantification within ±1.1%, while lower limit of detection upto 16 μg (in μg absolute) for the NMR detector. Finally the developed LC–UV–NMR method was applied to identify the APAs and AAPAs in real water samples, consequent to solid phase extraction and derivatization. The method is fast (total experiment time ∼2 h), sensitive, rugged and efficient.

Liquid–liquid–liquid microextraction of degradation products of nerve agents followed by liquid chromatography–tandem mass spectrometry

Alkyl alkylphosphonic acids (AAPAs) are important environmental markers of nerve agents. A simple hollow fiber-based liquid–liquid–liquid microextraction (HFLLLME) technique has been developed to enrich the AAPAs from water. AAPAs were extracted from acidified aqueous phase to organic phase present in pores of the hollow fiber, and then back extracted into the alkaline acceptor phase present in the lumen of the hollow fiber. Variables affecting the HFLLLME process were optimized using a Plackett–Burman design and a Doehlert design. Optimal experimental conditions were: organic solvent, 1-octanol; pH of acceptor phase, 14; extraction time, 60 min; pH of donor phase, 1; and NaCl concentration, 10% (w/v). Depending upon the alkyl substituent, lower limits of detection varied from 0.1 to 100 ng mL −1 (S/N ≥ 5). Repeatability of the method was observed with relative standard deviation of 1.49–9.83% ( n = 3). After validation, the method was applied to detect AAPAs present in the water sample provided by the Organization for Prohibition of Chemical Weapons (OPCW) during the 23rd official proficiency test. The added advantage of this method is that several successive extractions of AAPAs from the same water sample can be performed.

Field-amplified sample stacking for the detection of chemical warfare agent degradation products in low-conductivity matrices by capillary electrophoresis-mass spectrometry

Preconcentration of chemical warfare agent degradation products (alkylphosphonic acids and alkyl alkylphosphonic acids) in low-conductivity matrices (purified water, tap water and local river water) by field-amplified sample stacking (FASS) was developed for capillary electrophoresis (CE) coupled to ion trap mass spectrometry. FASS was performed by adding a mixture of HCOONH 4 and NH 4OH in appropriate concentrations to the sample. This allowed to control the conductivity and the pH of the sample in order to obtain FASS performances that are independent of analyte concentration. The influence of different parameters on FASS (sample to background electrolyte (BGE) conductivity ratio, injection volume and concentration of BGE) was studied to determine the optimal conditions and was rationalized by using the theoretical model developed by Burgi and Chien. A good correlation was obtained between the bulk electroosmotic velocity predicted by this model and the experimental value deduced from the migration time of the electroosmotic flow marker detected by mass spectrometry (MS). This newly developed method was successfully applied to the analysis of tap water and local river water fortified with the analytes and provided a 10-fold sensitivity enhancement in comparison to the signal obtained without preconcentration procedure. The quite satisfactory repeatability and linearity for peak areas obtained in the 0.5–5 μg mL −1 concentration range allow quantitative analysis to be implemented. Limits of detection of 0.25–0.5 μg mL −1 for the alkyl alkylphosphonic acids and of 0.35–5 μg mL −1 for the alkylphosphonic acids were reached in tap water and river water.

Determination of hydrolytic degradation products of nerve agents by injection port fluorination in gas chromatography/mass spectrometry for the verification of the Chemical Weapons Convention

Retrospective detection and identification of markers of chemical warfare agents are important aspects of verification of the Chemical Weapons Convention. Alkyl alkylphosphonic acids (AAPAs) and alkylphosphonic acids (APAs) are important markers of nerve agents. We describe the development and optimization of a new gas chromatography/mass spectrometry (GC/MS) injection port fluorination method for the derivatization of AAPAs and APAs. The process involved the injection of acids with trifluoroacetic anhydride in GC/MS, where acids are converted into their corresponding volatile fluorides. Various reaction conditions such as fluorinating agent, injection port temperature and splitless time were optimized. The maximum reaction efficiency of the acids with trifluoroacetic anhydride was observed at 230 degrees C injection port temperature with a splitless time of 2 min. APAs showed best analytical efficiencies at 400 degrees C injection port temperature, while the other conditions were similar to those of AAPAs. The linearities of response for APAs and AAPAs were in the range of 1-25 and 5-100 microg mL(-1), respectively, with limits of detection ranging from 500 pg to 800 ng mL(-1).

Extraction of acidic degradation products of organophosphorus chemical warfare agents: Comparison between silica and mixed-mode strong anion-exchange cartridges

The analysis of alkyl alkylphosphonic acids (AAPAs) and alkylphosphonic acids (APAs), the hydrolyzed products of nerve agents, constitutes an important aspect for verifying the compliance to the Chemical weapons convention (CWC). This work devotes on the development of solid-phase extraction method using polymeric mixed-mode strong anion-exchange (Oasis MAX) cartridges for extraction of AAPAs and APAs from water. The extracted analytes were analyzed by GC–MS under full scan and selected ion monitoring mode. The extraction efficiencies of MAX and silica-based anion-exchange cartridges were compared, and results revealed that MAX sorbents yielded better recoveries. Extraction parameters, such as loading capacity, extraction solvent, its volume, and washing solvent were optimized. Best recoveries were obtained using 1 mL of acidic methanol (0.1 M), and limits of detection could be achieved up to 5 × 10 −4 μg mL −1 (in SIM) and 0.05 μg mL −1 in full scan mode. The method was successfully employed for the detection and identification of alkylphosphonic acids present in soil sample sent by the Organization for Prohibition of Chemical Weapons (OPCW) in the official proficiency tests.

Separation and identification of isomeric acidic degradation products of organophosphorus chemical warfare agents by capillary electrophoresis-ion trap mass spectrometry

Capillary electrophoresis (CE) coupled to ion trap mass spectrometry (MS) was evaluated for the separation and identification of chemical warfare agent degradation products (alkylphosphonic acids and alkyl alkylphosphonic acids). Different analytical parameters were optimized in negative ionization mode such as electrolyte composition (15 mM CH 3COONH 4, pH 8.8), sheath liquid composition (MeOH/H 2O/NH 3, 75:25:2, v/v/v), nebulization and ion trapping conditions. A standard mixture of five alkylphosphonic (di)acids and five alkyl alkylphosphonic (mono)acids containing isomeric compounds was used in order to evaluate CE selectivity and MS identification capability. The obtained electropherograms revealed that CE selectivity was very limited in the case of alkyl alkylphosphonic acid positional isomers, whereas isomeric isopropylphosphonic and propylphosphonic acids were baseline-separated. CE–MS–MS experiments provided an unambiguous identification of each isomeric co-migrating alkyl alkylphosphonic acids thanks to the presence of specific fragment ions. On the other hand, CE separation was mandatory for the identification of isomeric alkylphosphonic acids, which led to the same fragment ion and could not be differentiated by MS–MS. The developed method was applied to the analysis of soil extracts spiked with the analytes (before or after extraction treatment) and appeared to be very promising since resolution and sensitivity were similar to those observed in deionized water. Especially, analytes were detected and identified in soil extract spiked at 5 μg mL −1 with each compound before extraction treatment.

Selective extraction of organophosphorus nerve agent degradation products by molecularly imprinted solid-phase extraction

The analysis of alkyl alkylphosphonic acids, the degradation products of V and G nerve agents as VX, Sarin or Soman, is an important task for the verification of compliance to the Chemical Weapons Convention. The detection of these contaminants at low concentration levels is often difficult in complex matrices due to the amount of interfering substances. Molecularly imprinted solid-phase extraction technique should allow a selective extraction of these compounds from complex samples, and thus make their detection easier. Two molecularly imprinted polymers (MIPs) prepared with methacrylic acid (MAA) as monomer and pinacolyl methylphosphonic acid (PMPA) as template molecule were synthesised and tested. The first polymer, MIP A, was prepared with ethylene glycol dimethacrylate (EGDMA) in dichloromethane. The second polymer, MIP B, was synthesised using trimethylolpropane trimethacrylate (TRIM) in acetonitrile. To evaluate the selectivity provided by these MIPs, the retention of the ethyl methylphosphonic acid (EMPA) target molecule was studied in parallel on a non-imprinted polymer (NIP). While MIP A does not show any difference compared to NIP A, a good selectivity was obtained for MIP B. After the optimisation of the extraction process, 60% of EMPA can be removed from the NIP B without affecting the retention on the MIP B. A recovery of extraction of 93% was then obtained on the MIP B. Its capacity was then measured and corresponds to 97 μg of EMPA per gram of MIP. Finally, the selectivity of MIP B was clearly demonstrated by applying it to the clean-up of a soil extract spiked with EMPA.

The trace analysis of alkyl alkylphosphonic acids in urine using gas chromatography–ion trap negative ion tandem mass spectrometry

A sensitive method has been developed for the trace analysis of alkyl alkylphosphonic acids, metabolites of nerve agents, in urine using a benchtop ion trap mass spectrometer. The acids were isolated from urine by simple solid phase extraction and converted to their pentafluorobenzyl esters. An ion trap mass spectrometer in selected reaction monitoring mode provided limits of detection of 0.1 ng/ml for isopropyl, isobutyl, pinacolyl and cyclohexyl methylphosphonic acids and for ethyl ethylphosphonic acid. The detection limit for ethyl methylphosphonic acid was higher (0.5 ng/ml) due to a lower recovery.

Selective extraction of hydrocarbons, phosphonates and phosphonic acids from soils by successive supercritical fluid and pressurized liquid extractions

Hydrocarbons, dialkyl alkylphosphonates and alkyl alkylphosphonic acids are selectively extracted from spiked soils by successive implementation of supercritical carbon dioxide, supercritical methanol–modified carbon dioxide and pressurized water. More than 95% of hydrocarbons are extracted during the first step (pure supercritical carbon dioxide extraction) whereas no organophosphorus compound is evidenced in this first extract. A quantitative extraction of phosphonates is achieved during the second step (methanol–modified supercritical carbon dioxide extraction). Polar phosphonic acids are extracted during a third step (pressurized water extraction) and analyzed by gas chromatography under methylated derivatives (diazomethane derivatization). Global recoveries for these compounds are close to 80%, a loss of about 20% occurring during the derivatization process (co-evaporation with solvent). The developed selective extraction method was successfully applied to a soil sample during an international collaborative exercise.

Rapid screening procedures for the hydrolysis products of chemical warfare agents using positive and negative ion liquid chromatography–mass spectrometry with atmospheric pressure chemical ionisation

Qualitative screening procedures have been developed for the rapid detection and identification of the hydrolysis products of chemical warfare agents in aqueous samples and extracts, using liquid chromatography–mass spectrometry with positive and negative atmospheric pressure chemical ionisation (APCI). Previously reported screening procedures, which used positive APCI or electrospray ionisation (ESI), were modified by using LC conditions that allowed acquisition of positive and negative ion mass spectra. APCI was generally found to be more robust than ESI, probably due to variable adduct ion formation with ESI, depending on the condition of the sample and the system. Negative APCI provided selective detection of acidic analytes and allowed facile differentiation of alkyl alkylphosphonic acids from isomeric dialkyl alkylphosphonates. The combination of positive and negative APCI, using a C 18 column and water–methanol mobile phase modified with ammonium formate, provides a rapid screening procedure for chemical warfare agent degradation products, with limits of detectability in the range 10–100 ng/ml. In the case of proficiency test samples, where analyte concentrations are in the range 1–10 ppm, introduction of the sample by infusion may provide an even faster preliminary screening procedure.

Identification of phosphonic acids by capillary electrophoresis–ionspray mass spectrometry

The identification of alkylphosphonic acids in spiked tap water has been investigated by on-line capillary electrophoresis–UV spectrometry–mass spectrometry (CE–UV–MS) in negative-ion mode. The 5 m M sorbic acid–ammonia electrolyte (pH 6.5) allows simultaneous indirect UV and MS (ionspray) detection. Several parameters (electrolyte pH, make-up chemical composition and make-up flow-rate) have been optimized and 5 mg/l limit of detection has been reached for these analytes in selected ion monitoring MS detection. MS–MS detection has also been investigated to reach a low detection limit (100 μg/l) for alkyl alkylphosphonic acids in spiked tap water. The mass spectra of these compounds exhibit a very abundant negative ion, [M−H] − (MS) with characteristic fragmentation (MS–MS) of acids and monoesters. Quantitative analysis achieved with CE–UV shows good correlation coefficients and allows accurate quantification of the analytes.

Application of liquid chromatography-atmospheric pressure chemical ionisation mass spectrometry, and tandem mass spectrometry, to the analysis and identification of degradation products of chemical warfare agents

A qualitative screening procedure was developed for the detection of the hydrolysis and related products of chemical warfare agents using liquid chromatography-mass spectrometry with atmospheric pressure chemical ionisation. A mixed C 8/C 18 reversed-phase column gave acceptable chromatography for the range of acidic, neutral and basic analytes. Detection limits for pure standards were less than 0.2 ng injected for the simple hydrolysis products of sulphur and nitrogen mustards, thiodiglycol sulphoxide, diisopropylaminoethanol and 3-quinuclidinol. Detection limits were in the range 0.2–8 ng injected for a series of alkylphosphonic acids and alkyl alkylphosphonic acids, and for benzilic acid. The methodology provides a rapid screening procedure for aqueous samples and extracts and was applied to the analysis of soil samples collected from bomb craters, and to spiked water and soil samples.