A. V. Braun

A validated LC-MS/MS method for rapid determination of methotrexate in human saliva and its application to an excretion evaluation study

A sensitive and simple method for the methotrexate quantification was developed using aminopterin as internal standard. Methotrexate is an anticancer agent that is widely used in a variety of human cancers including primary central nervous system lymphoma. The compound was quantified by liquid-chromatography coupled to electrospray ionization (positive ion-mode) low-energy collision dissociation-tandem mass spectrometry. Quantitative detection was by multiple reaction monitoring of the transitions of the [M+H]+ ion of MTX to its common product ion at m/z 308.4 and of aminopterin at m/z 441.2→m/z 294.0. The method demonstrated linearity over at least three orders of magnitude and had a detection limit of 1ng/ml for methotrexate. A run time of less than 8.0min for each sample made it possible to analyze a large number of human saliva samples per day. Application of this procedure was demonstrated to a saliva excretion study of methotrexate on the samples obtained after an intravenously administration of 1mg/kg/dose of methotrexate to six patients with acute lymphoblastic leukemia.

RAPID METHOD FOR THE DETECTION OF METABOLITE OF SULFUR MUSTARD 1,1′-SULFONYLBIS[2-S-(N-ACETYLCYSTEINYL)ETHANE] IN PLASMA AND URINE BY LIQUID CHROMATOGRAPHY-NEGATIVE ELECTROSPRAY-TANDEM MASS SPECTROMETRY

A method for detection of an important biological marker, 1,1′-sulfonylbis[2-S-(N-acetylcysteinyl)ethane] (SBSNAE) of sulfur mustard agent [bis-(2-chloroethyl)sulfide] (HD) in human plasma and rat urine to quantify HD exposure, is presented. It employs solid-phase extraction sample preparation, followed by liquid-chromatography-negative ion-electrospray ionization-tandem mass spectrometry. The method limit of detection (LOD) for SBSNAE is 0.05 ng/mL for rat urine samples with relative standard deviations of less than 10%. Application of this procedure was demonstrated in the HD animal exposure model. Rats were exposed intravenously to 5 mg/kg HD, and SBSNAE levels in urine samples were analyzed for 22 days post-exposure.

Lewisite metabolites detection in urine by liquid chromatography–tandem mass spectrometry

A sensitive and simple method for the quantification and for the detection of 2-chlorovinylarsonous (CVAA) and 2-chlorovinylarsonic (CVAOA) acids was developed. CVAA and CVOA are important biological markers in human and rat urine specific to lewisite (chlorovinylarsonous chloride compounds) exposure. The developed assay was based on the use of solid-phase extraction (SPE) followed by liquid-chromatography coupled to electrospray ionization (negative ion-mode) low-energy collision dissociation-tandem mass spectrometry (ESI-CID-MS/MS). The method demonstrated linearity over at least three orders of magnitude and had a detection limit (LOD) of 0.5 ng/ml for CVAA and 3 ng/ml for CVAOA. The relative standard deviations for the quality control samples ranged from 6 to 11%. Application of this procedure was demonstrated in the lewisite animals exposure model. Rats were exposed intravenously by no lethal doses of lewisite and markers levels in urine samples were analyzed for 21 days post-exposure.

Hydrophilic interaction liquid chromatography − tandem mass spectrometry methylphosponic and alkyl methylphosphonic acids determination in environmental samples after pre-column derivatization with p-bromophenacyl bromide

Once exposed to the environment organophosphate nerve agents readily degrade by rapid hydrolysis to the corresponding alkyl methylphosphonic acids which do not exist in nature. These alkyl methylphosphonic acids are finally slowly hydrolyzed to methylphosphonic acid. Methylphosphonic acid is the most stable hydrolysis product of organophosphate nerve agents, persisting in environment for a long time. A highly sensitive method of methylphosphonic acid and alkyl methylphosphonic acids detection in dust and ground mixed samples has been developed and validated. The fact that alkyl methylphosphonic acids unlike methylphosphonic acid did not react with p-bromophenacyl bromide under chosen conditions was discovered. This allowed simultaneous chromatographic separation and mass spectrometric detection of derivatized methylphosphonic acid and underivatized alkyl methylphosphonic acids using HILIC-MS/MS method. Very simple sample pretreatment with high recoveries for each analyte was developed. Methylphosphonic acid pre-column derivate and alkyl methylphosphonic acids were detected using tandem mass spectrometry with electrospray ionization after hydrophilic interaction liquid chromatography separation. The developed approach allows achieving ultra-low detection limits: 200 pg mL(-1) for methylphosphonic acid, 70 pg mL(-1) for ethyl methylphosphonic acid, 8 pg mL(-1) for i-propyl methylphosphonic acid, 8 pg mL(-1) for i-butyl methylphosphonic acid, 5 pg mL(-1) for pinacolyl methylphosphonic acid in the extracts of dust and ground mixed samples. This approach was successfully applied to the dust and ground mixed samples from decommissioned plant for the production of chemical weapons.

Single-run HPLC/ESI-LITMS profiling of ginsenosides in plant extracts and ginseng based products.

A rapid single-run analytical approach suitable to achieve a comprehensive characterization of ginsenosides - the main bioactive compounds present in plant materials from Panax species and ginseng-based products - was developed. The method is based on high-performance liquid chromatography coupled with electrospray positive ionization linear ion trap mass spectrometry (HPLC/ESI-LITMS). The main ions in the ESI-LITMS spectra were attributed to molecular adducts with sodium and potassium and fragments corresponding to cleavage of the glycosidic bonds. The simplicity of the approach allows laborious sample preparation and sophisticated spectral information-dependent acquisition to be avoided, and provides an opportunity for rapid screening. The method may replace existing HPLC-DAD profiling approaches. The results of this study indicate that HPLC/ESI-LITMS is applicable for quality control purposes on processed products and allows the rapid and direct identification of ginsenosides in crude plant extracts.

Detection of nerve agent markers by liquid chromatography-mass spectrometry

An approach to the detection of metabolites of organophosphorous agents (OPA), such as O-iso-propylmethylphosphonic acid (detection limit, 4 ng/mL), O-pinacolylmethylphosphonic acid (0.6 ng/mL), and O-isobutylmethylphosphonic acid (1 ng/mL), in plasma samples was developed using liquid chromatography-mass spectrometry. The curves of the elutionxcretion of OPA metabolites were obtained for the samples of biological material of rats exposed to toxic substances. Determination was performed by mass spectrometry with electrospray ionization in the negative ion mode, using deprotonated molecules for detection. The biological samples were analyzed by reversed-phase chromatography using hydrophilic end-capped adsorbents. Solid phase extraction on reversed-phase adsorption cartridges containing a copolymer of styrene and divinylbenzene was proposed for sample preparation.

The use of linear ion trap for qualitative analysis of phytochemicals in Korean ginseng tea

A new approach to qualitative analysis of ginsenosides in challenging matrices was developed on the basis of high-performance liquid chromatography/tandem mass spectrometry. Using the extracts from samples of ginseng tea, the approach was validated. Analysis of extracts was carried out using a reversed-phase chromatography with SB-C18 sorbent. For compound identification, electrospray ionization and a quadrupole/linear ion trap mass-spectrometer in different modes were used. A meticulous study of the fragmentation of ginsenosides in the linear ion trap and its application for analysis of these compounds was performed in this work. The accuracy of the identification was proven with standards of ginsenosides Rb1, Rg1, Re, Rf, Rd, Rb2, Rb3 and Rc.

Simultaneous determination of salidroside, rosavin, and rosarin in extracts from Rhodiola rosea by high performance liquid chromatography with tandem mass spectrometry detection

AbstractsAn approach is developed for the simultaneous determination of salidroside, rosarin, and rosavin in extracts from plant based on liquid chromatography/mass spectrometry; the detection limits of the method are 2, 4, and 6 ng/mL for salidroside, rosavin, and rosarin, respectively. The method was validated using real extracts from Rhodiola rosea rhizome. The analysis of extracts was accomplished by reversed-phase chromatography with a SB-C18 stationary phase. The analytes were detected by tandem mass spectrometry with electrospray ionization in the multiple reaction monitoring mode for negative ions.

Modern methods of identifying and determining ginsenosides

Ginsenosides are the main bioactive compounds of the Panax plant genus. Ginseng and its analogues are widely used to produce traditional medicines in China, Korea, Japan, United States, and the Russian Far East. For more than 40 years, many researchers developed methods of identifying and determining and ginsenosides in plant tissues, extracts, and commercial products. Various extraction methods were used to isolate these compounds from plant materials. The separation of ginsenosides was conducted with methods such as gas chromatography, thin-layer chromatography, and high-performance liquid chromatography (HPLC). The HPLC method was used predominantly. Spectrophotometric and fluorescent monitoring and, later, light scattering and mass-spectrometry coupled with HPLC were used to determine ginsenosides. The most recent variants of these methods are presented in this review, together with a critical evaluation of the published results.

Rapid IC–MS/MS determination of methylphosphonic acid in urine of rats exposed to organophosphorus nerve agents

A direct approach for the determination of a specific hydrolysis product of organophosphorus nerve agents such as methylphosphonic acid (MPA) in urine by ion chromatography and tandem mass spectrometry (IC-MS/MS) has been developed. The first advantage of the proposed approach is a rapid and simple sample preparation, which does not require a large sample volume, complicated and laborious preconcentration and derivatization steps, and takes less than 7min per sample. The second advantage is the fast and selective IC determination of MPA carried out on a noncommercial anion exchanger based on a poly(styrene-co-divinylbenzene) (PS-DVB) substrate with a high degree of crosslinking and a covalently-bonded branched functional layer, which enables complete resolution of MPA from major urine matrix components and allows one to overcome matrix effects. Hyphenation of IC with tandem mass spectrometry results in highly sensitive and reliable MPA determination with the lowest detection limit (4ngmL-1) reported so far for HPLC determination of MPA in urine. The proposed approach is successfully applied for the analysis of urine from rats exposed to nonlethal doses of organophosphorus nerve agents such as sarin, soman, and VR in up to 13days after initial exposure, which shows the possibility to verify the nerve agent exposure after a long period of time.