Rapid Communications in Mass Spectrometry

Maurel, Vincent; Ravanat, Jean‐Luc; Gambarelli, Serge

doi: 10.1002/rcm.2579pmid: 16810703

In this study, reactive free radicals derived from several nucleosides were spin trapped by 5,5‐dimethyl‐1‐pyrroline N‐oxide (DMPO) and then detected by high‐performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry (HPLC/ESI‐MS/MS). This method provides a specific detection of spin trapping adducts derived from nucleosides with a very high sensitivity: quantities as low as 0.5 picomoles of spin trapping adducts corresponding to concentrations of 2.5 10−8 mol . L−1 were detected. Different spin trapping adducts were characterized by HPLC/ESI‐MS/MS in three well‐known systems producing free radicals photochemically: the photolysis of 5‐halo‐2′‐deoxyuridines, the photolysis of 5‐thiophenylmethyl‐2′‐deoxyuridine and the photolysis of thymidine with menadione bisulfite as a photosensitizer. A new radical photoreactivity of uridine derivatives was also detected by this method both at the nucleoside and at the RNA level, showing that the method is also relevant for studying spin trapping adducts derived from DNA and RNA strands. Copyright © 2006 John Wiley & Sons, Ltd.

Assayag, Nelly; Rivé, Karine; Ader, Magali; Jézéquel, Didier; Agrinier, Pierre

doi: 10.1002/rcm.2585pmid: 16810706

We present here an improved and reliable method for measuring the concentration of dissolved inorganic carbon (DIC) and its isotope composition (δ13CDIC) in natural water samples. Our apparatus, a gas chromatograph coupled to an isotope ratio mass spectrometer (GCIRMS), runs in a quasi‐automated mode and is able to analyze about 50 water samples per day. The whole procedure (sample preparation, CO2(g)–CO2(aq) equilibration time and GCIRMS analysis) requires 2 days. It consists of injecting an aliquot of water into a H3PO4‐loaded and He‐flushed 12 mL glass tube. The H3PO4 reacts with the water and converts the DIC into aqueous and gaseous CO2. After a CO2(g)–CO2(aq) equilibration time of between 15 and 24 h, a portion of the headspace gas (mainly CO2+He) is introduced into the GCIRMS, to measure the carbon isotope ratio of the released CO2(g), from which the δ13CDIC is determined via a calibration procedure. For standard solutions with DIC concentrations ranging from 1 to 25 mmol · L−1 and solution volume of 1 mL (high DIC concentration samples) or 5 mL (low DIC concentration samples), δ13CDIC values are determined with a precision (1σ) better than 0.1‰. Compared with previously published headspace equilibration methods, the major improvement presented here is the development of a calibration procedure which takes the carbon isotope fractionation associated with the CO2(g)–CO2(aq) partition into account: the set of standard solutions and samples has to be prepared and analyzed with the same ‘gas/liquid’ and ‘H3PO4/water’ volume ratios. A set of natural water samples (lake, river and hydrothermal springs) was analyzed to demonstrate the utility of this new method. Copyright © 2006 John Wiley & Sons, Ltd.

Schänzer, Wilhelm; Geyer, Hans; Fußhöller, Gregor; Halatcheva, Natalia; Kohler, Maxie; Parr, Maria‐Kristina; Guddat, Sven; Thomas, Andreas; Thevis, Mario

doi: 10.1002/rcm.2587pmid: 16804957

Anabolic‐androgenic steroids are some of the most frequently detected drugs in amateur and professional sports. Doping control laboratories have developed numerous assays enabling the determination of administered drugs and/or their metabolic products that allow retrospectives with respect to pharmacokinetics and excretion profiles of steroids and their metabolites. A new metabolite generated from metandienone has been identified as 18‐nor‐17β‐hydroxymethyl,17α‐methyl‐androst‐1,4,13‐trien‐3‐one in excretion study urine samples providing a valuable tool for the long‐term detection of metandienone abuse by athletes in sports drug testing. The metabolite was characterized using gas chromatography/(tandem) mass spectrometry, liquid chromatography/tandem mass spectrometry and liquid chromatography/high‐resolution/high‐accuracy (tandem) mass spectrometry by characteristic fragmentation patterns representing the intact 3‐keto‐1,4‐diene structure in combination with typical product ions substantiating the proposed C/D‐ring structure of the steroid metabolite. In addition, structure confirmation was obtained by the analysis of excretion study urine specimens obtained after administration of 17‐CD3‐labeled metandienone providing the deuterated analogue to the newly identified metabolite. 18‐Nor‐17β‐hydroxymethyl,17α‐methyl‐androst‐1,4,13‐trien‐3‐one was determined in metandienone administration study urine specimens up to 19 days after application of a single dose of 5 mg, hence providing an extended detection period compared with commonly employed strategies. Copyright © 2006 John Wiley & Sons, Ltd.

Apollonio, Luigino G.; Whittall, Ian R.; Pianca, Dennis J.; Kyd, Jennelle M.; Maher, William A.

doi: 10.1002/rcm.2589pmid: 16810638

This paper describes the application of ultra‐performance liquid chromatography/tandem mass spectrometry (UPLC/MS/MS) technology to separate and identify amphetamine‐type substances (amphetamine, methamphetamine), common and novel designer analogues (MDA, MDMA, PMA, 4‐MTA, MBDB), and ketamine using Acquity UPLC/Micromass Quattro Micro API mass spectrometer instrumentation (Waters Corporation, USA). From injection of drug reference standards, it was demonstrated that these compounds can be identified by product ion mass spectra in less than 4 min total analysis time, indicating that the technological advancements associated with UPLC/MS/MS allow it to serve as a powerful analytical tool for high‐throughput testing. In addition to demonstrating the separation and response of these drug compounds under the stated UPLC/MS/MS conditions, we believe the acquired product ion spectra will be a beneficial reference to laboratories interested in incorporating the use of this technology in the routine analysis of drugs of abuse. Copyright © 2006 John Wiley & Sons, Ltd.

Attygalle, Athula B.; Kharbatia, Najeh; Bialecki, Jason; Ruzicka, Josef; Svatoš, Aleš; Stauber, Einar J.

doi: 10.1002/rcm.2582pmid: 16804956

The ion observed at m/z 145 when product ion spectra of iodobenzoate anions are recorded using ion‐trap mass spectrometers corresponds to the adduct ion [I(H2O)]−. The elements of water required for the formation of this adduct do not originate from the precursor ion but from traces of moisture present in the helium buffer gas. A collision‐induced decomposition (CID) spectrum recorded from the [M–H]− ion (m/z 251) derived from 3‐iodo[2,4,5,6‐2H4]benzoic acid also showed an ion at m/z 145. This observation confirmed that the m/z 145 is not a product ion resulting from a direct neutral loss from the carboxylate anion. 79Bromobenzoate anions produce similar results showing an ion at m/z 97 for [79Br(H2O)]−. The ion‐molecule reaction observed here is unique to ion‐trap mass spectrometers since a corresponding ion was not observed under our experimental conditions in spectra recorded with in‐space tandem mass spectrometers such as triple quadrupole or quadrupole time‐of‐flight instruments. Copyright © 2006 John Wiley & Sons, Ltd.

Fancy, Sally‐Ann; Beckonert, Olaf; Darbon, Gareth; Yabsley, Warren; Walley, Rosalind; Baker, David; Perkins, George L.; Pullen, Frank S.; Rumpel, Klaus

doi: 10.1002/rcm.2583pmid: 16810707

Seitz, Wolfram; Schulz, Wolfgang; Weber, Walter H.

doi: 10.1002/rcm.2554pmid: 16810705

Recent advances in the sensitivity of liquid chromatography/mass spectrometry (LC/MS) instrument technology provide the basis for the direct detection, i.e. without sample pre‐concentration, of organic contaminants in water in the ng/L range. Novel applications for the analysis of atrazine and some of its desalkylated and hydroxylated degradation products, the pharmaceutical compounds diclofenac and carbamazepine, sulfonylurea herbicides, and iodinated X‐ray contrast media have been developed. For each analyte a specific tandem mass spectrometric (MS/MS) transition has been selected and the corresponding mass spectrometric parameters optimised. All analytes could be analysed within three specific analytical runs including different high‐performance liquid chromatography (HPLC) conditions. Detection limits were determined to be better than 10 ng/L for the direct analysis of the compounds in water except for X‐ray contrast media, for which detection limits were found to be up to one order of magnitude higher. The methods have been successfully utilised for the analysis of natural waters. Matrix effects frequently occurring in LC/MS have shown to be low to moderate in the case of X‐ray contrast media. This work demonstrates that for the analysis of a large number of water contaminants, the sample pre‐concentration step could possibly be omitted. Copyright © 2006 John Wiley & Sons, Ltd.

Nirogi, Ramakrishna V. S.; Kandikere, Vishwottam N.; Shukla, Manoj; Mudigonda, Koteshwara; Maurya, Santosh

doi: 10.1002/rcm.2592pmid: 16810637

A simple, sensitive and rapid high‐performance liquid chromatography/positive ion electrospray tandem mass spectrometry (MS/MS) method was developed and validated for the assay of tizanidine in human plasma. Following liquid‐liquid extraction, the analytes were separated using an isocratic mobile phase on a reversed‐phase column and analyzed by MS/MS in the selected reaction monitoring mode. The assay exhibited a linear dynamic range of 50–5000 pg/mL for tizanidine in human plasma. The lower limit of quantification was 50 pg/mL with a relative standard deviation of less than 13%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 2.5 min for each sample made it possible to analyze more than 300 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies. Copyright © 2006 John Wiley & Sons, Ltd.

Schmidt, Anne‐Christine; Herzschuh, Rainer; Matysik, Frank‐Michael; Engewald, Werner

doi: 10.1002/rcm.2591pmid: 16810704

In order to develop a liquid chromatography/electrospray ionisation tandem mass spectrometry (LC/ESI‐MS/MS) method for identification and quantification of polar metabolites of explosives using a triple quadrupole system, the mass spectrometric ionisation and fragmentation behaviour of different nitrophenols, nitro‐ and aminonitrobenzoic acids, nitrotoluenesulfonic acids, and aminonitrotoluenes was investigated. Due to their different molecular structures, the substances concerned showed a very different ionisation efficiency in the ESI process. Interestingly, 2,4‐dinitrobenzoic acid yielded no mass signals in the Q1 scan suggesting a thermal decarboxylation in the ion source, whereas the corresponding 3,5‐isomer showed a high ionisation yield. Using negative ionisation polarity, carboxylic, phenolic, and sulfonic acid groups were deprotonated resulting in molecular anions, which could be fragmented in a collision cell. A pronounced dependency of the produced fragment ion series on the kind and position of substituents at the nitrobenzene ring (ortho effects) was observed and exploited for the development of substance‐specific detection methods in the multiple reaction monitoring mode. In case of benzoic and sulfonic acids, decarboxylation and desulfonation, respectively, were observed as the most frequent fragmentation reactions. Furthermore, besides loss of NO2, NO fragmentation occurred and preceded a decarbonylation of the benzene ring. The expulsion of the open‐shell molecules NO and NO2 led to a variety of distonic radical anions. Copyright © 2006 John Wiley & Sons, Ltd.

Wang, Su Jun; Zhang, Zhen Qing; Zhao, Yan Hong; Ruan, Jin Xiu; Li, Jing Lai

doi: 10.1002/rcm.2590pmid: 16810636

A simple, rapid and sensitive method was developed for the simultaneous quantification of chlorogenic acid (CGA) and caffeic acid (CA) in rat plasma using a high‐performance liquid chromatography system coupled to a negative ion electrospray mass spectrometric analysis. The plasma sample preparation was a simple deproteinization by the addition of two volumes of acetonitrile followed by centrifugation. The analytes and internal standard ferulic acid were separated on an Intersil C8‐3 column (5 mm; 250 × 2.1 mm) with acetonitrile/0.05% triethylamine solution (70:30, v/v) as mobile phase at a flow rate of 0.2 mL/min with an operating temperature of 30°C. Detection was performed on a quadrupole mass spectrometer equipped with an electrospray ionization (ESI) source operated in selected ion monitoring (SIM) mode. Negative ion ESI was used to form deprotonated molecules at m/z 353 for chlorogenic acid, m/z 179 for caffeic acid, and m/z 193 for the internal standard ferulic acid. Linear detection responses were obtained for CGA concentrations ranging from 0.005 to 2.0 µg/mL and for CA concentrations ranging from 0.010 to 2.0 µg/mL and the lower limits of quantitation (LLOQs) for CGA and CA were 0.005 and 0.01 µg/mL, respectively. The intra‐ and inter‐day precisions (RSD%) were within 9.0% for both analytes. Deviation of the assay accuracies was within ±10.0% for both analytes. Their average recoveries were greater than 88.0%. Both analytes were proved to be stable during all sample storage, preparation and analytic procedures. The method was successfully applied to the pharmacokinetic study of CGA and CA following an intravenous dose of 5 mL/kg mailuoning injection to rats. Copyright © 2006 John Wiley & Sons, Ltd.