Chromatography-atmospheric Pressure Chemical Ionization-mass Spectrometry Research Articles

High-performance liquid chromatography–atmospheric-pressure chemical ionization mass spectrometry as a new tool for the determination of the mycotoxin zearalenone in food and feed

A new method for the determination of the mycotoxin zearalenone (ZON) in food and feed, based on HPLC–MS with an atmospheric-pressure chemical ionization (APCI) interface after extraction from cereals and clean-up by either conventional solid-phase or immunoaffinity cartridges is presented. The APCI interface parameters are optimized to provide detection of ZON with maximum sensitivity after RP separation of ZON on a C 18 column with acetonitrile–water (40:60, v/v) at 1 ml/min column flow without split. Using APCI-MS detection, the sensitivity of the method was improved by a factor of ca. 50 in comparison to HPLC with fluorescence detection, allowing determination of ZON down to 0.12 μg/kg maize which is well below present threshold values. Due to the selectivity of MS detection, it also was possible to quantitatively determine ZON both in raw extracts without clean-up using a normal-size (100 mm) chromatographic column or using only a short (20 mm) chromatographic column, when a clean-up was done to minimize possible interferences.

Evaluation of ELISA Kits Followed by Liquid Chromatography-Atmospheric Pressure Chemical Ionization-Mass Spectrometry for the Determination of Organic Pollutants in Industrial Effluents

Contaminated industrial effluents often contain a variety of organic pollutants which are difficult to analyze by standard GC-MS methods since they often miss the more polar or nonvolatile of these organic compounds. The identification of highly polar analytes by chemical or rapid biological techniques is needed for characterization of the effluents. The present work evaluates the use of enzyme linked immunosorbent assays (ELISA) kits for determining pentachlorophenol, carcinogenic PAHs and BTEX (benzene, toluene, ethylbenzene, and o-, m-, and p-xylene) among the organic analytes present in various industrial effluents from Europe. The analytical protocol applied for the evaluation of the kits was based on the use of ELISA followed by solid-phase extraction (SPE) for the preconcentration of a variety of organic pollutants such as pentachlorophenol, phthalates, and nonylphenol and final determination with LC-MS characterization using an atmospheric pressure chemical ionization (APCI) interface in the positive and negative ionization modes. The developed protocol permitted the unequivocal identification of target analytes such as pentachlorophenol, nonylphenol, dibutylphthalate, dimethylphthalate, bis(2-ethylhexyl)phthalate 2-methylbenzenesulfonamide, and 2,2-dimethylbenzene-sulfonamide present in industrial effluents. The advantages and limitations of the three RaPID-magnetic particle-based ELISA kits applied to the characterization of industrial effluents are also reported.

Determination of glucuronides of molecules of toxicological interest by liquid chromatography negative-ion mass spectrometry with atmospheric pressure chemical ionization

A new chromatographic method for the direct determination of metabolites (glucuronide-conjugates) of molecules of toxicological relevance in biological media with the minimum sample pre-treatment has been developed. A high performance liquid chromatographyatmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS) system was used for this purpose. The separation of four glucuronides Aminophenylglucuronide (APhG), Phenylglucuronide (PhG),p-Nitrophenylglucuronide (NPhG) and α-Naphthylglucuronide (NG) was obtained under ion-suppressed reversed-phase chromatography conditions, by using high-speed (3 cm, 3 μm) columns and formic acid (2 mM) as the acid modifier in the mobile phase. Different C-18 stationary phases (partially endcapped and non-endcapped) were evaluated in order to obtain retention for these very polar, water soluble molecules. The ionization of the analytes was obtained in negativeion (NI) mode. Detection limits were in the range 1–5 mg L−1 and calibration curves were linear over two order of magnitude. Intra-day and inter-day precision were in the range 2.9–10.6% for all the compounds. The method was successfully applied for the determination of PhG in a urine sample of a European Quality Assurance Programme for Organic Solvent Metabolites.

Development of a method for quantitation of retinol and retinyl palmitate in human serum using high-performance liquid chromatography–atmospheric pressure chemical ionization–mass spectrometry

A method for the quantitative analysis of the vitamin A compounds all-trans-retinol and all-trans-retinyl palmitate was developed using high-performance liquid chromatography–atmospheric pressure chemical ionization–mass spectrometry (APCI–LC–MS). Unlike previous quantitative mass spectrometric methods for vitamin A, HPLC separations were carried out using a C30 reversed-phase column instead of GC separation. Because no sample hydrolysis or derivatization was necessary, retinyl palmitate was preserved for analysis instead of being hydrolyzed to retinol. Human serum was analyzed following simple hexane extraction without saponification or any additional purification. A comparison of APCI and electrospray ionization showed that only APCI produced a linear response over all four orders of magnitude of retinol and three orders of magnitude of retinyl palmitate concentrations. Selected ion monitoring of the fragment ion of m/z 269 was used for APCI quantitation of both retinol and retinyl palmitate, since it was the base peak and the only abundant ion in the mass spectra of both compounds and the internal standard, retinyl acetate. The ion of m/z 269 corresponded to loss of water, loss of palmitic acid, or elimination of acetic acid from the protonated molecules of retinol, retinyl palmitate and retinyl acetate, respectively. The limit of detection of APCI–LC–MS for all-trans-retinol and all-trans-retinyl palmitate was determined to be approximately 34 fmol/μl and 36 fmol/μl (0.670 pmol all-trans-retinol and 0.720 pmol all-trans-retinyl palmitate injected in 20 μl on-column), respectively. The limit of quantitation was approximately 500 fmol/μl and 250 fmol/μl (10 pmol and 5 pmol injected in 20 μl on-column) for retinol and retinyl palmitate, respectively.

Monitoring of pesticides in river water using fully automated on-line solid-pase extraction and liquid chromatography with diode array detection with a novel filtration device

A modification of the SAMOS (System for the Automated Monitoring of Organic pollutants in Surface water) has been performed, which includes the attachment of four refrigerated flasks prior to the preconcentration unit, which act as sample reservoirs. River water is driven to the SAMOS, where it is pumped inside the flasks and afterwards it is preconcentrated onto C18 precolumns of the Prospekt (Spark Holland, Netherlands), which is coupled on-line with liquid chromatography using the HP1090 equipped with a diode array detector (Hewlett Packard, Waldbron, Germany). The system is programmed in such a way that eight samples are analyzed on each day and it can be operated unattended for at least five days. In order to prevent clogging of the equipment due to the presence of particulate matter, a novel filtering device, comprising a 50-cm stainless steel cylindrical tube with 10 μm pores, connected to a 190-mm diameter glass fiber filter with 1 μm pores was installed. This configuration permitted satisfactory performance, with the unattended monitoring of Llobregat river water and with limits of detection (LODs) of 30–100 ng/l. Terbuthylazine was detected in Llobregat river effluent at a concentration of 13 μg/l. Confirmatory analyses were performed by liquid chromatography–atmospheric pressure chemical ionization–mass spectrometry. This latter technique was also used to screen organophosphorus pesticides in the Llobregat river.

High-performance liquid chromatography and fluorometric detection of arachidonylethanolamide (anandamide) and its analogues, derivatized with 4-(N-chloroformylmethyl-N-methyl)amino-7-N,N-dimethylaminosulp honyl-2,1 ,3- benzoxadiazole (DBD-COCl).

The endogeneous ligand for the cannabinoid receptor, arachidonylethanolamide (anandamide) and its analogues, oleinylethanolamide, palmitylethanolamide and eicosapentaenoylethanolamide, were derivatized with a fluorogenic reagent, 4-(N-chloroformylmethyl-N-methyl)amino-7-N,N-dimethylaminsulpho ny1-2,1,3- benzoxadiazole (DBD-COCl). They were separated on a reversed phase HPLC with a mobile phase of acetonitrile:water. The fluorometric detection of the derivatives was made at 560 nm with excitation at 450 nm and the detection limits for anandamide was 20 fmol on column. The structures of DBD-CO-ethanolamides were confirmed by liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS).

Liquid chromatographic—mass spectrometric analysis of N-acetylamino acids in human urine

Liquid chromatography—atmospheric pressure chemical-ionization mass spectrometry (LC—APCI-MS) was used for the analysis of N-acetylamino acids that could not be determined with an amino acid analyzer. LC—APCI-MS could directly detect the protonated molecular ions of various synthetic N-acetylamino acids, distinguishing N-acetylserine from O-acetylserine and N(α)-acetyllysine from N(ε)-acetyllysine. Furthermore, N-acetylasparagine, N-acetylaspartic acid, N-acetylglutamine and N-acetylglutamic acid were identified in normal human urine. The assay data for N-acetylaspartic acid agree with a previous report using gas chromatography—mass spectrometry. These results demonstrate the usefulness of the apparatus described above for the analysis of N-acetylamino acids in biological samples.

Qualitative and semi-quantitative analyses of cytokinins using LC/APCI-MS in combination with ELISA

Application of liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC/APCI-MS) for the analysis of cytokinins was examined. The fragmentation of cytokinins was studied using authentic trans-zeatin (t-Z), trans-zeatin riboside (t-ZR), isopentenyl adenine (i6Ade), isopentenyl adenosine (i6Ado), benzyl adenine (BAde), benzyl adenosine (BAdo), and kinetin. These cytokinins were effectively ionized by APCI in aqueous acetonitrile. t-Z, i6Ade, BAde, and kinetin showed prominent quasi-molecular ions of [M + H]+, and ribosylcytokinins clearly showed both [M + H]+ and a characteristic fragment ion ([M + H-ribose]+), giving some information about their structures. The qualitative and semi-quantitative analyses of cytokinins by LC/APCI-MS were validated in combination with enzyme-linked immunosorbent assay (ELISA) through the analysis of t-ZR in the teratoma of Nicotiana tobacum. t-ZR was conclusively identified and a semi-quantitative estimate of its endogenous levels were provided by the combination of LC/APCI-MS and ELISA. The quantified values obtained by LC/ APCI-MS (single ion detection) and ELISA are in close agreement.