Chromatography-atmospheric Pressure Chemical Ionization-tandem Mass Research Articles

Quantitative analysis of selegiline and three metabolites (N-desmethylselegiline, methamphetamine, and amphetamine) in human plasma by high-performance liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry.

This report describes a sensitive and specific high-performance liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry method for the detection of subnanogram concentrations of selegiline and its three principle metabolites, N-desmethylselegiline, methamphetamine, and amphetamine, in human plasma. The assay has a dynamic range of 0.1-20 ng/mL for selegiline and N-desmethylselegiline (norselegiline) and 0.2-20 ng/mL for methamphetamine and amphetamine. The inter- and intra-assay precision and accuracy varied by less than 11% for all analytes at 0.3, 2.5, and 15 ng/mL and less than 16% at the lower limit of quantitation (0.1 ng/mL for selegiline and norselegiline; and 0.2 ng/mL for methamphetamine and amphetamine). Selegiline and its metabolites showed no significant loss in quantitative accuracy after three freeze/thaw cycles or after up to 6 h at room temperature prior to extraction. Extracted plasma samples retained quantitative accuracy after storage for at least 7 days at -20 degrees C or up to 70 h at room temperature. Methanolic stock solutions were stable for at least 6 h when kept at room temperature or at least 90 days when kept at -20 degrees C.

Assay of TNP-470 and its two major metabolites in human plasma by high-performance liquid chromatography-mass spectrometry.

In this study, a sensitive, specific assay for the determination of TNP-470 and its two major metabolites M-IV (also know as AGM-1883) and M-II in human plasma is reported. The assay involves liquid-liquid extraction of acidified plasma followed by reversed-phase high-performance liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry. A liquid-liquid extraction using an organic solvent mixture (methyl-tert-butyl-ether-hexane, 1:1, v/v) is used in place of solid-phase extraction because it provides consistent recoveries for all analytes, including the internal standard. Retention times for the analytes and internal standard are less than 7 min. Within- and between-day precision is < or = 6.5% and < or = 13.3% relative standard deviation, respectively, for the three analytes. The lower limits of quantitation are 0.25, 0.5, and 1.0 ng/mL for TNP-470, M-IV, and M-II, respectively.

Simultaneous analysis of flunitrazepam and its major metabolites in human plasma by high performance liquid chromatography tandem mass spectrometry

A sensitive and specific high performance liquid chromatography–atmospheric pressure chemical ionization–tandem mass spectrometry (HPLC–APCI–MS–MS) method has been developed for the simultaneous determination of flunitrazepam and its major metabolites, 7-aminoflunitrazepam and N-desmethylflunitrazepam, in human plasma. After the addition of a deuterium labelled internal standard of flunitrazepam, plasma samples were extracted using Oasis® MCX solid phase extraction cartridges. The compounds were separated on a 5 μm Symmetry C18 (Waters) column (3.0×150 mm, i.d.) with a step gradient of acetonitrile-0.1% formic acid at a flow rate of 0.6 ml/min. The overall extraction efficiency was more than 89% for all three compounds. The limits of detection were 0.25 g/l for flunitrazepam, 0.5 μg/l for 7-aminoflunitrazepam, and 2.0 μg/l for N-desmethylflunitrazepam. Within-run accuracies for quality-control samples were between 92.5 and 101.3% of the target concentration, with coefficients of variation <8%. The proposed method enables the unambiguous identification and quantitation of flunitrazepam and its major metabolites in both clinical and forensic specimens.

A Reference Method for the Analysis of Aldosterone in Blood by High-Performance Liquid Chromatography–Atmospheric Pressure Chemical Ionization–Tandem Mass Spectrometry

A high-performance liquid chromatography–atmospheric pressure chemical ionization–tandem mass spectrometry (HPLC–APCI–MS/MS) reference method for the quantitation of aldosterone in serum and plasma has been developed. Samples were extracted with dichloromethane/diethyl ether, containing flumethasone as internal standard (IS). Chromatography was performed on a phenyl column using 50 mmammonium formate (pH 7.1)/methanol (50/50, v/v) as mobile phase. Analysis was in negative-ionization mode by selected reaction monitoring (aldosteronem/z359.2 → 331.2; ISm/z455.0 → 379.0). The assay was linear over the range 15–500 pg/mL, with limits of detection and quantitation of 10 and 15 pg/mL, respectively. Imprecisions of the assay at 15, 20, 150, and 450 pg/mL were 18.5, 8.8, 10.6, and 9.5%, respectively. The accuracy of the method ranged from 93.1 to 98.9% with absolute recoveries between 84.0 and 91.3% (aldosterone) and 88.0 and 92.3% (IS). We present a case study of a patient admitted, with suspected primary hyperaldosteronism, on the basis of a high radioimmunoassay (RIA) aldosterone concentration. The results suggest that RIA was unreliable, causing unnecessary patient discomfort and a costly 6-day hospital stay. The specific HPLC–API–MS/MS assay described offers the sensitivity and accuracy required to assess abnormal aldosterone production in hypertensive patients.

2,5-Dimethyl-4-hydroxy-3[2H]-furanone [formula omitted] in strawberry fruits

2,5-Dimethyl-4-hydroxy-3[2H]-furanone 6′-O- malonyl-β- d-glucopyranoside was isolated from a glycosidic extract of strawberry fruit ( Fragaria x ananassa, cv. Senga Sengana) by means of countercurrent chromatography and reverse-phase HPLC. Identification was achieved by comparison of chromatographic and 1H, 13C and 2D-NMR, as well as mass spectral data with those of the synthesized reference compound. In ripe strawberry fruit, a 1:1 ratio of the malonylated glucoside to its deacylated glucoconjugate was determined by on-line liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry.