Monitoring Mass Spectrometry Research Articles

A method for quantifying the unstable and highly polar drug nafamostat mesilate in human plasma with optimized solid-phase extraction and ESI-MS detection: more accurate evaluation for pharmacokinetic study

An advanced quantification method was developed with solid-phase extraction (SPE) and mass spectrometry (MS) determination for nafamostat, an unstable and highly polar drug, in human plasma. For unstable drugs with an ester group, the main analytical challenge is how to avoid the ester hydrolysis, and strong acid or alkaline conditions should be excluded during sample preparation. Considering that, we developed a relatively mild method with SPE for sample preparation without strong acid and alkaline treatment, which was optimized with different pHs and salt concentrations in phosphate-buffered saline treatment. The results indicated that pH 5 gave the most efficient extraction and 0.1 M salt concentration enhanced the extraction the most, with a minor effect on MS monitoring. The extraction method effectively avoided drug hydrolysis and achieved good drug enrichment over 82.2%. The linear range of quantification was 1.25-160 ng mL(-1). The stability of the drug in sample treatment was fully validated according to the sample processing procedure, including the stability in fresh blood, mobile phase, plasma and acidic methanol, and the results indicated that the drug remained stable during the whole sample preparation. Compared with a previous isotope-labeling method, more accurate and specific quantification of plasma concentration was achieved with liquid chromatography-electrospray ionization MS determination. With use of our method, nafamostat mesilate pharmacokinetics in 30 Chinese healthy volunteers was investigated with three doses via intravenous-drip infusion. The pharmacokinetic parameters were also estimated and compared with those of Japanese volunteers (slightly lower plasma concentration and longer terminal elimination half-life for Chinese volunteers). The difference in the pharmacokinetics may be ascribed to the quantification method, because previous isotope labeling may have overestimated the parent drug.

LC–APCI–MS–MS for the Determination of Celastrol in Human Whole Blood

A rapid, sensitive high-performance liquid chromatographic tandem mass spectrometric (HPLC–MS–MS) assay has been developed and validated for the quantitative determination of celastrol in human whole blood using hydrocortisone as an internal standard (I.S.). The celastrol and I.S. were extracted from human whole blood with ethyl acetate. The separation was performed by reversed-phase HPLC using an isocratic mobile phase consisting of 5 mmol L−1 aqueous ammonium acetate containing 0.05% acetic acid/methanol (25:75, v/v) on a XDB-C18 column (150 mm × 4.6 mm I.D., 5 μm). Detection was by negative atmospheric pressure chemical ionization (APCI) and ion trap tandem mass spectrometry in multiple reaction monitoring (MRM) mode with a transition of m/z 449.4 → 405.1 for celastrol, and 419.2 → 329.1 for I.S. The calibration curve was linear (r 2 = 0.9967) in the concentration range of 1.0–200.0 ng mL−1 in human whole blood with a limit of quantification of 1.0 ng mL−1. Intra-day and inter-day relative standard deviations (RSDs) were less than 8.5 and 10.1%. The mean extraction recovery was 89.2% for celastrol and 92.6% for I.S. This assay can be used to determine trace celastrol in human whole blood.

High-performance liquid chromatography–mass selective detection assay for adenine released from a synthetic RNA substrate by ricin A chain

High-performance liquid chromatography (HPLC) and selected ion monitoring mass spectrometry (MS) were used to develop a quantitative assay for adenine released from a synthetic RNA substrate by ricin A chain, which contains the toxin’s N-glycosidase activity. Because ricin and ricin A chain have potential applications as biotherapeutics and bioweapons, assays are needed to evaluate potency and potential inhibitors of activity. The detection limit for adenine was 0.02 μM (2.4 ng/ml), and the standard curve was linear up to 27.3 μM. The lower limit of quantitation was 0.27 μM and was reproducible throughout this range. Reaction characterization showed that most adenine was released by 5 h and that the reaction could not be fully stopped with formic acid concentrations up to 0.75 mM (the maximum typically used for HPLC-MS). Injections were made at 2-min intervals, 10 injections could be performed before the column was backflushed, and no ricin A chain was observed in the column effluent. This assay would also be useful for ricin since ricin A chain did not pass through the HPLC column. With minor modifications to this system, the assay should provide rapid, sensitive, selective, and quantitative assessment of the activitity of most ribosome-inactivating proteins. In addition, further chromatographic and mass spectrometric improvements could reduce sample requirements and analysis times.

Identification of cytochrome P450 enzymes involved in the metabolism of 4′-methoxy-α-pyrrolidinopropiophenone (MOPPP), a designer drug, in human liver microsomes

1. The metabolism of 4′-methoxy-α-pyrrolidinopropiophenone (MOPPP), a novel designer drug, to its demethylated major metabolite 4′-hydroxy-pyrrolidinopropio-phenone (HO-PPP) was studied in pooled human liver microsomes (HLM) and in cDNA-expressed human hepatic cytochrome P450 (CYP) enzymes.2. CYP2C19 catalysed the demethylation with apparent Km and Vmax values of 373.4 ± 45.1 μM and 6.0 ± 0.3 pmol min−1 pmol−1 CYP, respectively (mean ± SD). Both CYP2D6 and HLM exhibited clear biphasic profiles with apparent Km,1 values of 1.3 ± 0.4 and 22.0 ± 6.5 μM, respectively, and Vmax,1 values of 1.1 ± 0.1 pmol min−1 pmol−1 CYP and 169.1 ± 20.5 pmol min−1 mg−1 protein, respectively.3. Percentages of intrinsic clearances of MOPPP by particular CYPs were calculated using the relative activity factor (RAF) approach with (S)-mephenytoin-4′-hydroxylation or bufuralol-1′-hydroxylation as index reactions for CYP2C19 or CYP2D6, respectively.4. MOPPP, HO-PPP and the standard 3′,4′-methylenedioxy-pyrrolidinopropio-phenone (MDPPP) were separated and analysed by liquid chromatography–mass spectrometry in the selected-ion monitoring (SIM) mode.5. The CYP2D6 specific chemical inhibitor quinidine (3 μM) significantly ( p<0.0001) inhibited HO-PPP formation by 91.8 ± 0.5% (mean ± SEM) in incubation mixtures with HLM and 2 μM MOPPP.6. It can be concluded from the data obtained from kinetic and inhibition studies that polymorphically expressed CYP2D6 is the enzyme mainly responsible for MOPPP demethylation.

Comparison of UV and tandem mass spectrometric detection for the high-performance liquid chromatographic determination of diclofenac in microdialysis samples

High-performance liquid chromatography (HPLC) was used to analyze microdialysis samples obtained in vivo from human subcutaneous adipose tissue after topical application of the nonsteroidal anti-inflammatory drug diclofenac. For the reliable determination of diclofenac two different detection principles were applied in two different laboratories. One HPLC method utilized UV-detection at 280 nm, the other one used selected reaction monitoring mass spectrometry (MS). The HPLC-UV and -MS methods offered low limits of quantification of 10 and 1 ng/ml and an accuracy between 94.0–126.7 and 89.3–110.9%, respectively. However, a comparison showed that the HPLC-UV method failed to determine diclofenac in biological matrices, as both false negative and positive values were found. HPLC-MS is clearly superior to HPLC-UV due to a much more selective detection, increased sensitivity and shorter run times.

Application of high performance liquid chromatography/tandem mass spectrometry in the environmental and biological monitoring of health care personnel occupationally exposed to cyclophosphamide and ifosfamide

Application of high performance liquid chromatography/tandem mass spectrometry in the environmental and biological monitoring of health care personnel occupationally exposed to cyclophosphamide and ifosfamide

Rapid semi-on-line monitoring of Fmoc solid-phase peptide synthesis by matrix-assisted laser desorption/ionization mass spectrometry

A simple yet highly effective application of matrix-assisted laser desorption/ionization massspectrometry (MALDI-MS) for the rapid monitoring of Fmoc solid-phase peptide synthesisis described. A few beads of the resin are removed at any desired step during synthesis, thefully protected peptide is cleaved from the resin and an MS spectrum of the analytes presentis produced. Some standard side-chain protecting groups may be cleaved off during samplepreparation for MS analysis; however, these cleavages are readily identified. Using thisapproach, incomplete amino acid acylations are readily detected in approximately the sametime as by traditional tests such as ninhydrin. The semi-on-line method also lends itself toready optimization of synthesis protocols and to the examination of resin-bound peptide sidereactions which may not be detectable by chemical means.

Preconcentration of triazine herbicides from water by an ion chromatography column and determination by gas chromatography-mass spectrometry

The efficiency of ion chromatography columns packed with styrene-divinylbenzene copolymer containing quaternary ammonium groups to preconcentrate triazine herbicides and their degradation products below μg/l levels has been established. Retention is studied for different types of water. Pure methanol was used in a one-step elution. Enrichment factors of at least 4000 are achieved. Determination was carried out by using gas chromatography-single-ion monitoring mass spectrometry. Recoveries for run-off agricultural water were between 67–100% and close to 100% for ground water. The maximum admissible concentration ion drinking water (0.1 μg/l) and the alert and alarm threshold values in surface water (1 and 3 μg/l, respectively) dictated by the European Union can be measured.

Enantiomeric resolution of persistent compounds of technical toxaphene (CTTs) on t-butyldimethylsilylated β-cyclodextrin phases

Eight of the most important single compounds of technical toxaphene were separated on t-butyldimethylsilylated β-cyclodextrin (β-BSCD) diluted in a medium polar phase using gas chromatography with electron capture detectors (GC-ECD). The enantiomeric resolution of all compounds was obtained in one GC run. The β-BSCD phase also separated the enantiomers of oxychlordane, cis- and trans-chlordane as well as α-HCH. Problems in the enantioselective determination of CTTs in biological samples are discussed. Finally, the enantioselective determination of the two most recalcitrant CTTs in biological samples was achieved using electron capture negative ionization mass spectrometry (GC-ECNI-MS) in the single ion monitoring (SIM) mode.

Determination of 9,10-epoxystearate ester in intravenous solutions stored in poly(vinyl chloride) bags, using gas chromatography-single-ion monitoring mass spectrometry.

Determination of 9,10-epoxystearate ester in intravenous solutions stored in poly(vinyl chloride) bags, using gas chromatography-single-ion monitoring mass spectrometry.

An automated system for simultaneous thermal analysis and mass spectrometry. Part 1

An automated system which permits differential thermal analysis (DTA), thermogravimetry (TG), derivative thermogravimetry (DTG) and mass spectrometry (MS) measurements to be performed simultaneously on a test sample under an inert or oxidative atmosphere has been developed. The system involves interfacing a Mettler thermoanalyzer, model TA-1, with a Hewlett-Packard 5992 quadrupole mass spectrometer. Data acquisiton from the thermoanalyzer and the mass spectrometer is handled by two separate Hewlett-Packard 9825 calculators with the mass spectral data stored on a floppy disk. Computer software has also been developed to permit: (1) continuous MS monitoring of volatiles, (2) screening of the acquired MS data, (3) tabulation of mass spectra, (4) subtraction of mass spectra acquired at two different times and (5) thermograms and mass ion profiles plotted together as a function of temperature or time.

Some simple modifications to the LKB 9000S to achieve improved selected ion monitoring and repetitive scanning performance.

Modifications are described to the electron multiplier pre-amplifier, selected ion monitor, magnet current supply and scan controller of the LKB 9000S gas chromatograph mass spectrometer. The modifications described have secured an improved performance for specific ion monitoring and fast repetitive scanning gas chromatography mass spectrometry.

The Use of Mass Spectrometry for Off-Line Monitoring of Liquid Chromatography

The Use of Mass Spectrometry for Off-Line Monitoring of Liquid Chromatography