A major challenge in selecting an appropriate stationary phase for diastereomeric separation is that it is difficult to predict which of the commercially available stationary phases could achieve the required liquid chromatographic (LC) separation. This work describes the selection and evaluation of a porous graphitic carbon (PGC) column coupled with tandem mass spectrometry (MS/MS) for the simultaneous quantitation of an experimental drug candidate (I), its two diastereomeric metabolites (II and III), and its demethylated metabolite (IV) in rat plasma. In addition, we investigated the PGC column for the separation of another drug candidate (VI), its two diastereomeric metabolites (VII and VIII) and its ketone metabolite (IX). The PGC column showed excellent chromatographic resolution for the two diastereomers II and III, as well as for VII and VIII. In contrast, the required resolution for the diastereomers II and III could not be achieved using silica-bonded C(18), C(30), phenyl, perfluorinated, polar embedded and polar end-capped phases. The PGC column showed ruggedness with excellent reproducibility of retention times, peak symmetry and response over a period of more than 400 injections of a plasma acetonitrile-precipitation extract. Excellent accuracy and precision were achieved, with accuracy of 94-108% and intra- and inter-run precision within 9%. This work indicates that PGC is a valuable addition to the repertoire of LC columns used for quantitative LC/MS/MS bioanalysis, especially where the separation and quantitation of diastereomeric analytes is involved.

A sensitive, simple and rapid high performance liquid chromatography-tandem mass spectrometry (HPLC–MS/MS) method was developed and fully validated for the simultaneous quantification of buprenorphine (BUP) and its N-dealkylated metabolite norbuprenorphine (NBUP) in 200 μL human plasma. Human plasma samples were prepared using liquid–liquid extraction, and then separated on a Shiseido MG C18 (5 μm, 2.0 mm×50 mm) via 4.1 min gradient elution. Following electrospray ionization, the analytes were quantified on a triple–quadrupole mass spectrometer in multiple-reaction-monitoring (MRM) positive ion mode. Linearity was achieved from 25.0 to 10000 pg/mL for buprenorphine, from 20.0 to 8000 pg/mL for norbuprenorphine with r2>0.99. The method was demonstrated with acceptable accuracy, precision and specificity for the detection of buprenorphine and norbuprenorphine. Recovery was 81.8–88.8% for buprenorphine and 77.0–84.6% for norbuprenorphine, and the matrix effect was 95.6–97.4% for buprenorphine and 94.0–96.9% for norbuprenorphine; all were not concentration dependent. With validated matrix and autosampler stability data, this method was successfully applied in a bioequivalence study to support abbreviated new drug application.

A sensitive and high-throughput chiral liquid chromatography-tandem mass spectrometry method was developed and validated for the quantification of R-pantoprazole and S-pantoprazole in human plasma. Sample extraction was carried out by using ethyl acetate liquid-liquid extraction in 96-well plate format. The separation of pantoprazole enantiomers was performed on a CHIRALCEL OJ-RH column and an overlapping injection mode was used to achieve a run time of 5.0 min/sample. The mobile phase consisted of 1) 10 mM ammonium acetate in methanol: acetonitrile (1:1, v/v) and 2) 20 mM ammonium acetate in water. Isocratic elution was used with flow rate at 500 μL/min. The enantiomers were quantified on a triple-quadrupole mass spectrometer under multiple reaction monitoring (MRM) mode with m/z 382.1/230.0 for pantoprazole and m/z 388.4/230.1 for pantoprazole-d7. Linearity from 20.0 to 5000 ng/mL was established for each enantiomer (r(2) > 0.99). Extraction recovery ranged from 91.7% to 96.4% for R-pantoprazole and from 92.5% to 96.5% for S-pantoprazole and the IS-normalized matrix factor was 0.98 to 1.07 for R-pantoprazole and S-pantoprazole, respectively. The method was demonstrated with acceptable accuracy, precision, selectivity, and stability and the method was applied to support a pharmacokinetic study of a phase I clinical trial of racemic pantoprazole in healthy Chinese subjects. Chirality 28:569-575, 2016. © 2016 Wiley Periodicals, Inc.

No abstract

No abstract