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Abstract: A sensitive and selective liquid chromatographic–tandem mass spectrometric (LC–MS/MS) method was developed for the determination of serial chiral novel anticholinergic compounds of phencynonate in rat plasma. After a simple protein-precipitation using methanol, the post-treatment samples were separated on a CAPCELL UG120 column with a mobile phase of a mixture of methanol and water (35:65) containing 0.1% formic acid. The serial chiral analytes and internal standard (IS) were all detected by the use of selected reaction monitoring mode (SRM). The method of all serial chiral analytes developed was validated in rat plasma with a daily working range of 0.5–100ng/ml with correlation coefficient, R 2 ≥0.99 and a sensitivity of 0.5ng/ml as lower limit of quantification, respectively. This method was fully validated for the accuracy, precision and stability studies for all serial chiral analytes. The method proved to be accurate and specific, and was applied to the pharmacokinetic study of serial chiral novel anticholinergic compounds of phencynonate in rat plasma. [Copyright &y& Elsevier]

Abstract: A sensitive and specific gas chromatographic–mass spectrometry with the extracted ion chromatograms (GC–MS/EIC) method has been developed and validated for the identification and quantification of penehyclidine (PH) in human and animal blood. The chromatography was on HP-5 capillary column (12m×0.2mm×0.3μm). PH-d5 was selected as the internal standard (IS). Simultaneous MS detection of PH and IS was performed at m/z 315 (PH) and m/z 320 (PH-d5), and the EIC of the two compounds was at 175 and 180. PH and PH-d5 eluted at approximately 8.2min and no endogenous materials interfered with their measurement. Linearity was obtained over the concentration range of 0.1–100ng/ml in the blood. The lower limit of quantification (LLOQ) was reproducible at 50pg/ml in both human and animal blood. The within-day and between-day precisions were no more than 9.1%. This method was successfully applied to quantification and pharmacokinetic studies of PH in the subjects. The concentration–time profiles of PH in humans, rabbits and mice were all fitted to first order absorption two-compartment open model after i.m. a single dose. The differences in absorption, distribution and elimination of PH among the species were found. The results provided the important information for developing a novel anti-cholinergic drug and for obtaining a more effectual remedy in clinical practice. [Copyright &y& Elsevier]

Highlights: [•] The LLOQ of this method is better than previous published methods. [•] The esterification derivatization was used to increase the HPLC retention time and ionization efficiency in this article. [•] Peramivir is a novel anti-influenza drug which was shown to be safe and have well activity. [Copyright &y& Elsevier]

Abstract: Peramivir is a novel influenza neuraminidase inhibitor. In this article, hydrophilic interaction chromatography coupled with tandem mass spectrometry was developed to determine peramivir in human plasma. The positive ion MRM mode was performed and the precursor to the product ion transitions of m/z 329→100 and 285→138 were used to measure peramivir and Ro 64-0802 (I.S.). Chromatographic separation was performed on an Amide-80 column with acetonitrile–water–formic acid (70:30:0.1, v/v/v, 0.5mL/min). The method was linear over a concentration range of 10–10,000ng/mL. The average inter-day/intra-day precision values were 3.7±1.8% and 4.3±1.8%, respectively, while the accuracy values were 97.0±4.8%. This method has been successfully applied to Phase I of clinical research of peramivir. [Copyright &y& Elsevier]

Abstract: A sensitive and reproducible high-performance liquid chromatography (HPLC)–UV method for the determination of Z24, a tumorigenesis and angiogenesis inhibitor, has been developed and validated in mouse whole blood. Blood samples were extracted with ether, evaporated, and the residue was reconstituted in mobile phase. An aliquot was separated by isocratic reversed-phase HPLC on a Hypersil ODS-2 column and quantified using UV detection at 390nm. The mobile phase was 50% (v/v) acetonitrile/water with a flow rate of 0.8ml/min. A linear curve over the concentration range of 0.05–6μg/ml (r 2 =0.9976) was obtained. The coefficient of the variation for the intra- and inter-day precision ranged from 3.0 to 10.9% and 5.7 to 10.3%, respectively. The absolute recovery of Z24 was 89.2–108.5%. The method is simple, economical and sufficient for in vivo pharmacokinetic studies on Z24. Nonlinear pharmacokinetics was found in mice at doses from 20 to 80mg/kg. [Copyright &y& Elsevier]

Abstract: The structural elucidation of metabolites of penehyclidine in rats, a novel anti-cholinergic drug, by the method of liquid chromatography–electrospray ionization mass spectrometry, gas chromatography–mass spectrometry with electron impact ion source and stable isotope ion cluster was described. Identification and elucidation of the phase I and phase II metabolites were performed by comparing the daughter ion pairs of stable isotope cluster, changes of the protonated molecular masses, full scan MS n spectra and retention times with those of the parent drug, penehyclidine and penehyclidine deuterium-labeled. Penehyclidine was easily biotransformed by the pathways of oxidative, hydroxylated, methoxylated and phase II conjugated reactions to form several metabolites that retained the some features of the parent molecules. Twelve metabolites (penehyclidine monoxide, hydroxypenehyclidine, penehyclidine dioxide, hydroxypenehyclidine monoxide, dihydroxypenehyclidine, dihydroxypenehyclidine (position isomer), dihydroxypenehyclidine monoxide, trihydroxypenehyclidine, methoxypenehyclidine dioxide, dimethoxypenehyclidine, trihydroxymethoxypenehyclidine and glucuronide conjugated hydroxypenehyclidine) were identified. The results from electrospray ion and electron impact ion data with the stable isotope cluster showed the qualitative differences in the mass spectral patterns, suggesting that these technologies should be used in parallel to ensure comprehensive metabolites detection and characterization. The described method has wide applicability to rapidly screen and provide structural information of metabolites. [Copyright &y& Elsevier]

Abstract: A simple, sensitive and reliable method was developed to determine simultaneously the concentrations of thienorphine and its metabolite thienorphine glucuronide conjugate in rat plasma by liquid chromatography–tandem mass spectrometry (LC–MS/MS). The metabolite was identified by MS: thienorphine glucuronide conjugate. Sample preparation involved protein precipitation with methanol. Analytes were separated on Finnigan BetaBasic-18 column (150mm×2.1mm i.d., 5μm) using methanol: water: formic acid (56:44:0.1, v/v/v) as mobile phase at a flow rate of 0.2ml/min. The method had a linear calibration curve over the concentration range of 0.1–50ng/ml for thienorphine and 2–1000ng/ml for thienorphine glucuronide conjugate, respectively. LOQ of thienorphine and thienorphine glucuronide conjugate was 0.1 and 2ng/ml, respectively. The intra- and inter-batch precisions were less than 12% and their recoveries were greater than 80%. Pharmacokinetic data of thienorphine and its metabolite thienorphine glucuronide conjugate obtained with this method following a single oral dose of 3mg/kg thienorphine to rats were also reported for the first time. [Copyright &y& Elsevier]

Abstract: A sensitive and specific high-performance liquid chromatographic assay with electrospray ionization mass spectrometry detection (LC-ESI-MS) has been developed and validated for the identification and quantification of the novel anticholinergic drug phencynonate in rat blood and urine. The sample pretreatment involves basification and iterative liquid–liquid extraction with ethyl ether–dichloromethane (2:1, v/v) solution, followed by LC separation and positive electrospray ionization mass spectrometry detection. The chromatography was on BetaBasic-18 column (150mm×2.1mm i.d., 3μm). The mobile phase was composed of methanol–water (85:15, v/v), containing 0.5‰ formic acid, which was pumped at a flow-rate of 0.2ml/min. Thiencynonate was selected as the internal standard (IS). Simultaneous MS detection of phencynonate and IS was performed at m/z 358.4 (phencynonate), m/z 364 (thiencynonate), and the selected reaction ion monitoring (SRM) of the two compounds was at 156. Phencynonate eluted at approximately 5.25min, thiencynonate eluted at approximately 5.10min and no endogenous materials interfered with their measurement. Linearity was obtained over the concentration range of 1–100ng/ml in rat blood and 1–500ng/ml in rat urine. The lower limit of quantification (LLOQ) was reproducible at 1ng/ml in both of rat blood and urine. The precision measured was obtained from 2.92 to 9.76% in rat blood and 4.17 to 9.76% in rat urine. Extraction recoveries were in the range of 69.57–79.49% in blood and 56.85–64.86% in urine. This method was successfully applied to the identification and quantification of phencynonate in pharmacokinetic studies. [Copyright &y& Elsevier]