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Development and validation of samples stabilization strategy and LC-MS/MS method for simultaneous determination of clevidipine and its primary metabolite in human plasma: Application to clinical pharmacokinetic study in Chinese healthy volunteers.
Zhang, Y, Zhao, S, Zhou, H, Ding, L
Journal of chromatography. B, Analytical technologies in the biomedical and life sciences. 2020;:122448
Abstract
A feasible LC-MS/MS method with reliable stabilizers consisted of sodium fluoride, ascorbic acid and formic acid was developed and validated for the determination of clevidipine and its primary metabolite (H152/81) in human plasma. Sodium fluoride existing in the vacutainer tubes was used to inhibit esterase activity to protect the clevidipine from hydrolysis as soon as blood was collected. Ascorbic acid and formic acid were added to the separated plasma samples to avoid the oxidation and further hydrolysis of clevidipine and H152/81. The further sample preparation was accomplished through a single step liquid-liquid extraction (LLE) by ethyl acetate. The chromatography separation was carried out on an ACE Excel 3 μm SuperC18 (2.1 × 50 mm, id, ACE, United Kingdom) column with gradient elution using 10 mM ammonium acetate water solution and methanol as the mobile phase. Detection was performed in the negative ion electrospray ionization mode using multiple reaction monitoring (clevidipine: m/z 454.1 → 234.0; clevidipine-d7: m/z 461.1 → 240.1; H152/81: m/z 354.0 → 208.0; H152/81-13CD3: m/z 358.0 → 212.0). The method exhibited good linearity over the concentration ranges of 0.100 to 40.0 ng/mL for clevidipine and 5.00 to 400 ng/mL for H152/81. The intra- and inter-batch precision and accuracy of clevidipine and H152/81 were all within the acceptable criteria. The method was successfully applied to a pharmacokinetic study of clevidipine and H152/81 in healthy Chinese volunteers following 8 mg/h intravenous infusion of clevidipine butyrate injectable emulsion for 0.5 h. The results showed that clevidipine was rapidly eliminated with a short half-life time of 0.244 ± 0.125 h and a maximum concentration of 25.2 ± 7.09 ng/mL. H152/81 was detectable in the plasma samples up to 48.5 h with a half-life time of 10.7 ± 2.30 h and a maximum plasma concentration of 301 ± 38.1 ng/mL.
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Simultaneous quantification of alpha-aminoadipic semialdehyde, piperideine-6-carboxylate, pipecolic acid and alpha-aminoadipic acid in pyridoxine-dependent epilepsy.
Xue, J, Wang, J, Gong, P, Wu, M, Yang, W, Jiang, S, Wu, Y, Jiang, Y, Zhang, Y, Yuzyuk, T, et al
Scientific reports. 2019;(1):11371
Abstract
The measurements of lysine metabolites provide valuable information for the rapid diagnosis of pyridoxine-dependent epilepsy (PDE). Here, we aimed to develop a sensitive method to simultaneously quantify multiple lysine metabolites in PDE, including α-aminoadipic semialdehyde (a-AASA), piperideine-6-carboxylate (P6C), pipecolic acid (PA) and α-aminoadipic acid (α-AAA) in plasma, serum, dried blood spots (DBS), urine and dried urine spots (DUS). Fifteen patients with molecularly confirmed PDE were detected using liquid chromatography-mass spectrometry (LC-MS/MS) method. Compared to the control groups, the concentrations of a-AASA, P6C and the sum of a-AASA and P6C (AASA-P6C) in all types of samples from PDE patients were markedly elevated. The PA and a-AAA concentrations ranges overlapped partially between PDE patients and control groups. The concentrations of all the analytes in plasma and serum, as well as in urine and DUS were highly correlated. Our study provided more options for the diverse sample collection in the biochemical tests according to practical requirements. With treatment modality of newly triple therapy investigated, biomarker study might play important role not only on diagnosis but also on treatment monitoring and fine tuning the diet. The persistently elevated analytes with good correlation between plasma and DBS, as well as urine and DUS made neonatal screening using DBS and DUS possible.
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UHPLC-MS/MS method for determination of atorvastatin calcium in human plasma: Application to a pharmacokinetic study based on healthy volunteers with specific genotype.
Xia, B, Li, Y, Zhang, Y, Xue, M, Li, X, Xu, P, Xia, T, Chen, S
Journal of pharmaceutical and biomedical analysis. 2018;:428-435
Abstract
A rapid, selective and sensitive ultra high performance liquid chromatography coupled with tandem triple quaternary mass spectrometry (UHPLC-MS/MS) method was developed and validated for the quantitative determination of atorvastatin calcium (AC) in human plasma. Separation of AC and rosuvastatin calcium (internal standard, IS) were achieved on a Dikma Leapsil C18 reversed phase column (100 × 2.1 mm, 2.7 μm) with gradient elution using 0.2% (v/v) formic acid in water and acetonitrile as mobile phases, at the flow rate of 0.3 mL/min. AC and IS were detected using MS/MS with turbo ion pray source in negative mode by monitoring the precursor-to-product ion transitions m/z 557.0→453.0 for AC and m/z 480.0→418.0 for IS. The calibration curves were linear from 0.05 to 50 ng/mL with a correlation coefficient ( r2) of 0.9992 or better. Thereafter, 187 healthy candidates were checked to the genetic polymorphism analysis of SLCO1B1 521T>C(rs4149056), SLCO1B1 388A>G(rs2306283), CYP3A4 1*B(rs2740574), CYP3A4 1*G(rs2242480) and CYP3A5*3(rs776746) using fluorescence in situ hybridization technology. The genotype frequencies of wild-type homozygote, mutant heterozygote and mutant homozygote were 62.57%(TT), 34.22%(TC) and 3.21%(CC) for SLCO1B1 521T>C, and 8.56%(AA), 33.69%(AG) and 57.75%(GG) for SLCO1B1 388A>G, and 62.57%(CC), 34.22%(CT) and 3.21%(TT) for CYP3A4 1 G, and 58.29%(GG), 34.76%(GA) and 6.95%(AA) for CYP3A5*3, respectively. Furthermore, each tested genotype of CYP3A4 1B was wild type. Finally, 5 candidates with specific genotype described above were recruited to carry out the clinical pharmacokinetics of AC (n = 5). The validated UHPLC-MS/MS method was implemented in a high-throughput setting, capable of analyzing up to 288 samples per day, and was successfully applied to the pharmacokinetic study of AC based on healthy volunteers with specific genotype. The Cmax of AC in human volunteers with the specific genotype was nearly 10 times higher than that previous reported, indicating that genetic polymorphisms of these specific genotypes have significant influence on pharmacokinetics of atorvastatin.
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Simultaneous quantification of EVT201, a novel partial positive allosteric GABAA receptor modulator, and its two metabolites in human plasma by UHPLC/MS/MS.
Zhang, X, Zhou, C, Zhang, Y, Zhu, S, Cai, X, Pan, C, Wang, C, Zhai, S, Yang, L
Journal of pharmaceutical and biomedical analysis. 2018;:282-290
Abstract
A sensitive, accurate and rapid UHPLC-MS/MS method was developed and validated for the simultaneous determination of EVT201 and its two metabolites, Ro46-1927 and Ro18-5528, in human plasma. D6-EVT201 was used as the internal standard (IS). Plasma samples were extracted using ethyl acetate after being alkalized with saturated sodium carbonate solution. Chromatographic separation was carried out on an Acquity BEH C18 column (2.1 × 50 mm, 1.7 μm) with a gradient mobile phase at a flow-rate of 0.50 mL/min. The analytical run time was 5.5 min. For mass spectrometric detection, multiple reaction monitoring (MRM) was used. The MRM ion transitions were m/z 373.2 → 58.0 for EVT201, m/z 359.2 → 316.1 for Ro46-1927, m/z 291.2 → 274.1 for Ro18-5528 and m/z 379.3 → 64.0 for the IS. The linear range was 0.2-200 ng/mL for each analyte, with a correlation coefficient (r) over 0.9900. The intra-/inter- precision was within 7.9% and 4.5% for EVT201, 13.2% and 6.3% for Ro46-1927, 3.7% and 4.1% for Ro18-5528. For the accuracy, the relative bias of intra-/inter- run was no more than 6.4% and 4.6% for EVT201, 9.4% and 7.9% for Ro46-1927, -6.9% and -7.5% for Ro18-5528. The validated method was successfully applied to the analysis of more than 1500 samples from a Phase I clinical trial. The incurred sample reanalysis (ISR) of 146 samples from the study was evaluated and the results met the acceptance criteria. It indicated that the method could be used for the pharmacokinetic study of EVT201.