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Evolocumab Effects on Lipoproteins, Measured by High-Performance Liquid Chromatography.
Masuda, D, Kiyosue, A, Hirayama, A, Shimauchi, J, López, JAG, Miyawaki, K, Yamashita, S
Journal of atherosclerosis and thrombosis. 2020;(11):1183-1207
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Abstract
AIMS: Profiling of lipoproteins can predict risk of cardiovascular disease; gel permeation high-performance liquid chromatography (HPLC) improves prediction accuracy by providing detailed data for specific lipoprotein subclasses. This study applied HPLC to examine the effects of evolocumab, which effectively treats hyperlipidemia and mixed dyslipidemia, on lipoprotein subclasses, specifically the number and size of lipoprotein particles. METHODS This post-hoc analysis used patient blood samples from YUKAWA-2, a phase 3 trial evaluating the efficacy of evolocumab in Japanese adult patients with hyperlipidemia or mixed dyslipidemia and at high risk for cardiovascular disease. We used HPLC to assess observed values and percent change from baseline in cholesterol and triglyceride (TG) concentrations, number of particles in lipoprotein subclasses to week 12, and mean observed values and mean percent change from baseline in variables to weeks 10 and 12. HPLC was also compared with conventional methods in assessing low-density lipoprotein (LDL) cholesterol (LDL-C) values. RESULTS Data for all 404 patients were analyzed. Evolocumab significantly decreased cholesterol and TG concentrations, and total particle count, in very low-density lipoprotein (VLDL) and LDL subclasses. Particle size increased slightly in LDL, high-density lipoprotein (HDL), and VLDL, but data varied widely. At very low LDL-C, HPLC measurements were higher than those from conventional methods. CONCLUSION This research used HPLC to assess the effects of evolocumab in 20 lipid subclasses. By lowering lipid content and improving the lipid profile, evolocumab may reduce atherogenicity. This reduction is better quantified by HPLC than by conventional methods in the very low LDL-C range.
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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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HPLC with fluorescence detection assay of perampanel, a novel AMPA receptor antagonist, in human plasma for clinical pharmacokinetic studies.
Mano, Y, Takenaka, O, Kusano, K
Biomedical chromatography : BMC. 2015;(10):1589-93
Abstract
Perampanel (Fycompa®), a novel α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist, is registered for the adjunctive treatment of patients (aged ≥12 years) with refractory partial-onset seizures. To support therapeutic drug monitoring, a simple high-performance liquid chromatography (HPLC) assay with fluorescence detection was developed to determine perampanel concentrations in human plasma and validated to support clinical trials. Human plasma samples (1.0 mL) were processed by liquid extraction using diethyl ether, followed by chromatographic separation on a YMC Pack Pro C18 column (150 × 4.6 mm i.d., 5 µm) with isocratic elution of acetonitrile-water-acetic acid-sodium acetate (840:560:3:1.8, v/v/v/w) at a flow rate of 1.0 mL/min. Column eluent was monitored at excitation and emission wavelengths of 290 and 430 nm, respectively. The assay was linear (range 1.0-500 ng/mL) and this could be extended to 25 µg/mL by 50-fold dilution integrity. No endogenous peaks were detected in the elution of analytes in drug-free blank human plasma from six individuals and no interference was observed with co-medications tested. Intra- and inter-batch reproducibility studies demonstrated accuracy and precision within the acceptance criteria of bioanalytical guidelines. Validation data demonstrated that our assay is simple, selective, reproducible and suitable for therapeutic drug monitoring of perampanel.
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Validation and pharmacokinetic application of a high-performance liquid chromatographic technique for determining the concentrations of amodiaquine and its metabolite in plasma of patients treated with oral fixed-dose amodiaquine-artesunate combination in areas of malaria endemicity.
Adedeji, ON, Bolaji, OO, Falade, CO, Osonuga, OA, Ademowo, OG
Antimicrobial agents and chemotherapy. 2015;(9):5114-22
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Abstract
Artemisinin-based combination therapies (ACTs) have been adopted by most African countries, including Nigeria, as first-line treatments for uncomplicated falciparum malaria. Fixed-dose combinations of these ACTs, amodiaquine-artesunate (FDC AQAS) and artemether-lumefantrine (AL), were introduced in Nigeria to improve compliance and achieve positive outcomes of malaria treatment. In order to achieve clinical success with AQAS, we developed and validated a simple and sensitive high-performance liquid chromatography (HPLC) method with UV detection for determination of amodiaquine (AQ) and desethylamodiaquine (DAQ) in plasma using liquid-liquid extraction of the drugs with diethyl ether following protein precipitation with acetonitrile. Chromatographic separation was achieved using an Agilent Zorbax C18 column and a mobile phase consisting of distilled water-methanol (80:20 [vol/vol]) with 2% (vol/vol) triethylamine, pH 2.2, at a flow rate of 1 ml/min. Calibration curves in spiked plasma were linear from 100 to 1,000 ng/ml (r > 0.99) for both AQ and DAQ. The limit of detection was 1 ng (sample size, 20 μl). The intra- and interday coefficients of variation at 150, 300, and 900 ng/ml ranged from 1.3 to 4.8%, and the biases were between 6.4 and 9.5%. The mean extraction recoveries of AQ and DAQ were 80.0% and 68.9%, respectively. The results for the pharmacokinetic parameters of DAQ following oral administration of FDC AQAS (612/200 mg) for 3 days in female and male patients with uncomplicated falciparum malaria showed that the maximum plasma concentrations (C max) (740 ± 197 versus 767 ± 185 ng/ml), areas under the plasma concentration-time curve (AUC) (185,080 ± 20,813 versus 184,940 ± 16,370 h · ng/ml), and elimination half-life values (T 1/2) (212 ± 1.14 versus 214 ± 0.84 h) were similar (P > 0.05).
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Validated HILIC-MS/MS assay for determination of vindesine in human plasma: Application to a population pharmacokinetic study.
Zhu, RH, Li, HD, Cai, HL, Jiang, ZP, Xu, P, Dai, LB, Peng, WX
Journal of pharmaceutical and biomedical analysis. 2014;:31-6
Abstract
The first HILIC-tandem mass spectrometry (MS/MS) method for determination of vindesine (VDS) in human plasma using vinorelbine as an internal standard (IS) has been developed and validated. Plasma samples clean-up consisted of solid phase extraction with a strata™-X column. The compounds were separated on a HILIC column with an isocratic mobile phase consisting of acetonitrile and 15mM ammonium acetate buffer containing 0.15% formic acid (80:20, v/v). The detection was performed on a triple quadrupole tandem mass spectrometer via electrospray positive ionization (ESI(+)). The ion transitions recorded in multiple reaction monitoring mode were m/z 754.6→123.8 for VDS and 779.4→323.3 for IS, respectively. Linear calibration curves were obtained in the concentration range of 0.3-28ng/ml and the lower limit of quantification for VDS was 0.3ng/ml. The coefficient of variation of the assay precision was less than 13%, and the accuracy exceeded 96%. The developed assay method was successfully applied for the evaluation of population pharmacokinetics of VDS after intravenous infusion of Xi Ai Ke Vial(®) (3mg of Vindesine Sulfate for Injection) to Chinese Han subjects with hematological malignant disorders.
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Metabolic profile of keratoconic cornea.
Kryczka, T, Ehlers, N, Nielsen, K, Wylegala, E, Dobrowolski, D, Midelfart, A
Current eye research. 2013;(2):305-9
Abstract
PURPOSE To investigate the difference in metabolic profile of keratoconic and normal corneas using two different analysis methods. METHODS Keratoconic corneas were obtained from patients (aged 19-27) during transplantation surgery. Control samples were obtained from human donors (aged 61-75) 1-8 h post-mortem. The metabolic profile of tissues was investigated with high-resolution magic angle spinning (1)H nuclear magnetic resonance (NMR) spectroscopy and high performance liquid chromatography (HPLC). RESULTS Nine amino acids and 20 metabolites were assigned with HPLC and NMR spectroscopy, respectively. No significant biochemical difference was revealed between keratoconic and control samples, which represent distant age groups. CONCLUSIONS It suggests that development of keratoconus might be related to the accelerated ageing of the cornea. This issue warrants further studies.
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Determination of piperphentonamine and metabolites M1 and M6 in human plasma and urine by LC/MS/MS and its application in a pharmacokinetics study in Chinese healthy volunteers.
Shi, A, Hu, X, Li, K, Li, J, Han, L, Wan, H, Li, R
Journal of chromatography. B, Analytical technologies in the biomedical and life sciences. 2012;:87-93
Abstract
Piperphentonamine hydrochloride (PPTA) is a new calcium sensitizer. A liquid chromatography-tandem mass spectrometry (LC/MS/MS) method for determination of piperphentonamine and its metabolites M1 and M6 was developed for the first time and applied to a pharmacokinetics study. Protein precipitation was used for pre-treatment of plasma samples, and solid phase extraction method was used for pre-treatment of urine samples. The chromatographic separation was achieved on a C(18) column using gradient elution in this study: A: 1% acetic acid aqueous solution, and B: acetonitrile. The whole analysis lasted for 10.5min and the gradient flow rate was 0.25mL/min constantly. The detection was performed of a triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode via a positive electrospray ionization source. The results were that the m/z ratios of monitored precursor ions and product ions of PPTA, M1 and M6 were 354.0→191.8, 356.0→148.7 and 358.0→148.7, respectively. From the standard curve, the concentration ranges of both PPTA and M1 in blood and urine samples were 0.1-500ng/mL and 0.1-200ng/mL, respectively; the concentration ranges of M6 in blood sample and urine sample were 0.2-500ng/mL and 0.2-200ng/mL, respectively; and the correlation coefficient of standard curve was r>0.99. A total of 31 healthy Chinese subjects participated in the pharmacokinetic study of single bolus intravenous injection of piperphentonamine hydrochloride. They were divided into three dosage groups and given 0.2, 0.4 and 0.6mg/kg of PPTA. After drug administration, concentrations of PPTA, M1 and M6 in human plasma and urine samples were determined to evaluation the pharmacokinetic characteristics of PPTA and its metabolites M1 and M6.
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A validated HPLC assay for the determination of R-(-)-gossypol in human plasma and its application in clinical pharmacokinetic studies.
Lin, H, Gounder, MK, Bertino, JR, Kong, AN, DiPaola, RS, Stein, MN
Journal of pharmaceutical and biomedical analysis. 2012;:371-5
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Abstract
R-(-)-gossypol acetic acid (AT-101), a natural BH3 mimetic, is investigated in a Phase I/II clinical trial for the treatment of advanced solid tumor malignancies. Gossypol undergoes rapid degradation in solution phase, which causes major technical difficulty for its quantitation in plasma. We developed and validated a sensitive HPLC assay for pharmacokinetic evaluation of gossypol. Acetonitrile deproteinization method was chosen for sample preparation and Schiff's base derivative, R-(-)-gossypol-diamino-propanol (GDP), was used as internal standard. Chromatographic separation of gossypol in plasma was performed using a Zorbax Eclipse XDB column C(18) at 30 °C. The mobile phase consists of 10 mmol/L KH(2)PO(4) (pH 3.0) and acetonitrile (20:80) at 1.0 mL/min flow rate. Linearity ranged over 56-3585 ng/mL (R(2)=0.9997±0.0003, n=4), and the limit of detection was 28 ng/mL. The intra- and inter-assay precision was less than 13.7% and the bias ranged from -7.4 to 7.0%. The method was successfully applied to characterize the pharmacokinetics of AT-101 in a Phase I clinical trial. The validated assay is accurate, and sensitive with minimum loss and rapid analysis time and suitable for quantification of gossypol for pharmacokinetics evaluation.
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Population pharmacokinetic model of canrenone after intravenous administration of potassium canrenoate to paediatric patients.
Suyagh, M, Hawwa, AF, Collier, PS, Millership, JS, Kole, P, Millar, M, Shields, MD, Halliday, HL, McElnay, JC
British journal of clinical pharmacology. 2012;(5):864-72
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Abstract
WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT Little is known about the pharmacokinetics of potassium canrenoate/canrenone in paediatric patients WHAT THIS STUDY ADDS A population pharmacokinetic model has been developed to evaluate the pharmacokinetics of canrenone in paediatric patients who received potassium canrenoate as part of their therapy in the intensive care unit. AIMS To characterize the population pharmacokinetics of canrenone following administration of potassium canrenoate to paediatric patients. METHODS Data were collected prospectively from 23 paediatric patients (2 days to 10 years of age; median weight 4 kg, range 2.16-28.0 kg) who received intravenous potassium canrenoate (K-canrenoate) as part of their intensive care therapy for removal of retained fluids, e.g. in pulmonary oedema due to chronic lung disease and for the management of congestive heart failure. Plasma samples were analyzed by HPLC for determination of canrenone (the major metabolite and pharmacologically active moiety) and the data subjected to pharmacokinetic analysis using NONMEM. RESULTS A one compartment model best described the data. The only significant covariate was weight (WT). The final population models for canrenone clearance (CL/F) and volume of distribution (V/F) were CL/F (l h(-1) ) = 11.4 × (WT/70.0)(0.75) and V/F (l) = 374.2 × (WT/70) where WT is in kg. The values of CL/F and V/F in a 4 kg child would be 1.33 l h(-1) and 21.4 l, respectively, resulting in an elimination half-life of 11.2 h. CONCLUSIONS The range of estimated CL/F in the study population was 0.67-7.38 l h(-1) . The data suggest that adjustment of K-canrenoate dosage according to body weight is appropriate in paediatric patients.
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Caffeine and paraxanthine HPLC assay for CYP1A2 phenotype assessment using saliva and plasma.
Perera, V, Gross, AS, McLachlan, AJ
Biomedical chromatography : BMC. 2010;(10):1136-44
Abstract
Caffeine has been extensively used as a probe to measure CYP1A2 activity in humans with caffeine clearance or the paraxanthine (major metabolite of caffeine) to caffeine concentration ratio being regarded as the preferred metric. A simple reverse-phased C(18) HPLC assay using ethyl acetate liquid-liquid extraction was developed to quantitate caffeine and paraxanthine concentrations in saliva and plasma. The mobile phase consisted of acetonitrile-acetic acid-H(2)O (100:1:899) and analytes were quantitated with UV detection at 280 nm. The extraction recovery for paraxanthine and caffeine was approximately 70% in both saliva and plasma. The assay was linear over the concentration ranges 0.05-2.50 and 0.05-5.00 µg/mL, for paraxanthine and caffeine, respectively, in saliva. In plasma the assay was linear over the ranges 0.025-2.50 and 0.025-5.00 µg/mL for paraxanthine and caffeine, respectively. Intra- and inter-assay precision and accuracy were less than 15%. Detection limits were 0.015 µg/mL for paraxanthine and caffeine in saliva, while it was 0.005 µg/mL for paraxanthine and caffeine in plasma. Utility was established in samples collected from two healthy volunteers who abstained from caffeine for 24 h and received a single 100 mg oral dose of caffeine. The assay developed is a robust, simple and precise technique to measure caffeine and paraxanthine in saliva and plasma of healthy volunteers after a single oral dose of caffeine.