Development of a joint population pharmacokinetic model of ezetimibe and its conjugated metabolite.

Ezetimibe (EZE) is an extensively used antihyperlipidemic drug with an important cholesterol lowering activity. It undergoes extensive first-pass metabolism to form its active glucuronide metabolite (EZEG). Both drugs exhibit complex pharmacokinetic profiles attributed mainly to repetitive enterohepatic kinetics. The aim of the present study was the investigation of EZE and EZEG pharmacokinetics (PK), through the development of a joint population pharmacokinetic model able to characterize their kinetic processes and enterohepatic recirculation simultaneously. Concentration-time data derived from a bioequivalence study in 28 healthy subjects were used for the analysis. Population PK modeling was performed on the obtained data using nonlinear mixed effect modeling approach, where different methodologies were applied for the description of the complex metabolism and recirculation processes of the two compounds. EZE and EZEG concentrations were best described by a population PK model incorporating first-pass metabolism and an enterohepatic recirculation loop, accounting for the recycling process of the two moieties. This is the first joint population pharmacokinetic model describing the kinetics of both EZE and EZEG.

Cetyl-alcohol-reinforced hollow fiber solid/liquid-phase microextraction and HPLC-DAD analysis of ezetimibe and simvastatin in human plasma and urine.

A new cetyl-alcohol-reinforced hollow fiber solid/liquid-phase microextraction (CA-HF-SLPME) followed by high-performance liquid chromatography-diode array detection (HPLC-DAD) method was developed for simultaneous determination of ezetimibe and simvastatin in human plasma and urine samples. To prepare the CA-HF-SLPME device, the cetyl-alcohol was immobilized into the pores of a 2.5 cm hollow fiber micro-tube and the lumen of the micro-tube was filled with 1-octanol with the two ends sealed. Afterwards, the prepared device was introduced into 10 mL of the sample solution containing the analytes with agitation. Under optimized conditions, calibration curves plotted in spiked plasma and urine samples were linear in the ranges of 0.363-25/0.49-25 µg L-1 for ezetimibe/simvastatin and 0.193-25/0.312-25 µg L-1 for ezetimibe/simvastatin in plasma and urine samples, respectively. The limit of detection was 0.109/0.174 µg L-1 for ezetimibe/simvastatin in plasma and 0.058/0.093 µg L-1 for ezetimibe/simvastatin in urine. As a potential application, the proposed method was applied to determine the concentration of selected analytes in patient plasma and urine samples after medication and satisfactory results were achieved. In comparison with reference methods, the CA-HF-SLPME-HPLC-DAD method demonstrates considerable potential in the biopharmaceutical analysis of selected drugs.

Simultaneous quantitation of rosuvastatin and ezetimibe in human plasma by LC-MS/MS: Pharmacokinetic study of fixed-dose formulation and separate tablets.

A simple, high-throughput and highly sensitive liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method has been developed for the simultaneous estimation of rosuvastatin and free ezetimibe. Liquid-liquid extraction was carried out using methyl-tert butyl ether after prior acidification from 300 µL human plasma. The recovery for both the analytes and their deuterated internal standards (ISs) ranged from 95.7 to 99.8%. Rosuvastatin and ezetimibe were separated on Symmetry C18 column using acetonitrile and ammonium formate buffer, pH 3.5 (30:70, v/v) as the mobile phase. The analytes were well resolved with a resolution factor of 3.8. Detection and quantitation were performed under multiple reaction monitoring using ESI(+) for rosuvastatin (m/z 482.0 → 258.1) and ESI(-) for ezetimibe (m/z 407.9 → 271.1). A linear response function was established in the concentration ranges of 0.05-50.0 ng/mL and 0.01-10.0 ng/mL for rosuvastatin and ezetimibe, respectively, with correlation coefficient, r2 ≥ 0.9991. The IS-normalized matrix factors for the analytes ranged from 0.963 to 1.023. The developed method was successfully used to compare the pharmacokinetics of a fixed-dose combination tablet of rosuvastatin-ezetimibe and co-administered rosuvastatin and ezetimibe as separate tablets to 24 healthy subjects. The reliability of the assay was also assessed by reanalysis of 115 subject samples.

A rapid and sensitive supercritical fluid chromatography/tandem mass spectrometry method for detection of ezetimibein dog plasma and its application in pharmacokinetic studies.

The aim of this study is to develop and validate a rapid, high-selective and sensitive supercritical fluid chromatography/tandem mass spectrometry (SFC-MS/MS) with a multiple reactions monitoring (MRM) mode method for the detection of ezetimibe in dog plasma. Several conditions were optimized systematically as follows: lipid-lipid extraction (LLE) performances were used to extract analytes from dog plasma; an ACQUITY HSS C18 SB (1.8 µm, 3.0 × 100 mm) column was employed to separate the target compounds; the triple-quadrupole mass spectrometry equipped with electrospray ionization (ESI) source was applied to detect ezetimibe. The method, which required a relatively small volume of plasma (100 µL), was obtained at concentration ranging from 1.0 to 100 ng/mL(r2 > 0.99). The lower limit of quantification (LLOQ)for ezetimibe was found to be as low as 1.0 ng/mL. In addition, the validations of the methodology including sensitivity, recovery, matrix effect, intra- and inter-day precision, accuracy and stability were all within acceptable limits. The Cmax, AUC0-inf and Tmax values obtained in our study were 52.2 ±â€¯6.3, 820.6 ±â€¯4.3 and 1.25 ±â€¯0.35 for reference formulation; 61.8 ±â€¯12.6, 924.2 ±â€¯4.7 and 2.00 ±â€¯0 for test formulation. In conclusion, the method developed in this study can be successfully applied to pharmacokinetic studies after oral administration of ezetimibe in dogs.

Development and Validation of an LC-MS-MS Method for the Simultaneous Determination of Simvastatin, Simvastatin Acid and Ezetimibe in Human Plasma and Its Application to Pharmacokinetic Study in the Indian Population.

A simple, selective, sensitive and high-throughput liquid chromatography-tandem mass spectrometry (LC-MS-MS) method has been developed and validated for the simultaneous quantification of simvastatin (SS), simvastatin acid (SSA, active metabolite of SS) and ezetimibe (EZM) in K2 EDTA containing human plasma, using simvastatin D6, simvastatin acid D3 and ezetimibe D4 as internal standards (ISTDs), respectively. A volume of plasma sample of only 400 µL was processed by the solid phase extraction technique; then 20 µL of processed sample was run on a Phenomenex, Kinetix XB C18, 150 × 4.6 mm, 5 µm column using an isocratic mobile phase consisting of 10 mM ammonium formate buffer (pH 4.0 ± 0.3): acetonitrile (27 : 73, v/v) with a run time of 6.3 min. The precursor and product ions of SSA, EZM and their ISTDs were monitored on a triple quadrupole instrument operated in the negative ionization mode, and SS was monitored in the positive mode. The method was validated over a concentration range of 0.2-80 ng/mL for SS, 0.1-60 ng/mL for SSA and 0.05-15 ng/mL for EZM. The method has been successfully applied in clinical pharmacokinetic study in the Indian population. The Cmax, AUC0-inf and Tmax values obtained in our study were 10.61 ± 5.287, 77.58 ± 29.367 and 1.62 ± 0.436 for EZM; 69.74 ± 45.274, 190.71 ± 107.271 and 1.74 ± 0.480 for SS; and 25.36 ± 23.576, 139.24 ± 131.653 and 3.95 ± 0.671 for SSA, respectively.

Simultaneous determination of ezetimibe and its glucuronide metabolite in human plasma by solid phase extraction and liquid chromatography-tandem mass spectrometry.

A liquid chromatography-tandem mass spectrometry method (LC-MS/MS) was developed to quantify ezetimibe (EZM) and its major glucuronide (ezetimibe glucuronide, EZM-G) in human plasma simultaneously. The analytes were purified by solid phase extraction (SPE) without hydrolysis. Separation of the analytes was achieved using acetonitrile-water (0.08% formic acid) (70:30, v/v) as the mobile phase at a flow rate of 0.8 mL/min on an Agilent Extend C18 column. The analytes were detected by LC-MS/MS using negative ionization in multiple reaction monitoring (MRM) mode. The mass transition pairs of m/z 408.4→271.0 and m/z 584.5→271.0 were used to detect EZM and EZM-G, respectively. The analytical method was linear over the concentration range of 0.1-20 ng/mL for EZM and 0.5-200 ng/mL for EZM-G. Within- and between-run precision for EZM was no more than 8.6% and 12.8%; and for EZM-G was no more than 9.0% and 8.7%, respectively. This method was reproducible and reliable, and was successfully used to analyze human plasma samples for application in a bioequivalence study.