High-Performance Liquid Chromatographic Method for Quantitative Determination of Amlodipine in Human Plasma and Pharmaceutical Dosage Form and its Application to Pharmacokinetic Studies (original) (raw)
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Journal of chromatographic science, 2009
An accurate, sensitive, and reproducible high-performance liquid chromatographic method for the quantitation of amlodipine besylate in human plasma has been developed and validated. The drug, internal standard, and major metabolite were eluted from a C 18 hypersil HyPurity column (3 µm, 3.9 mm i.d. × × 150 mm) at room temperature with a mobile phase consisting of acetonitrile-potassium dihydrogen phosphate buffer (0.05 M) and acetic acid (62:38:0.1) with the pH adjusted to 3.5 using phosphoric acid. The flow-rate was 1.8 mL/min. The limit of detection was 1.0 ng/mL, and the limit of quantification of amlodipine besylate in plasma was 10 ng/mL. The intra-and inter-day precisions showed coefficients of variation ranging from 5.98-11.4% and from 5.60-11.74%, respectively at three different levels of concentration. The averages of the absolute and relative recoveries were found to be 96.74-98.51% and 95.96-100.71%, respectively. Stability studies showed that amlodipine besylate is stable for at least 2 months in plasma after freezing at -20°C. The method was successfully applied for a pharmacokinetic study and for the determination of commercial amlodipine tablet content. . Chemical structure of amlodipine.
Il Farmaco, 2005
A rapid, simple and sensitive high-performance liquid chromatography (HPLC) method has been developed for quantification of amlodipine in plasma. The assay enables the measurement of amlodipine for therapeutic drug monitoring with a minimum detectable limit of 0.2 ng ml −1 . The method involves simple, one-step extraction procedure and analytical recovery was about 97%. The separation was performed on an analytical 125 × 4.6 mm i.d. Nucleosil C 8 column. The wavelength was set at 239 nm. The mobile phase was a mixture of 0.01 M sodium dihydrogen phosphate buffer and acetonitrile (63:37, v/v) adjusted to pH 3.5 at a flow rate of 1.5 ml min -1 . The calibration curve was linear over the concentration range 0.5-16 ng ml −1 . The coefficients of variation for inter-day and intra-day assay were found to be less than 10%.
Biomedical Chromatography, 2006
A simple, sensitive and rapid high-performance liquid chromatography/electrospray ionization tandem mass spectrometry method was developed and validated for the assay of amlodipine in human plasma. Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reverse-phase C 18 column and analyzed by MS in the multiple reaction monitoring mode using the respective [M+H] + ions, m/z 409/238 for amlodipine and m/z 409/228 for the IS. The assay exhibited a linear dynamic range of 50-10,000 pg/mL for amlodipine in human plasma. The lower limit of quantification was 50 pg/mL with a relative standard deviation of less than 8%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. The average absolute recoveries of amlodipine and the IS from spiked plasma samples were 74.7 ± 4.6 and 72.1 ± 2.0%, respectively. A run time of 1.5 min for each sample made it possible to analyze more than 400 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies. The observed maximum plasma concentration (C max) of amlodipine (2.5 mg oral dose) was 1425 pg/mL, time to observed maximum plasma concentration (T max) was 8.1 h and elimination half-life (T 1/2) was 50.
A simple and validated liquid chromatographic-mass spectrometric method (LC-MS) for amlodipine in human plasma was quantifi ed using LC-MS (ESI). Chromatography was performed on a C 18 analytical column, the mobile phase used was Acetonitrile-10mM Ammonium acetate in the ratio of 90:10%v/v and the retention times were 0.829 and 1.281 min for azithromycin (Internal standard) and amlodipine respectively. The ionization was optimized using ESI (+) and enhanced selectivity was achieved. The method is validated as per FDA guidelines. The analyte was shown to be stable over the timescale of the whole procedure. The pharmacokinetic parameters such as peak plasma concentration (C max ), Time to peak Concentration (t max ), Area under the plasma concentration-time curve (AUC 0-t & AUC 0-∞ ), elimination rate constant (K eli ), Elimination half-life (t ½ ) were calculated. Log transferred values were compared by Analysis of Variance (ANOVA) followed by classical 90% confi dence interval for C max AUC 0-t .and AUC 0-∞ and was found to be within the range. These results indicated that the Test and Reference formulation is bioequivalent.
Pharmaceutical Sciences Asia, 2017
A sensitive liquid chromatography tandem mass spectrometry method was developed to quantify amlodipine in human plasma. Alkalinized plasma spiked with desipramine, an internal standard, was extracted by liquid-liquid extraction and evaporated an organic part to dryness. The residue was reconstituted and injected into an Acquity Ultra Performance LC TM , (Waters, Co., Ltd. USA) with C 18 column. The isocratic elution of mobile phase was performed by 90% of acetonitrile and 10% of 10 mM ammonium acetate pH 4 at flow rate of 0.20 mL/min with 5 minutes of total run time. Mass spectrometric analysis was performed using a Quattro Premier XE mass spectrometer, (Micromass Technologies, UK) coupled with an electrospray ionization (ESI) source in the positive ion mode. The MRM transitions of m/z 409.17>238.19 and 409.17>294.09 were selected for amlodipine and 267.09>208.06 for desipramine. The retention times were 1.63 and 1.69 minutes for amlodipine and desipramine, respectively. The linearity of the method revealed a correlation coefficient of >0.998 within the concentration range of 0.05-20 ng/mL. This work has been fully validated according to the Guidance for Industry: Bioanalytical Method Validation (USFDA CDER, 2001, BP) with high degree of accuracy and precision. This method was applied to quantify amlodipine concentrations in human plasma samples in a bioequivalence study.
Amlodipine is a dihydropyridine calcium antagonist that inhibits the movement of calcium ions into vascular smooth muscle cells and cardiac muscle cells. The aim of this work to developed simple analytical method to determine amlodipine in rat plasma using High Performance Liquid Chromatography with Ultraviolet Detector (HPLC-UV). HPLC columns Phenomenex ® C18 (Phenomenex, CA, USA), 250 mm × 4.6 mm, 5 μm particle size and guard column (C18, 4.0 X 2.0mm, Shimadazu, Japan) were used for analyzing blood samples. The compounds were separated isocratically with a mobile phase consisting of acetonetrile-phosphate buffer (0.05 M) with pH 2.8 ± 0.2 in the proportion of (40/60, v/v) at a flow rate 1.0 mL/min with injection volume 20 μL. The interday/Intraday precisions were expressed as CV% and were below 15% and the accuracy was between 80 % and 120 %, which complies with the FDA regulations [12]. The extraction procedure showed good sensitivity, specificity, precision, accuracy, recovery, and linearity, and hence the method was successfully implemented for the analysis of blood samples. In conclusion, the proposed extraction procedure showed good sensitivity, specificity, precision, accuracy, recovery, and linearity, and hence the method can be implemented for the analysis of blood samples for Pharmacokinetics or Bioequivalence study.
QUANTITATION OF AMLODIPINE IN HUMAN PLASMA BY LCMS/MS ASSAY
International Journal of Pharmacy and Pharmaceutical Sciences, 2016
Objective: To develop and validate a simple, precise, and rapid liquid chromatographic-tandem mass spectrometric (LC-MS/MS) method for quantification of amlodipine in human plasma. Methods: Chromatographic analysis was performed on Atlantis dC18 column (2.1 x 100 mm, 3 µm) with a mobile phase consisting of acetonitrile and 10 mM formic acid (80:20, v: v) that was delivered at a flow rate of 0.3 ml/min. The eluents were monitored using electrospray ionization in the positive ion mode set at transition 409 → 238.4 and 254.3 → 43.9 for amlodipine and tizanidine hydrochloride (IS), respectively. The method was validated for linearity, accuracy, precision, and recovery as per US-FDA guidelines. Results: The retention times of amlodipine and tizanidine (IS) were 1.26 and 1.22 respectively. The relationship between amlodipine concentration and peak height ratio of amlodipine to the IS was linear (R 2 Conclusion: The proposed method is simple, precise, and accurate for rapid measurement of amlodipine level using 0.5 ml human plasma. Further, the assay was successfully applied to determine amlodipine level in human plasma samples obtained from a healthy volunteer. ≥ 0.9868) in the range of 0.2-20 ng/ml, and the intra-and inter-day coefficient of variations and bias were ≤14.4% and ≤13.6% and ≤13.7% and ≤11.2%, respectively.
Analytical method development and validation of Amlodipine besylate in tablet dosage form
Journal of Drug Delivery and Therapeutics
The accurate and precise HPLC analytical method validated for the determination of Amlodipine besylate in pharmaceutical dosage form.The chromatographic separation is carried out on shimadzu HPLC system (LC-2010 CHT) with UV Vissible detector and C18(150mm x3.9 mm) 5 μm Column. The Mobile phase consists of Acetonitrile: Methanol: PH 3.0 Buffer (15 V: 35 V: 50 V) , at the flow rate of 1.0 ml/min and elutes were monitoring at 237 nm. The observed retention time for Amlodipine besylate was 10.8 min. The % RSD for system precision was 0.41 % and Method precision was 0.58 %. The method was found to linear (R=0.99996) in the Concentration range of 35-105 μg/ml (50 to 150%). The accuracy was in between 99.50-99.91%. Keywords: HPLC, Correlation coefficient, System suitability, Bias, % RSD and ICH guidelines
Biopharmaceutics & Drug Disposition, 2001
Objective—To assess the bioequivalence of two amlodipine tablet formulations (Amlodipine® 5 mg tablet from Merck S.A. Indústrias Químicas, Brazil as test formulation and Norvasc® 5 mg tablet from Laboratórios Pfizer Ltd., Brazil as reference formulation) in 24 healthy volunteers of both sexes.Methods—The study was conducted using an open, randomized two‐period crossover design with a 4‐week washout interval. Plasma samples were obtained over a 144 h period. Plasma amlodipine concentrations were analyzed by combined liquid chromatography coupled to tandem mass spectrometry (LC‐MS‐MS) with positive ion electrospray ionization using multiple reaction monitoring (MRM). From the amlodipine plasma concentration vs time curves, the following pharmacokinetic parameters were obtained: AUClast, AUC0–inf and Cmax. The statistical interval proposed was 80–125% according to the US Food and Drug Administration Agency.Results—The limit of quantification was 0.1 ng/ml for plasma amlodipine analysis...