Liquid Chromatography–Mass Spectrometry Method for the Determination of Amlodipine in Human Plasma and its Application in a Bioequivalence Study

Abstract

A sensitive and selective liquid chromatographic-mass spectrometric (LC-MS) method for the determination of amlodipine (CAS 88150-42-9) in human plasma has been developed. Sample preparation was based on liquid-liquid extraction using NaOH and cyclohexane. Analytical determination was carried out on a C18 column interfaced with a single quadrupole mass spectrometer. Positive electrospray ionization was employed as the ionization source. The mobile phase was 10 mmol/L ammonium acetate solution-methanol (30 : 70, v/v) at the flow rate of 0.2 mL/min. The method was linear in the concentration range of 0.2–20 ng/mL. The lower limit of quantification was 0.2 ng/mL. Amlodipine was sensitive to UV light. The method was validated in terms of accuracy, precision, absolute recovery, and stability. The intra- and inter-day relative standard deviation across three validation runs over the entire concentration range was less than 8.17%. The accuracy determined at three concentrations (0.4, 2.0 and 10 ng/mL for amlodipine maleate) was within ± 3.17% in terms of relative error. The method herein described was successfully applied for the evaluation of pharmacokinetic profiles of amlodipine maleate tablets in 20 healthy volunteers. The results showed that AUC, T_max, C_max and T_1/2 between the test and reference formulation have no significant difference (P > 0.05). The relative bioavailability was 103.7 ± 12.3%.

Key words

Amlodipine maleate, liquid chromatographic-mass spectrometric determination, pharmacokinetics - Calcium antagonists - CAS 88150-42-9

• References

• 1 Arrowsmith JE, Campbell SF, Cross PE, Stubbs JK, Burges RA, Gardiner DG, Blackburn KL. Long-acting dihydropyridine calcium antagonists. 1. 2-Alkoxymethyl derivatives incorporating basic substituents. J Med Chem. 1986; 29: 1696-702
• 2 Burges R, Moisey D. Unique pharmacologic properties of amlodipine. Am J Cardiol. 1994; 73: 2A-9A
• 3 Meredith PA, Elliott HL. Clinical pharmacokinetics of amlodipine. Clin Pharmacokinet. 1992; 22: 22-31
• 4 EMEA/CPMC opinion following an article 29 referral. International non-proprietary name: Amlodipine. London; 2004.
• 5 Park JY, Kim KA, Lee GS, Park PW, Kim SL, Lee YS et al. Randomized, openlabel, two-period crossover comparison of the pharmacokinetic and pharmacodynamic properties of two amlodipine formulations in healthy adult male Korean subjects. Clin Ther. 2004; 26: 715-23
• 6 Hong SJ, Ahn TH, Baek SH, Cho WH, Jeon HK, Kyun J et al. Comparison of efficacy and tolerability of amlodipine orotate versus amlodipine besylate in adult patients with mild to moderate hypertension: A multicenter, randomized, double-blind, placebo-controlled, parallel-group, eight-week follow-up, noninferiority trial. Clin Ther. 2006; 28: 537-55
• 7 Zhang H, Wang SM. Comparative study of amlodipine maleate and amlodipine benzene sulfonate on hypertension. Modern Med Health. 2004; 16: 1648-9
• 8 Stopher DA, Beresford AP, Macrae PV, Humphrey MJ. The metabolism and pharmacokinetics of amlodipine in human and animals. J Cardiovasc Pharmacol. 1988; 12: S55-9
• 9 Murdoch D, Heel RC. Amlodipine: a review of its pharmacodynamic and pharmacokinetic properties, and therapeutic use in cardiovascular disease. Drugs. 1991; 41: 478-505
• 10 Zarghi A, Foroutan SM, Shafaati A, Khoddam A. Validated HPLC method for determination of amlodipine in human plasma and its application to pharmacokinetic studies. Il Farmaco. 2005; 60: 789-92
• 11 Bahrami GH, Mirzaeei SH. Simple and rapid HPLC method for determination of amlodipine in human serum with fluorescence detection and its use in pharmacokinetic studies. J Pharm Biomed Anal. 2004; 36: 163-8
• 12 Josefsson M, Zackrisson AL, Norlander B. Sensitive high-performance liquid chromatographic analysis of amlodipine in human plasma with amperometric detection and a single step solid-phase sample preparation. J Chromatogr B Biomed Appl. 1995; 672: 310-3
• 13 Monkman SC, Ellis JS, Cholerton S, Thomason JM, Seymour RA, Idle JR. Automated gas chromatographic assay for amlodipine in plasma and gingival crevicular fluid. J Chromatogr B Biomed Appl. 1996; 678: 360-4
• 14 Suchanova B, Kostiainen R, Setola RA. Characterization of the in vitro metabolic profile of amlodipine in rat using liquid chromatography–mass spectrometry. Eur J Pharm Sci. 2008; 33: 91-9
• 15 Carvalho M, Oliveira CH, Mendes GD, Sucupira M, Moraes ME, De Nucci G. Amlodipine bioequivalence study: quantification by liquid chromatography coupled to tandem mass spectrometry. Biopharm Drug Dispos. 2001; 22: 383-90
• 16 Abad-Santos F, Novalbos J, Gálvez-Múgica MA, Gallego-Sandín S, Almeida S, Vallée F, García AG. Assessment of sex differences in pharmacokinetics and pharmacodynamics of amlodipine in a bioequivalence study. Pharmacol Res. 2005; 51: 445-52
• 17 Massaroti P, Moraes LAB, Marchioretto MAM, Cassiano NM, Bernasconi G, Calafatti SA et al. Development and validation of a selective and robust LC-MS/MS method for quantifying amlodipine in human plasma. Anal Bioanal Chem. 2005; 382: 1049-54
• 18 Nirogi RVS, Kandikere VN, Mudigonda K, Shukla M, Maurya S. Sensitive and rapid liquid chromatography/tandem mass spectrometry assay for the quantification of amlodipine in human plasma. Biomed. Chromatogr. 2006; 20: 833-42
• 19 Bhatt J, Singh S, Subbaiah G, Shah B, Kambli S, Ameta S. A rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the estimation of amlodipine in human plasma. Biomed Chromatogr. 2007; 21: 169-75
• 20 Yuanyuan Ma, Feng Qin, Xiaohong Sun, Xiumei Lu, Famei Li. Determination and pharmacokinetic study of amlodipine in humanplasma by ultra performance liquid chromatography-electrospray ionization mass spectrometry. J Pharm Biomed Anal. 2007; 43: 1540-5
• 21 Mignini F, Tomassoni D, Traini E, Amenta F. Single-Dose, Randomized, Crossover Bioequivalence Study of Amlodipine Maleate versus Amlodipine Besylate in Healthy Volunteers. Clin Exp Hypertens. 2007; 29: 539-52
• 22 Ragno G, Cione E, Garofalo A, Genchi G, Ioele G, Risoli A et al. Design and monitoring of photostability systems for amlodipine dosage forms. Int J Pharm. 2003; 265: 125-32
• 23 Naidu KR, Kale UN, Shingare MS. Stability indicating RP-HPLC method for simultaneous determination of amlodipine and benezepril hydrochloride from their combination drug product. J Pharm Biomed Anal. 2005; 39: 147-55
• 24 Chaudhari BG, Patel NM, Shah PB. Stability indicating RP-HPLC method for simultaneous determination of atorvastatin and amlodipine from their combination drug products. Chem Pharm Bull. 2007; 55: 241-6