ABSTRACT
Gliflozins and gliptins represent two different pharmacological drug classes that exert different and potentially complementary glucose-lowering effect in patients with type II diabetes mellitus. A novel, selective, and sensitive HPLC method was developed for the determination of canagliflozin, empaglifozin, linagliptin, and metformin in pure form, in laboratory prepared mixtures, and in pharmaceutical dosage form. Experimental design optimization was applied by using Plackett-Burman and face-centered composite designs to achieve the best resolution with minimum experimental trials. Three significant variables affecting optimization, namely buffer pH, percentage of methanol, and percentage of acetonitrile, were studied. Chromatographic separation was achieved using an Agilent Eclipse C8 column, and column temperature was kept at 45°C. The mobile phase was composed of dipotassium hydrogen phosphate buffer (0.05 M, adjusted to pH 6 using o-phosphoric acid):acetonitrile:methanol (50:25:25, v/v/v) at a flow rate of 1.5 mL/min. Sharp and well-resolved peaks of the cited drugs were obtained. The method was fully validated in terms of linearity, accuracy, precision, selectivity and robustness in agreement with the International Council of Harmonization (ICH) guidelines Q2 (R1). Satisfactory results were obtained by the analysis of tablets through applying the developed method. Therefore, it could be performed for the analysis of the cited drugs in quality control laboratories.
Subject(s)
Chromatography, High Pressure Liquid/methods , Dipeptidyl-Peptidase IV Inhibitors/analysis , Sodium-Glucose Transporter 2 Inhibitors/analysis , Benzhydryl Compounds/analysis , Benzhydryl Compounds/chemistry , Benzhydryl Compounds/isolation & purification , Canagliflozin/analysis , Canagliflozin/chemistry , Canagliflozin/isolation & purification , Dipeptidyl-Peptidase IV Inhibitors/chemistry , Dipeptidyl-Peptidase IV Inhibitors/isolation & purification , Glucosides/analysis , Glucosides/chemistry , Glucosides/isolation & purification , Limit of Detection , Linagliptin/analysis , Linagliptin/chemistry , Linagliptin/isolation & purification , Linear Models , Metformin/analysis , Metformin/chemistry , Metformin/isolation & purification , Reproducibility of Results , Research Design , Sodium-Glucose Transporter 2 Inhibitors/chemistry , Sodium-Glucose Transporter 2 Inhibitors/isolation & purification , TabletsABSTRACT
A novel, highly sensitive ultrasound-assisted dispersive liquid-liquid microextraction (UA-DLLME) high performance liquid chromatography with diode-array detection (HPLC/DAD) method was developed for the determination of empagliflozin, dapagliflozin and canagliflozin in human plasma using methanol as protein precipitating agent/disperser and 1-dodecanol as extracting solvent. The analytes were eluted with an isocratic mobile phase consisting of acetonitrile:aqueous 0.1% trifluoroacetic acid pH 2.5, (40:60, v/v), at a flow rate of 1 mL/min and UV detection at 210 nm. The microextraction conditions were optimized regarding type and volume of extractant, type of disperser, sample pH, extraction time and centrifugation time. Under the optimal conditions, the enrichment factors were 19 for empagliflozin, 27 for dapagliflozin and 50 for canagliflozin. Linearity ranges were 2-2500 ng/mL, 3.5-2500 ng/mL and 1.1-2500 ng/mL for empagliflozin, dapagliflozin and canagliflozin, respectively. The developed method employs very small volumes of organic solvents in sample extraction and allows determination of small concetrations of gliflozins in human plasma.
