Development of HPLC method for the determination of enalapril in tablets using salts of chaotropic anions

Abstract

Leading pharmacopoeias of the world regulate performing chromatographic determination of enalapril in the substance using gradient elution (for 30 min) and a chromatographic column of 4.1 mm × 15 cm, 5 μm, class L21, at a temperature of 70 °C, with a flow rate of 1.5 ml/min, and in tablets on a column of class L7, at a temperature of 50 °C, with a flow rate of 2.0 ml/min, using isocratic elution with acetonitrile (ACN) and buffer (sodium dihydrogen phosphate solution in water, adjusted with phosphoric acid to pH 2.2) (250:750). In scientific literature, researchers have proposed HPLC methods for determining enalapril in pharmaceutical products; however, such methods have drawbacks (asymmetrical peaks, elution near the “dead volume”, etc.). We propose the use of salts of chaotropic anions in the mobile phase on a C18 chromatographic column as a promising approach to obtain a symmetrical enalapril peak that elutes not near the “dead volume”.

The aim of the work was to develop a green, rapid, and simple HPLC method for the determination of enalapril in tablets using salts of chaotropic anions.

Materials and methods. The study used a Shimadzu LC-2050C and Agilent 1260 liquid chromatograph with a diode array detector; LabSolutions software was used for obtaining chromatograms and integrating results. Other analytical equipment included analytical electronic laboratory balances “RAD WAG AS 200/C”; ultrasonic bath (Elmasonic Easy 40 H, Germany); pH meter (Mettler-Toledo, model LE438, Switzerland). The chromatographic column Luna C18 (100 × 4.6 mm, 3 µm) was purchased from Phenomenex. Enalapril maleate reference standard (purity ≥99 %) was acquired from Sigma-Aldrich Chemicals Co., and “Enap” tablets (20 mg, KRKA, Slovenia) were used.

Results. The use of salts of chaotropic anions in the mobile phase during HPLC determination of enalapril made it possible to reduce analysis time (without performing the determination near the dead volume) and to improve the symmetry of the enalapril peak. During the conducted experimental studies, the optimal chromatographic conditions for the quantitative determination of enalapril in tablets were established: Luna C18 chromatographic column (100 × 4.6 mm, 3 μm), mobile phase – 2 % ACN and 98 % KPF6 buffer solution 40 mM, pH 2.43, column temperature – 30 °C, detection at a wavelength of 210 nm. The method was linear in the concentration range of 40–120 μg/mL. Regression equation: y = 14166x – 9589.2, correlation coefficient R² = 0.9972. Low limit of determination (LOD) – 8.52 μg/mL, low limit of quantification (LOQ) – 25.80 μg/mL. Modern tools for assessing greenness, AGREE (score 0.74), MoGAPI (score 81), Complex MoGAPI (score 81), AGSA (score 77.78), CaFRI (score 82), and CACI (score 79), confirmed that the proposed HPLC method is green.

Conclusions. A green, rapid, and simple HPLC method for the determination of enalapril in tablets using chaotropic anion salts has been developed. The proposed approach enabled the production of symmetrical peaks with excellent chromatographic system parameters and allowed analysis in a short time. In addition, reducing the mobile phase flow rate to 0.6 ml/min made the analysis greener. The developed HPLC method for determining enalapril in tablets can be applied both in routine pharmaceutical analysis and in testing by independent laboratories.