Development and validation of a fast HPLC method for methyldopa enantiomers using superficially porous particle based macrocyclic glycopeptide stationary phase

Abstract

Methyldopa is a centrally acting antihypertensive agent, whose pharmacological activity is attributed to the S-α-methyldopa enantiomer. The assessment of enantiomeric purity is an essential quality control test for chiral drugs, as the presence of certain impurities in a drug can interfere with the desired pharmacological effect. The present study was undertaken to evaluate different chiral stationary phases composed of macrocyclic glycopeptides (vancomycin, teicoplanin and aglycone teicoplanin) and the use of superficially porous particles for the efficient separation of methyldopa enantiomers. The reversed-phase and polar ionic elution modes were evaluated, with different proportions of solvents and additives. The influence of the flow rate and temperature on the analysis were also evaluated. Different stationary phases in different elution modes exhibited suitable selectivity and fast analysis. Total separation of the enantiomers were obtained with the teicoplanin and teicoplanin aglycone stationary phases. The teicoplanin aglycone column produced excellent resolution values and very fast separation analyzes. The use of the teicoplanin aglycone chiral stationary phase for the analysis of MDopa enantiomers is innovative and has not yet been explored in the literature. The best method was obtained and validated with teicoplanin aglycone in the reversed-phase mode using 20 mM ammonium acetate buffer pH 4.0/MeOH (20:80), column temperature of 45 °C and flow rate of 1.0 mL/min, which presented a total run time less than 5 min, a resolution of 5.05 and good peak symmetry. The method can be used in pharmaceutical quality control laboratories and support the stereospecific synthesis of the drug.