Improvement of Electrophoretic Enantioseparation of Amlodipine by Polybrene

Document Type: Research article

Authors

1 School of Pharmacy, Zabol University of Medical Sciences, Zabol, Iran.

2 School of Pharmacy, Shaheed Beheshti University of Medical Sciences, Tehran, Iran.

3 Department of Chemistry, University of Alberta, Canada.

Abstract


   In chiral and non-chiral electrophoretic resolution of basic drugs, adsorption of analytes to negatively charged capillary wall could lead to poor repeatability of migration time and peak area. In addition, chiral resolutions of basic drugs are commonly performed in low pH buffers. Therefore, longer analysis time due to suppression of electroosmotic flow (EOF) is another dilemma. In this work the improvement effect of polybrene (PB), a cationic polymer, on chiral separation of a model basic drug, amlodipine (AML), was investigated. PB both as a semi-permanent coating agent and as an additive in the running buffer was utilized. Better results were obtained with PB as a buffer additive. Compare to untreated bare silica without using PB in running buffer, addition of 0.0005% PB to buffer decreased analysis time downed to 3 folds; efficiency improved up to 5 folds; limit of detection (LOD) and limit of quantification (LOQ) downed to 8 folds and within-day migration time and peak area repeatabilities, in terms of relative standard deviations (RSD) downed to 5 and 20 folds, respectively.

Keywords


 

Introduction

   About 40% of drugs in use are chiral (1, 2) and in most cases two enantiomers of chiral drugs exhibit different pharmacological, toxicological or pharmacokinetic properties (3). Therefore, development of analytical methods for enantiomer separation for controlling synthesis, enantiomer purity check, and for pharmacodynamic studies is attracting area of research. Thus, different analytical techniques have been applied such as high-performance liquid chromatography (HPLC), thin-layer chromatography (TLC), gas chromatography (GC), supercritical fluid chromatography (SFC) and capillary electrophoresis (CE).

   CE has matured to a powerful technique, especially for analytical enantioseparations. This is primarily due to high efficiency and high flexibility with regard to analytes and to the separation conditions, as well as the low consumption of chemicals and solvents. According to a recent discussion forum on the application of CE in the pharmaceutical industry, the major application of the technique is chiral separations (4, 5).

   However, CE has some limitations, especially for chiral and non-chiral resolution of basic drugs. Basic analytes are positively charged and electrostatically attracted to the negatively charged capillary walls. This irreproducible adsorption can lead to migration time and peak area variations (6), and consequently to lower precision. Peak tailing can also mask impurity peaks and may also deteriorate resolution of enantiomers in which two analytes migrate closely after each other. Furthermore, analyte adsorption could raise limit of detection (LOD) and limit of quantification (LOQ) of analysis.

   There are several strategies to avoid undesirable interactions of the analytes with the capillary wall. These include: use of high salt concentration, extremes of pH, buffer additives, and coated capillaries (6). Different approaches have been applied for coating of capillaries: covalently/ cross-linked polymer (7), adsorbed cationic polymers (8-11), adsorbed nonionic polymers (12) and adsorbed surfactants (13-17). Dynamic coatings are more attractive compared to covalently bonded polymer coatings, due to their simplicity, versatility, and low cost. The use of dynamically coated capillaries allows for the rapid, precise, and reproducible separation of moderate to strong basic analytes (pKa > 5) at pH 2.5 (18).

   In this work, the effect of a cationic polymer, polybrene (PB), on the improvement of chiral resolution of a model basic drug, amlodipine (AML), was studied. AML (Figure 1) is a potent dihydropyridine calcium antagonist used for the treatment of hypertension and angina pectoris. Only the S (-) isomer of the drug exerts vasodilating action (19).

 

Figure 1. AML structure , pka 8.6 (18). 

 

Experimental

  All solutions were prepared in Nanopure 18 MΩ ultrapure water (Barnstead, Chicago, IL). Sodium phosphate buffer pH 2.5 was prepared from sodium phosphate monobasic monohydrate (EM sciences, Fort Washington, PA), and adjusting the pH with orthophosphoric acid (BDH, Darmstadt, Germany). PB