Large volume sample stacking of antiepileptic drugs in counter current electrophoresis performed in PAMAPTAC coated capillary

Abstract

Electrophoretic stacking is developed for sensitive determination of three zwitterionic antiepileptics, namely vigabatrin, pregabalin and gabapentin, in human serum. CE separation is performed in a 25 μm fused silica capillary covalently coated with the copolymer of acrylamide with 5% content of permanently charged 3-acrylamidopropyl trimethylammonium chloride (PAMAPTAC). In background electrolyte of 500 mM acetic acid, the 5% PAMAPTAC generates an anodic electro-osmotic flow with a magnitude of (-18.6 ± 0.5) · 10^-9 m²V^-1s^-1, which acts against the direction of the electrophoretic migration of the analytes. A sample of the antiepileptic prepared in a 25% v/v infusion solution and 75% v/v acetonitrile is injected into the capillary in a large volume attaining a zone length of up to 270 mm. After turning on the separation voltage, the antiepileptics are isotachophoretically focussed behind the zone of Na⁺ ions with a sensitivity enhancement factor of 78. For the clinical determination of antiepileptics, the human serum is diluted with acetonitrile in a ratio of 1:3 v/v and a zone with a length of 90 mm is injected into the capillary. The method is linear in the 0.025-2.5 μg/mL concentration range; the attained limit of quantification is in the range 18.3-22.8 nmol/L; the within-day precision for the migration time is 0.8-1.2% and for the peak area 1.5-2.4%.

Keywords: Antiepileptic; Capillary electrophoresis; Coating; Contactless conductivity detection; Sample pretreatment; Serum; Stacking.