Application of micellar electrokinetic capillary chromatography to forensic analysis of barbiturates in biological fluids.

Micellar electrokinetic capillary chromatography (MECC) is a form of capillary zone electrophoresis. Addition of a surfactant produces micelles in an aqueous/organic buffer. Separation of drugs is obtained via differences in the electrophoretic mobilities of the analytes within the capillary, resulting from their electrophoretic velocity and the electroosmotic flow of the buffer in a given electric field. The migration order is determined by the differential partitioning of the drugs between the micelles and the aqueous/organic phase. Barbiturates were extracted from various biological fluids at pH 4.5 with TOXI-TUBES B. MECC analyses were performed using a Waters Quanta 4000 Capillary Electrophoretic System with a 745 Data Module with a 75 microns x 60 cm capillary and an aqueous/organic buffer of 85% 10 mM borate, 10 mM phosphate, 100 mM sodium dodecyl sulfate and 15% acetonitrile at a pH of 8.5 with a voltage of 20 kV using ultraviolet absorption detection at 214 nm. Migration times were: phenobarbital, 7.78 min.; butalbital, 8.01 min.; butabarbital, 8.23 min.; mephobarbital (internal standard), 8.88 min.; amobarbital, 9.41 min.; pentobarbital, 10.03 min. and secobarbital, 10.79 min. Correlation coefficients (r) between peak areas and concentration ranges of 3 to 60 micrograms/mL were from 0.964 to 0.999. Coefficients of variation (CV) ranged from 2.6 to 8.6% between days and 2.3 to 9.8% within day. Application of this methodology to four forensic cases of butalbital intoxication detected concentrations of 0.7 to 12.7 micrograms/mL in blood; 0.8 to 1.9 micrograms/mL in vitreous humor and 1.5 to 7.6 micrograms/mL in urine. MECC is applicable to forensic analysis of barbiturates extracted from biological fluids.