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Abstract
A high performance liquid chromatographic (HPLC) method was developed for the determination of phenobarbital in human breast milk and plasma. The chromatographic behaviors of phenobarbital and other barbiturate derivatives on various sorbents, including octadecyl (C18), octyl (C8), methyl (C1), cyanopropyl (CN), phenyl (Ph), and dimethyl tert-butyl (tert-But), were investigated as stationary phases, and then chromatographic separation was achieved by C8 analytical column using a potassium dihydrogenphosphate buffer (pH 3.5 for milk, pH 6.5 for plasma) / acetonitrile (70 : 30, v/v) as the mobile phase because interference peaks affected differently in each specimen. Phenobarbital and 5-methyl-phenobarbital as an internal standard were detected by ultraviolet detection method at 230 nm. Phenobarbital was extracted by a rapid and simple procedure based on C18 bonded-solid phase extraction in breast milk and plasma.
Determination of phenobarbital in human breast milk and plasma was possible in the concentration range of 0.05 - 30μg/mL. The recoveries of phenobarbital added to human breast milk and plasma were 83.8 - 100.6 % and 95.6 - 102.0 %, respectively, with coefficient of variation of less than 6.6 % and 8.4 %.
This method was used for drug-level monitoring in human breast milk and plasma from patients who were being treated with phenobarbital. The mean concentration of phenobarbital in breast milk and plasma was 6.05 ± 1.2 μg/mL and 14.0 ± 9.0 μg/mL, respectively. The average ratio between the breast milk concentration versus plasma concentration (M/P ratio) was 0.35 ± 0.1.