Simultaneous analysis of antipyrine and lorazepam by high-performance liquid chromatography

A novel analytical methodology was developed for simultaneous high-performance liquid chromatographic quantitation of antipyrine and lorazepam in biological fluids using phenacetin and flunitrazepam as internal standards, respectively. A gradient solvent system was used to achieve specificity and sensitivity for each of the compounds in biological fluids. The between-run coefficients of variation for replicate analysis of antipyrine and lorazepam in identical plasma samples were 9.4 and 6.4% at 60 micrograms/ml and 5.5 micrograms/ml, respectively, for antipyrine, and 9.1 and 9.9% at 110 ng/ml and 10.5 ng/ml, respectively, for lorazepam. Accuracy ranged from 99.7 to 101.4% at concentrations of 8, 45 and 105 micrograms/ml for antipyrine and from 94.8 to 95.8% at concentrations of 12, 55 and 125 ng/ml for lorazepam. This method requires only one organic extraction procedure for all four compounds, allows quantitation of these two drugs in samples also containing indocyanine green, and sample throughput following extraction is totally automated.