Application of Substitution Method of Reference Substance in the Rapid Determination of Sedative Drugs by HPLC

OBJECTIVE To establish a rapid HPLC method for sedative drugs (chlordiazepoxide，midazolam，nitrazepam，estazolam, oxazepam, lorazepam and alprazolam) by substitution method of reference substance. METHODS The Agilent ZORBAX Eclipse Plus C₈ column(4.6 mm×150 mm, 5 μm) was adopted. The mobile phase was 0.01 mol·L^-1 potassium didydrogen phosphate(pH 2.5)-(methanol-acetonitrile 1∶1)(57∶43). The flow rate was 1.0 mL·min^-1 and the column temperature was 30 ℃. The qualitation was measured by relative retention time and spectral similarity. The relative correction factor for quantification analysis was used for content determination. RESULTS The 7 sedative drugs got complete separation. The qualitation was more accurate by applying the double indicator. The relative correction factor method for quantification which required less reference substances in the on-site screening enabled rapid drug-testing. CONCLUSION This method is rapid, simple and reliable for the rapid determination of sedative drugs.