[The simultaneous determination of ketamine and midazolam using high pressure liquid chromatography and UV detection...

In order to prevent some negative side effects of ketamine during "dissociative anaesthesia", this substance is regularly combined with benzodiazepines, e.g., midazolam. This study was undertaken to develop a simple and effective method for the simultaneous detection of ketamine and midazolam in plasma by high-pressure liquid chromatography with UV-detection (HPLC/UV). METHODS. The chromatographic system consisted of a 300 x 3.9-mm C18 column for reversed-phase chromatography and a mobile phase of 30% acetonitrile and 70% 0.05 M sodium phosphate buffer, pH 4.45. The flow rate was 1 ml/min. UV-detection took place at a wavelength of 210 nm. Blood samples were preferably taken from a central venous or arterial line. After plasma separation, 1 microgram (100 microliters) etidocaine was added to 1 ml plasma as an internal standard. The samples were alkalinised and extracted with ether, followed by back-extraction of the organic phase in 250 microliters 0.05 N sulphuric acid; 50 microliters of this solution was injected into the system. RESULTS. Sample preparation by trained personnel was reliable and led to adequate results. Separation of ketamine and midazolam was very satisfactory. Standard curves for both drugs were linear from 15-4000 ng/ml (ketamine r = 0.9998; midazolam r = 0.9996). Recovery rates for ketamine in plasma were above 95%, for midazolam 70%-75%. Coefficients of variation for ketamine in plasma were between 0.6% and 1.3%, for midazolam between 2.0% and 2.6%. The detection limit was lower than 5 ng/ml. CONCLUSION. The method is suitable for clinical practice and allows the simultaneous detection of two anaesthetics in wide-spread combined use.