|Budget Amount *help
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1996: ¥300,000 (Direct Cost: ¥300,000)
Fiscal Year 1995: ¥1,700,000 (Direct Cost: ¥1,700,000)
1 The methods of determination of the racemic and enatiomers of ketamine (K) and norketamine (NrK) in plasma were established.
Instruments : (HITACHI) L-6000 pump, L-7300 column oven, L-7400 UV Detector, D-2500 Recorder.
Column : NBS-C-18 (0.46*250cm) for racemic type.
DAICEL-CHIRICELL-OD (0.46*250cm) for enatiomer type.
Extraction and Quantitation : Ketamine and norketamine were extracted from 0.5ml plasma that added a known amount of Br-ketamine as internal standard with 2.5ml of 5% isopropyl alcohol and 95% n-butyl chloride. The upper organic layrs were transfered to disposable glass culture tubes (12*75mm) and evaporated to dryness by a cintrifugal vacuum evaporator. The residues were disolved with the 0.15ml of HPLC buffer and purified with millipor centrifuge filters. The volume of injected sample into HPLC were 0.05ml respectivery.
The mobile phase folw-rate was 0.5/ml/min in 30ﾟC of column temperature, and the UV detector was set at 215nm.
Chromatography and Selectivity
(A) Rasemic K and NrK : The retenntion time of NrK,K and Br-k were 9.4,10.9,13.9min. respectivery. The coefficient variation (CV) were K=4.0%, Nrk=5.3% in conc.of 125-1000ng/ml. Minimum detectable conc.was 5ng/ml.
(B) Enatiomer (R-, S-) K and NrK : The retention time of R-K,S-K,S-NrK and R-Nrk were 18.1,20.7,32.8,33.8min, respectivery. Br-K as internal standard were 19.3 and 23.4min. The CV of R-K,S-K,S-Nrk and R-NrK were 2.8,2.9,7.9 and 7.3% (125-500ng/ml), respectivery. Minumum detectable conc.was 10ng/ml.
2 Determination of K and Nrk during DFK anesthesia (10 patients)
During 2 hours of DFK Anesthesia, there was no stastic difference observed between enatiomer of K and NrK.
Although long time anesthesia over 6 hours (5 patients), the results was same.
3 Discussion : In limited budget, we could not buy a autosampler injector. It took long time to measure the samples, so we stocked in freezer many kinds of samples. In the ferture, we have to estimate these samples.