Preparation of Hydromorphone-Ketamine Solutions
Three independently prepared combinations of the 2 drugs were evaluated in each of 3 types of container. Stock solutions of the combination of hydromorphone (Sandoz Canada, Boucherville, Quebec; lot 137324, expiry January 2009) and ketamine (Sandoz Canada; lot 139132, expiry April 2009) were prepared from commercially available solutions, diluted in NS (Baxter Corporation, Toronto, Ontario; lot W7J24CO, expiry May 2009) to concentrations of 0.2 and 0.2 mg/mL, 0.2 and 0.6 mg/mL, and 0.2 and 1.0 mg/mL, respectively. Portions of each combined solution were transferred to brown glass bottles (Richards Distribution, Richmond, British Columbia; 100-mL samples), plastic syringes (Becton Dickinson, Oakville, Ontario; lot 8058879; 50-mL samples), and IV bags (Baxter Corporation; lot W7L17CO; 50-mL samples). All solutions were kept in the dark at room temperature (25°C).
The physical characteristics of the solutions were evaluated qualitatively at the time of preparation and on days 1, 2, 3, and 7. As samples were collected for analysis during the study period, each solution was tested by the same individual (D.D.) and was visually examined for changes in colour (against white and black backgrounds) and for formation of precipitate. Three aliquots from each bottle, syringe, and IV bag were collected at each time point for determination of pH. The pH meter was calibrated using commercially available standards at the beginning of each testing session. Immediately following the physical observations, 1.5-mL samples were transferred to 2-mL polypropylene vials (VWR, Edmonton, Alberta; lot 108069978), which were immediately stored at —85°C until analysis up to 6 weeks later by stability-indicating high-performance liquid chromatography (HPLC) with ultraviolet detection. Kamagra Oral Jelly
Preparation of Stocks, Standards, and Standard Curve
Stock solutions were prepared as follows: hydromorphone 2 mg/mL and ketamine 10 mg/mL were each diluted in NS to a concentration of 1 mg/mL. The internal standard, naloxone 1 mg/mL (Sandoz Canada; lot 138390), was diluted in NS to a concentration of 0.5 mg/mL. Standard solutions containing 0.05 mg/mL of the internal standard were prepared in NS with the following concentrations of hydromorphone and ketamine, respectively (mg/mL): 0.26 and 0.26, 0.23 and 0.23, 0.20 and 0.20, 0.17 and 0.17, 0.10 and 0.13, 0.05 and 0.09, and 0.03 and 0.08. Each standard was passed through a 0.45-^m microfilter to prevent injection of impurities onto the column.
A 7-point calibration curve was prepared with a blank (NS) at the beginning of each run, to ensure that there was no carry-over from one run to the next. The range of this calibration curve (from 0.26 and 0.26 mg/mL to 0.03 and 0.08 mg/mL hydromorphone and ketamine, respectively) encompassed the diluted test concentrations of hydromor- phone 0.2 mg/mL with ketamine 0.2 to 1.0 mg/mL. The calibration curve was generated by least-squares regression of the peak area ratio of hydromorphone—ketamine to naloxone (internal standard) and the concentration of each standard (hydromorphone—ketamine). Accuracy of the assay was calculated as the mean deviation between nominal and observed concentrations on 4 replicate samples (hydromorphone 0.15 mg/mL and ketamine 0.15 mg/mL). The precision of the assay was evaluated by intraday and interday validation methods. Intraday variability was determined by running the following stock solutions in quadruplicate throughout a single day: hydromorphone and ketamine 0.25 and 0.25, 0.15 and 0.15, 0.05 and 0.09, and 0.03 and 0.08 mg/mL. Interday variability was determined by running the same concentrations (as in the testing for intraday variability) in quadruplicate daily for 4 days. The means, standard deviations, and coefficients of variation were then calculated. Acceptable limits for the coefficients of variation were defined a priori as less than 10%.
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