A LC-Electrospray Tandem MS Method for the Analysis of Naltrexone in Canine Plasma Employing a Molecular Model to Demonstrate the Absence of Internal Standard Deuterium Isotope Effects

Abstract

A simple and sensitive method is described for the determination of naltrexone (NAL), an opioid antagonist, in dog plasma. Sample processing involved a single step liquid-liquid extraction, followed by evaporation of the supernatant, and reconstitution of the residue prior to injection into the liquid chromatograph. The peak height ratio of NAL to [15,15,16-(2)H] naltrexone (NAL-d(3)) was used for quantitation. Observation of the chromatograms for NAL and NAL-d(3) revealed that the mean retention times of the compounds were 1.32 and 1.31 min, respectively. The almost identical retention times possibly accounted for the absence of matrix effects influencing quantitation. Molecular mechanics calculations using SYBYL software were carried out to qualitatively and quantitatively assess analyte and isotopic internal standard stationary phase interactions. Binding energy values of -10.22 and -10.26 kcal/mole were obtained for NAL and NAL-d(3), respectively. These data predict, semi-quantitatively, the absence of deuterium isotope effects that may influence quantitation. Calibration curves were linear from 10 pg/mL to 5014 pg/mL with a weighting factor of 1/x. Precision and accuracy and reverse predicted concentration residuals were within 15%. The method has been used successfully for the analysis of plasma samples from a pilot subcutaneous implantation study in dog.