Analysis of Tamoxifen and Its Metabolites by On-Line Capillary Electrophoresis−Electrospray Ionization Mass Spectrometry Employing Nonaqueous Media Containing Surfactants

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Abstract
On-line capillary electrophoresis−electrospray ionization mass spectrometry (CE−ESMS) has been employed for the analysis of metabolites of the anticancer drug tamoxifen. Nonaqueous (methanol) CE electrolyte provided better resolution and detection sensitivity compared to aqueous systems or highly aqueous water−methanol electrolyte mixtures. Nonaqueous methanol also permitted the use of lower ES voltages presumably owing to its lower surface tension, which facilitated droplet breakup. This decreased the tendency to produce electric discharges, thus improving the stability of electrospray conditions. The relative ease of methanol solvent evaporation may contribute to an improved yield of protonated analytes as compared to highly aqueous solutions. Enhanced CE resolution can be at least partially attributed to the improved solubility of analytes in methanol relative to water. Higher solubility implies less aggregation of hydrophobic analytes, thus improving homogeneity in solution. Moreover, electroosmotic flow toward the detector decreased in methanol relative to water. The reduction of this force pushing all analytes through the capillary, but not aiding in separation, implies that other factors such as slight differences in electrophoretic mobilities are more apt to lead to successful separations. Surfactants were employed as nonaqueous CE−ESMS buffer additives. An SDS concentration of 7 mM lowered the ESMS signal response for N-desmethyltamoxifen by a factor of ∼3. However, separation of tamoxifen metabolites using 7 mM SDS was augmented relative to the unadulterated methanol electrolyte. This enabled the separation of α-hydroxytamoxifen and 4-hydroxytamoxifen, which were not resolvable in methanol electrolyte devoid of SDS. The methanol−surfactant electrolyte system has been successfully used to determine metabolites formed after incubation of tamoxifen with mouse hepatocytes.