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ABSTRACT
A simple and accurate ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for the first time as a confirmatory method for the simultaneous determination of stilbenes – hexestrol and diethylstibestrol in serum. Extraction was based on a simple acid denaturation of protein followed by liquid–liquid extraction using methyl tert butyl ether. Extracts were directly injected into the UHPLC-MS/MS without further purification. Excellent recoveries in the range 82–99% and 91–128% were obtained for hexestrol and diethylstibestrol, respectively. Both within-day repeatability and between-day reproducibility were generally satisfactory with RSD <20%. The linearity of the internal standard based matrix-matched calibration curve measured as the coefficient of regression (r2) was generally >0.99 for both hexestrol and diethylstibestrol. Both matrix effect and uncertainties associated with sample preparation and instrumental analysis were significantly reduced with the use of a deuterated compound (hexestrol-d4) as internal standard. The LOD and LOQ were 0.09 and 0.08 ng/ml, and 0.28 and 0.25 ng/ml, respectively, for hexestrol and diethylstibestrol. The method was found to be suitable for the simultaneous determination of hexestrol and diethylstibestrol in serum.
Conflict of interest
Ovokeroye Abafe declares that he has no conflict of interest. Pumza Gatyeni declares that she has no conflict of interest. Tlou Chokwe declares that he has no conflict of interest.
Ethical approval
All applicable national guidelines for the care and use of animals were followed. This article does not contain any studies with human participants performed by any of the authors.