Quantitative measurement of 4-hydroxy tamoxifen in human plasma and mammary tumours by combined gas chromatography/negative chemical ionization mass spectrometry

A highly sensitive and specific assay was developed for the quantitative measurement of 4-hydroxy tamoxifen (4-OH Tam) at the femtomole level in human plasma and mammary tumours. The drug and deuterated internal standard (4-OH Tam D4) were measured by gas chromatography/negative chemical ionization mass spectrometry with methane as the reactant gas. The two compounds of interest were isolated from the complex biological matrices using a solid-phase extraction procedure with Extrelut 1 columns. Soft operating conditions were required to convert 4-OH Tam to the fluorinated derivatives with pentafluorobenzyl chloride. The mass spectrometer was tuned to monitor the abundant and stable molecular ions at m/z 581 and 585 which were generated in the ion source by an electron capture process. This assay required only 0.5 ml of plasma or 0.5 g of mammary tissue, and the quantification limits of the method were 20 pg ml−1 for the body fluids or 100 pg g−1 for the tissue samples. The very low relative standard deviation and mean percentage error calculated during the different within-day or day-to-day repeatability assays have clearly demonstrated the ruggedness of the technique for routine analysis of 4-OH Tam.