An UPLC–MS/MS method for separation and accurate quantification of tamoxifen and its metabolites isomers

•Development and validation of an UPLC–MS/MS assay of tamoxifen and metabolites.•E-, Z-, Z′-endoxifen, 4-, 4′-hydroxytamoxifen resolution.•Tamoxifen and metabolites quantification in patient plasma samples. A selective and accurate analytical method is needed to quantify tamoxifen and its phase I metabolites in a prospective clinical protocol, for evaluation of pharmacokinetic parameters of tamoxifen and its metabolites in adjuvant treatment of breast cancer. The selectivity of the analytical method is a fundamental criteria to allow the quantification of the main active metabolites (Z)-isomers from (Z)′-isomers. An UPLC–MS/MS method was developed and validated for the quantification of (Z)-tamoxifen, (Z)-endoxifen, (E)-endoxifen, Z′-endoxifen, (Z)′-endoxifen, (Z)-4-hydroxytamoxifen, (Z)-4′-hydroxytamoxifen, N-desmethyl tamoxifen, and tamoxifen-N-oxide. The validation range was set between 0.5ng/mL and 125ng/mL for 4-hydroxytamoxifen and endoxifen isomers, and between 12.5ng/mL and 300ng/mL for tamoxifen, tamoxifen N-desmethyl and tamoxifen-N-oxide. The application to patient plasma samples was performed.