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  • Title: Comparison of a stable isotope-labeled and an analog internal standard for the quantification of everolimus by a liquid chromatography-tandem mass spectrometry method.
    Author: Heideloff C, Payto D, Wang S.
    Journal: Ther Drug Monit; 2013 Apr; 35(2):246-50. PubMed ID: 23503452.
    Abstract:
    BACKGROUND: Everolimus is an immunosuppressant drug used in solid organ transplantation. Immunoassays and liquid chromatography-mass spectrometry (LC-MS) methods have been used for therapeutic drug monitoring of this drug. In LC-tandem mass spectrometry (MS/MS) methods, both 32-desmethoxyrapamycin and everolimus-d4 have been used as internal standards. OBJECTIVES: To compare 2 internal standards (32-desmethoxyrapamycin and everolimus-d4) for the quantification of everolimus by an LC-MS/MS method. METHODS: Both 32-desmethoxyrapamycin and everolimus-d4 were introduced in the method validation process with 2 transitions simultaneously monitored for everolimus (975.6 → 908.7 as the quantifier and 975.6 → 926.9 as the qualifier) by an established LC-MS/MS method. The key performance characteristics were lower limit of quantification, accuracy, precision, and comparison with an LC-MS/MS method offered by another laboratory. RESULTS: The lower limit of quantification (LLOQ) was 1.0 ng/mL using either internal standard with an analytical recovery of 98.3%-108.1% across the linear range. The total coefficient of variation for everolimus was 4.3%-7.2% with no significant difference between the 2 internal standards. In comparison with an independent LC-MS/MS method, though everolimus-d4 offered a better slope (0.95 versus 0.83), both internal standards showed acceptable results and had a coefficient of correlation r > 0.98 in the tested concentration range of 1.2-12.7 ng/mL. CONCLUSIONS: Although everolimus-d4 offered a more favorable comparison with an independent LC-MS/MS method, both everolimus-d4 and 32-desmethoxyrapamycin had acceptable performance as the internal standards for everolimus quantification by the LC-MS/MS method.
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