High-performance liquid chromatography with mass-selective detection is recognized as the «gold standard» for therapeutic drug monitoring in the leading laboratories of the world. One of the drugs that require monitoring of concentrations at a high level is a proliferative signal inhibitor — everolimus, prescribed to patients after transplantation of solid organs [1, 2].
THE AIM OF THE STUDY
Was to develop an analytical technique for the quantitative determination of everolimus in human whole blood by high-performance liquid chromatography with mass selective detection that meets the requirements for methods for therapeutic drug monitoring.
MATERIALS AND METHODS
To create a method of quantitative determination of everolimus in human whole blood by high-performance liquid chromatography with mass selective detection, a laboratory and instrumental complex including a highly efficient liquid chromatograph «Agilent Technologies» with a reversed-phase chromatographic column Poroshell 120 EC-C18 [50 mm/3.0 mm/2.7 micron] was used. The mass spectrometer of the triple quadrupole type with the electrospray ionization system was used as a detector. Buffer solutions based on deionized water and methanol were used as the mobile phase. Standard pharmacological substance of everolimus was used for selection of optimal analytical conditions, as an internal standard acted deuterated everolimus. Testing of analytical methods for suitability for use in clinical practice is carried out in accordance with international requirements for validation of bioanalytic methods [3, 4].
RESULTS
The method of preliminary preparation of whole blood for the study by high-performance liquid chromatography with mass-selective detection was experimentally selected. The conditions of separation of the biological mixture on the chromatographic column. The conditions of detection of everolimus and internal standard, using a three-quadrupole mass analyzer with ionization system «electrospray». The technique was tested in accordance with international requirements for validation of bioanalytic techniques and was tested on real samples of patients after heart transplantation, taking everolimus with parallel measurement of drug concentrations in the reference laboratory.
SUMMARY
The developed method of quantitative determination of everolimus in human whole blood by high-effect liquid chromatography with mass-selective detection meets international requirements for validation of bioanalytic techniques, demonstrating high sensitivity, specificity, reproducibility, and accuracy. Allows you to build a linear calibration relationship in the desired range of concentrations. The technique has been successfully implemented in clinical and laboratory practice of the Almazov National Medical Research Centre and demonstrated good results of external quality control.
