Automated, fast and sensitive quantification of drugs in blood by liquid chromatography–mass spectrometry with on-line extraction: immunosuppressants

We developed a universal LC–mass spectrometry assay with automated online extraction (LC/LC–MS) to quantify the immunosuppressants cyclosporine, tacrolimus, sirolimus and SDZ-RAD alone or in combination in whole blood. After protein precipitation, samples were loaded on a C18 extraction column, were washed and, after activation of the column-switching valve, were backflushed onto the C8 analytical column. [M+Na]+ ions were detected in the selected ion mode. For tacrolimus, sirolimus and SDZ-RAD, the assay was linear from 0.25 to 100 μg/l and for cyclosporine from 7.5 to 1250 μg/l (all r2>0.99). Analytical recovery was >85% and, in general, inter-day, intra-day variability for precision and accuracy were <10%.