Measurement of tacrolimus in blood with the method of high performance liquid chromatography-mass spectrometry

Abstract

To develop a simple, precise, and specific method for measurement of tacrolimus in whole blood. Tacrolimus was extracted from the blood samples of 40 patients treated with tacrolimus by liquid-liquid extraction. Then high performance liquid chromatography-mass spectrometry (HPLC/MS) was used, with ascomycin as internal marker, to isolate the tacrolimus and measure the concentration thereof. The standard curve was drawn. The linearity of quantitative measurement was determined. Detection limits test, CV test, and recovery test were performed. ELISA was used simultaneously. The results of these 2 methods were compared. A good standard curve was drawn based on the results of HPLC/MS assay (y = 0.2464x + 0.0082, R2 = 0.9996). The detected data were highly related to the detected concentrations. The linearity range was 0.100 - 40.000 ng/ml (y = 1.0294x-0.035, R2 = 0.9998). The detection limit of the assay was 0.100 ng/ml. The CV values by this assay were basically < 5%. The recovery rate ranged 95.67% - 98.30% for tacrolimus over the range 0.300 - 16.016 ng/ml. There was a linear correlation (y = 1.0172x + 0.3742, R2 = 0.9630) between the assay results by HPLC/MS to ELISA in blood. The measurement value of ELISA was (19 +/- 9), significantly higher than that of HPLC/MS (18 +/- 9, P < 0.01). This newly developed method of HPLC/MS is simple, precise, and specific and with lower cost, it can be used in the clinical practice and experimental study on tacrolimus.