A high performance liquid chromatography tandem mass spectrometry for the quantification of tacrolimus in human bile in liver transplant recipients

作者：Tron Camille^*; Rayar Michel; Petitcollin Antoine; Beaurepaire Jean Marie; Cusumano Caterina; Verdier Marie Clemence; Houssel Debry Pauline; Camus Christophe; Boudjema Karim; Bellissant Eric; Lemaitre Florian

来源：Journal of Chromatography A, 2016, 1475: 55-63.

DOI：10.1016/j.chroma.2016.10.075

• Tacrolimus
• High performance liquid chromatography
• Mass spectrometry
• Bile
• Alternative matrix
• Therapeutic drug monitoring

Tacrolimus whole-blood concentrations imperfectly reflect concentrations at the effect site. Tacrolimus concentrations in the transplanted organ could be more relevant to predict rejection events. Because liver biopsy cannot be repeatedly performed after liver transplantation, we suggested measuring tacrolimus in the bile to have a cost-effective and clinically implementable surrogate marker of intra-hepatic tacrolimus concentration. We developed and fully validated a liquid chromatography-tandem mass spectrometry method for the determination of tacrolimus in human bile. Sample purification was achieved using protein precipitation and liquid-liquid extraction with ethyl-acetate. Gradient elution was performed using a C18 analytical column with a 5 min run-time. The method was linear from 0.5 ng/mL to 20 ng/mL. In this concentration range, within-day and between-day precisions as well as overall bias were within 15%. Matrix effect was fully corrected by the internal standard (ascomycin). The assay was optimized to achieve good selectivity in this complex biological matrix. Tacrolimus was found to be stable in bile stored 6 months at -80 degrees C, after 3 freeze and thaw cycles, 20 h at room temperature and 24 h in extracts kept at 15 degrees C in the auto-sampler. The method was applied to quantify tacrolimus in bile from liver transplant recipients. It allowed getting preliminary data about tacrolimus excretion profile in bile and showed the lack of correlation between tacrolimus whole blood concentration and tacrolimus liver exposition. This alternative and innovative analytical approach of tacrolimus bio-analysis appears suitable for further studies evaluating relevance of biliary tacrolimus concentration as a new pharmacological marker of immunosuppressive activity.

• 出版日期2016-12-2