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. 2022 Mar 12;14(3):632.
doi: 10.3390/pharmaceutics14030632.

Development and Validation of the New Liquid Chromatography-Tandem Mass Spectrometry Method for the Determination of Unbound Tacrolimus in the Plasma Ultrafiltrate of Transplant Recipients

Magdalena Bodnar-Broniarczyk  1 , Karola Warzyszyńska  2 , Katarzyna Czerwińska  3 , Dorota Marszałek  1 , Natalia Dziewa  2 , Maciej Kosieradzki  2 , Tomasz Pawiński  1
Affiliations

Development and Validation of the New Liquid Chromatography-Tandem Mass Spectrometry Method for the Determination of Unbound Tacrolimus in the Plasma Ultrafiltrate of Transplant Recipients

Magdalena Bodnar-Broniarczyk et al. Pharmaceutics. .

Abstract

(1) Background: Only unbound tacrolimus particles are considered to be active and capable of crossing cellular membranes. Thus, the free-drug concentration might be better associated with clinical effects than the total drug concentration used for dosage adjustment. We propose a new, fully validated online liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for unbound tacrolimus concentration measurement. (2) Methods: The determination of the unbound tacrolimus concentration in plasma ultrafiltrate was performed with the Nexera LC system with LCMS-8050 triple quadrupole MS using ascomycin as an internal standard. Chromatographic separation was made using a HypurityC18 analytical column. MS/MS with electrospray ionization and positive-ion multiple-reaction monitoring was used. The unbound tacrolimus level was determined in 36 patients after solid organ transplantation (n = 140). (3) Results: A lower limit of quantification 0.1 pg/mL was achieved, and the assay was linear between 0.1 and 20 pg/mL (R2 = 0.991). No carry-over was detected. The within-run and between-run accuracies ranged between 97.8-109.7% and 98.3-107.1%, while the greatest imprecision was 10.6% and 10.7%, respectively. Free tacrolimus in patients' plasma ultrafiltrate varied between 0.06 and 18.25 pg/mL (median: 0.98 pg/mL). (4) Conclusions: The proposed method can be easily implemented. The significance of the unbound tacrolimus concentration needs to be investigated. This may facilitate the individualization and optimization of immunosuppressive treatment.

Keywords: LC–MS/MS; TDM; tacrolimus; ultrafiltration; unbound concentration; validation.

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Conflict of interest statement

The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Figures

Figure 1
Figure 1
Representative LC-MS/MS chromatograms of (a) blank plasma ultrafiltrate; (b) tacrolimus at the lower limit of quantification (0.1 pg/mL); (c) unbound tacrolimus in a patient ultrafiltrate sample. * Tacrolimus reference standard (99.1% purity). ** Internal standard ascomycin.
See this image and copyright information in PMC

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