Efficient Quantification of Tacrolimus in Rat Plasma With Ultra‐High Performance LC–Q–Orbitrap MS: A Precision Pharmacokinetic Analysis Tool

ABSTRACT The narrow therapeutic index of tacrolimus demands precise dose optimization to maintain therapeutic efficacy while minimizing toxicity, underscoring the necessity for reliable analytical techniques to accurately measure drug concentrations in biological matrices. In this study, we developed an ultrahigh‐performance liquid chromatography coupled with quadrupole‐orbitrap mass spectrometry method for quantifying tacrolimus in plasma. This approach employed an Acquity UPLC BEH C 18 column, leveraging a gradient elution of methanol (A) and water with 0.1% formic acid (B) to ensure meticulous separation of tacrolimus and the internal standard, ascomycin. Operating in positive ionization mode, the system utilized full MS/dd‐MS2 scans, capturing comprehensive data for heightened analytical accuracy and precision. The method demonstrated a broad linear range of 0.5 to 500 μg/L, with a low quantification limit of 0.5 μg/L. Notably, it achieved exceptional reproducibility and accuracy, with relative standard deviations and errors below 6%, complemented by consistent extraction efficiencies for tacrolimus at 84.78% to 97.15%. This study successfully showcases an advanced analytical platform for the meticulous quantification of tacrolimus, seamlessly integrating into pharmacokinetic investigations following a 1.2 mg/kg oral dose in rats. The method's deployment revealed its exceptional suitability for capturing intricate pharmacokinetic profiles, underscoring its value in preclinical drug disposition studies.