Development, Validation, and Clinical Application of a Simultaneous Quantification Method of Seventeen Antituberculosis Drugs in Plasma, Serum, or Saliva Using LC-MS/MS.

BACKGROUND

Tuberculosis (TB) remains a global health issue. Treatment involves multiple anti-TB drugs, and pharmacokinetic variability may lead to toxicity or ineffective treatment. Therapeutic drug monitoring may help address these challenges. This study aimed to develop a rapid method to analyze anti-TB drugs in plasma, serum, and saliva.

METHODS

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for simultaneous quantification of anti-TB drugs in plasma, serum, and saliva. The method was validated according to European Medicines Agency and Food and Drug Administration guidelines, assessing linearity, selectivity, carry-over, accuracy, precision, dilution integrity, matrix effect, recovery, and stability. Performance was evaluated across plasma, serum, and saliva and in an external quality control program.

RESULTS

A method with a runtime of 1.7 minutes was developed to analyze 17 anti-TB drugs and 6 metabolites: (N-desmethyl-)bedaquiline, clofazimine, delamanid, ethambutol, ethionamide, gatifloxacin, (acetyl-)isoniazid, (nor)levofloxacin, linezolid, moxifloxacin, pretomanid, protionamide, pyrazinamide, (desacetyl-)rifabutin, (desacetyl-)rifampicin, (desacetyl-)rifapentine, and tedizolid. The method demonstrated linearity, selectivity, and minimal carry-over. Accuracy and precision were within acceptable limits for all drugs and metabolites, with the largest biases observed for rifabutin (-13.5%) and rifapentine (11.6%). Most drugs were stable, although delamanid and protionamide required immediate processing. All analytes met the precision criteria (CV <15%) in serum and saliva, whereas matrix-specific calibrators were required for some analytes to achieve acceptable accuracy (bias <15%).

CONCLUSIONS

This validated LC-MS/MS method allows simultaneous analysis of multiple anti-TB drugs in plasma, serum, and saliva, facilitating clinical and research applications. The method has been successfully applied in clinical practice.