Background <p>The first-line anti-tuberculosis (anti-TB) drugs require long-term administration, and their plasma concentrations are closely associated with therapeutic efficacy and safety. The primary objective of this study was to analyze interindividual and interdrug variability in plasma concentrations of first-line anti-TB drugs in pediatric patients and to investigate factors influencing drug concentrations.</p> Methods <p>A retrospective analysis was conducted on 110 pediatric tuberculosis (TB) patients who underwent therapeutic drug monitoring (TDM) for anti-TB drugs between January 2022 and November 2023. Patients with complete TDM data were included, defined as those receiving the tablet formulation at doses consistent with the 2010 World Health Organization recommendations, with blood samples collected 5–7 days after drug administration. Accordingly, TDM data were analyzed for isoniazid (INH, <i>n</i> = 105), rifampicin (RFP, <i>n</i> = 83), and pyrazinamide (PZA, <i>n</i> = 103).</p> Results <p>Two hours after administration, the proportions of children with plasma concentrations below the target range were 31.4% (33/105) for INH, 30.1% (25/83) for RFP, and 3.9% (4/103) for PZA. Multivariate logistic regression analysis revealed that higher doses (mg/kg) were significantly associated with achieving a therapeutic plasma concentration (<i>P</i> &lt; 0.05).</p> Conclusion <p>Subtherapeutic plasma concentrations of RFP and INH are common in pediatric TB patients receiving guideline-recommended doses of first-line anti-tuberculosis drugs. The administered dose (mg/kg) significantly influences plasma drug concentrations, underscoring the necessity of TDM for first-line anti-tuberculosis drugs in children and individualized treatment plan adjustments to optimize TB management.</p> Clinical trial <p>Not applicable.</p>