BACKGROUND: The effects of sclerotic bone on anti-tuberculosis (anti-TB) drug distribution in the blood and in spinal tuberculosis (STB) lesions were investigated. METHODS: Fifty-six patients with STB were prospectively enrolled from January 2020 to March 2023 and were divided into 2 groups: a group with sclerotic bone and a group without sclerotic bone, as identified on preoperative computed tomography (CT) scans. Individuals in the sclerotic bone group were further divided into fragmentary and non-fragmentary sclerotic bone groups. The patients underwent surgery, and blood was collected along with normal vertebral and STB-lesion-containing bone tissue samples. Following treatment, the samples were processed by a pharmacological laboratory in order to detect the concentrations of anti-TB drugs, including pyrazinamide, rifampicin, isoniazid, and ethambutol. RESULTS: Twenty-seven East Asian female and 29 East Asian male patients with STB were included in this study. The levels of anti-TB drugs showed a progressive decrease with increased circulatory distance, from blood to normal vertebral tissue to TB lesions, across all patient groups. Drug concentrations in TB lesions in the sclerotic bone group were significantly lower than those in the non-sclerotic bone group, as were concentrations in TB lesions in the non-fragmentary sclerotic bone group relative to those in the fragmentary sclerotic bone group. Drug levels in the blood and in normal vertebral bone tissue did not significantly differ between the sclerotic and non-sclerotic groups, nor between the fragmentary and non-fragmentary groups. Drug levels in the blood were linearly correlated with those in TB lesions in both the non-sclerotic bone group and the fragmentary sclerotic bone group. CONCLUSIONS: These results indicate that sclerotic bone negatively affects the dissemination of anti-TB drugs, with non-fragmentary sclerotic bone posing a greater obstacle than fragmentary sclerotic bone. In patients with STB without sclerotic bone or with fragmentary sclerotic bone, anti-TB drug levels in the blood were linearly correlated with drug levels in STB lesions. LEVEL OF EVIDENCE: Prognostic Level II. See Instructions for Authors for a complete description of levels of evidence.