Few data on azathioprine (AZA) therapy for inflammatory bowel disease (IBD) exist for children. We evaluated whether the 6-thioguanine nucleotides (6-TGN) level predicts AZA refractoriness in children with IBD and whether children benefit an AZA dose escalation. Seventy-eight children with IBD initially treated with an AZA dose of 1.5–2.5 mg/kg/day were retrospectively included. The dose was adjusted based on the clinical status. The receiver operating characteristic curve and logistic regression were used to determine predictors for AZA resistance. Initially, 18 of 40 (45%) patients receiving a dose of <2 mg/kg/day and 11 of 38 (28.9%) patients receiving a dose of 2–2.5 mg/kg/day achieved remission. The 6-TGN level above 250 pmol/8.108 RBCs was associated with a higher remission rate, though non-significant. Among 35 patients with a dose escalation due to treatment failure, 12 (34.3%) achieved remission (the median 6-TGN level increased from 260 to 394 pmol/8.108 RBCs [P = .002]), 23 (67.6%) were AZA refractory. A 6-TGN level above 405 pmol/8.108 RBCs was the only predictor for AZA resistance (sensitivity 78.3%, specificity 75%, OR 10.8 [95% CI: 2.1–55.7, P = .004]). Serial metabolite monitoring is useful to identify children with IBD resistant to AZA. Children who cannot achieve remission despite a 6-TGN level above 405 pmol/8.108 RBCs should receive alternative therapies than dose increase.