Intestinal damage (ID) leads to bacterial translocation and bloodstream infections-the common cause of non-relapse mortality in childhood acute lymphoblastic leukemia (ALL). This study evaluated ID over ALL induction and the significance of body mass index (BMI) for its development and identified biomarkers reflecting chemotherapy-induced ID. The composite biomarker panel included 37 plasma amino acids, urea, ammonia, fecal calprotectin (fCLP), absolute neutrophil count (ANC), C-reactive protein, and albumin. We prospectively assessed 45 children treated according to the ALLTogether protocol in 2020-2024. Analysis and sample collection were performed on days 1, 8, 15, 22, and 29 of the protocol. The obtained values were compared between the ID and non-ID groups. 40% of patients (18/45) had grade I-III ID which was more pronounced on day 22 of induction when the ANC increased from its lowest point. Age younger than 5.5 years at a diagnosis was a significant prognostic factor for ID. Decreasing BMI and concentrations of citrulline, taurine, cystine, phosphoethanolamine, A-aminobutyric acid, B-alanine, and albumin suggest progressive ID in children treated due to ALL. No difference in ANC and fCLP was found between patients with and without ID, but fCLP levels start to rise simultaneously as the most intense ID is observed. In conclusion, assessing nutritional status and prospective evaluation of biomarkers may provide valuable information on treatment-related ID.