Abstract Background Acute kidney injury (AKI) subtypes combining kidney functional parameters and injury biomarkers may have prognostic value. We aimed to determine whether neutrophil gelatinase-associated lipocalin (NGAL)/hepcidin-25 ratio (urinary concentrations of NGAL divided by that of hepcidin-25) defined subtypes are of prognostic relevance in cardiac surgery patients. Methods We studied 198 higher-risk cardiac surgery patients. We allocated patients to four groups: Kidney Disease Improving Global Outcomes (KDIGO)-AKI-negative and NGAL/hepcidin-25 ratio-negative (no AKI), KDIGO AKI-negative and NGAL/hepcidin-25 ratio-positive (subclinical AKI), KDIGO AKI-positive and NGAL/hepcidin-25 ratio-negative (clinical AKI), KDIGO AKI-positive and NGAL/hepcidin-25 ratio-positive (combined AKI). Outcomes included in-hospital mortality (primary) and long-term mortality (secondary). Results We identified 127 (61.6%) patients with no AKI, 13 (6.6%) with subclinical, 40 (20.2%) with clinical and 18 (9.1%) with combined AKI. Subclinical AKI patients had a 23-fold greater in-hospital mortality than no AKI patients. For combined AKI vs. no AKI or clinical AKI, findings were stronger (odds ratios (ORs): 126 and 39, respectively). After adjusting for EuroScore, volume of intraoperative packed red blood cells, and aortic cross-clamp time, subclinical and combined AKI remained associated with greater in-hospital mortality than no AKI and clinical AKI (adjusted ORs: 28.118, 95% CI 1.465–539.703; 3.737, 95% CI 1.746–7.998). Cox proportional hazard models found a significant association of biomarker-informed AKI subtypes with long-term survival compared with no AKI (adjusted ORs: pooled subclinical and clinical AKI: 1.885, 95% CI 1.003–3.542; combined AKI: 1.792, 95% CI 1.367–2.350). Conclusions In the presence or absence of KDIGO clinical criteria for AKI, the urinary NGAL/hepcidin-25-ratio appears to detect prognostically relevant AKI subtypes. Trial registration number NCT00672334, clinicaltrials.gov, date of registration: 6th May 2008, https://clinicaltrials.gov/ct2/show/NCT00672334. Graphic abstract Definition of AKI subtypes: subclinical AKI (KDIGO negative AND Ratio-positive), clinical AKI (KDIGO positive AND Ratio-negative) and combined AKI (KDIGO positive AND Ratio-positive) with urinary NGAL/hepcidin-25 ratio-positive cut-off at 85% specificity for in-hospital death. AKI, acute kidney injury. AUC, area under the curve. NGAL, neutrophil gelatinase-associated lipocalin. KDIGO, Kidney Disease Improving Global Outcomes Initiative AKI definition.