Myocardial protection during cardiopulmonary bypass (CPB) can be achieved using cardioplegic solutions. Although, acute kidney injury (AKI) is a common complication following CPB, the effects of cardioplegic solutions on AKI have rarely been investigated. Within this study, the effects of the cardioplegic solutions histidine‐tryptophan‐ketoglutarate (HTK; Custodiol) and HTK‐N (Custodiol‐N) on AKI in a large animal model were compared. Therefore, Landrace pigs underwent median sternotomy, CPB at 34°C, 90 minutes of cardiac arrest and 120 minutes of reperfusion. Animals were randomized for single‐shot cardioplegia with either HTK (n = 10) or HTK‐N (n = 10). Renal biopsies and sera were analyzed to determine AKI biomarkers and apoptosis. Compared to HTK, HTK‐N induced a decreased extent of proximal tubule swelling (48.3 ± 1.6 µm vs 52.3 ± 1.1 µm, P = .05) and decreased cytochrome c release (0.26 ± 0.04 vs 0.46 ± 0.08, P = .04) without reaching statistical significance due to Bonferroni correction. Comparing baseline and postreperfusion levels, the hemoglobin (Hb) and blood calcium levels were lower in HTK‐N (Hb_baseline: 6.0 ± 0.6 mmol/L, Hb_reperfusion: 6.2 ± 0.7 mmol/L, P = .12; Ca²⁺_baseline: 1.36 ± 0.05 mmol/L, Ca²⁺_reperfusion: 1.28 ± 0.05 mmol/L, P = .16) compared to the HTK group (Hb_baseline: 5.9 ± 0.4 mmol/L, Hb_reperfusion: 4.7 ± 0.8 mmol/L, P < .01; Ca²⁺_baseline: 1.34 ± 0.07 mmol/L, Ca²⁺_reperfusion: 1.24 ± 0.06 mmol/L, P < .01). The present study showed that HTK‐N could positively affect the kidney during CPB. Hb and calcium levels were stabilized. A statistical trend was found showing that AKI‐related proximal tubule swelling and cytochrome c release were diminished.