High-motility group box protein 1 (HMGB1) has an important role in autophagy; however, its exact role in acute necrotizing pancreatitis (ANP) remains unknown. The present study aimed to investigate the expression pattern of HMGB1 in ANP, and to determine its association with autophagy. Sprague Dawley rats (weight, 350±30 g, n=48) were randomly divided into control (n=12) and experimental (n=36) groups. Experimental rats were retrogradely injected with 5% sodium taurocholate into the biliopancreatic duct to induce ANP. Control rats received an equal amount of saline. Serum amylase levels were used to determine whether the model had been successfully generated. Autophagosomes in pancreatic acinar cells were observed under electron microscopy. The expression levels of HMGB1 and Beclin 1 were detected in pancreatic tissues by western blotting, quantitative polymerase chain reaction and immunohistochemistry. HMGB1 levels were also determined in the serum and in isolated nuclei. The results demonstrated that autophagy was detected at 3 h post‑ANP induction; however, HMGB1 expression remained unaltered during the early stage (0‑6 h; P>0.05). HMGB1 expression was significantly increased at 12 h, and was still increasing at 24 h (P<0.05). Notably, HMGB1 was increased in the nuclei compared with in the cytoplasm at 3‑6 h. Furthermore, serum HMGB1 levels began to increase at 3 h, and reached the highest levels at 24 h in the ANP group. In conclusion, in an ANP model, HMGB1 was initially increased in the nuclei to initiate autophagy. Subsequently, it moved into the cytoplasm, where it interacted with Beclin 1 to enhance autophagy, and HMGB1 was released into the blood, leading to the deterioration of ANP.