Objective To investigate the early diagnostic value of combined detection of serum TIR?domain?containing adapter?inducing interferon?β （TRIF）， cyclophilin A （CyPA）， ubiquitin carboxyl?terminal hydrolase L1 （UCH?L1）， and hepcidin （Hepc） in brain injury among premature infants （BIPI）. Methods Clinical data from 105 infants with brain injury following perinatal asphyxia （BIPI） admitted to our hospital between January 2022 and December 2024 were retrospectively collected and assigned to the case group， which was further subdivided into a severe subgroup （n = 31） and a mild subgroup （n = 74） based on the extent of brain injury. As controls， clinical data from 105 healthy preterm infants born in our hospital during the same period were retrospectively enrolled in a 1∶1 ratio. Serum levels of TRIF， CyPA， UCH?L1， and Hepc， along with clinical characteristics， were compared between the groups. Pearson correlation analysis was performed to evaluate the association between serum biomarker levels and neonatal behavioral neurological assessment （NBNA） scores. The diagnostic performance of these biomarkers for early detection of BIPI was assessed by constructing receiver operating characteristic （ROC） curves and calculating the area under the curve （AUC）. Results Serum levels of TRIF， CyPA， UCH?L1， and Hepc were significantly higher in the case group than in the control group （P < 0.05）， while NBNA scores were significantly lower in the case group （P < 0.05）. Similarly， serum levels of these biomarkers were significantly elevated in the severe group compared to the mild group （P < 0.05）， accompanied by significantly lower NBNA scores in the severe group （P < 0.05）. Pearson correlation analysis revealed that serum levels of TRIF， CyPA， UCH?L1， and Hepc were negatively correlated with NBNA scores （r = -0.579， -0.514， -0.609， -0.588； all P < 0.05）. ROC analysis demonstrated that the combined detection of these four markers yielded an AUC of 0.927 for early diagnosis of BIPI， which was significantly higher than the AUCs of individual markers （0.819， 0.803， 0.776， and 0.767， respectively； P < 0.05）. Conclusions The serum levels of TRIF， CyPA， UCH?L1， and Hepc in children with BIPI were significantly elevated. These biomarkers were closely associated with disease progression and neurological development， and their combined measurement demonstrated superior performance in the early diagnosis of BIPI.