The burden of hypoglycemia is high in resource limited countries, such as Ethiopia. However, there are no sufficient studies conducted in Ethiopia in general and in the study setting in particular in the previous era. Hence, this study aims to assess the proportion of hypoglycemia and associated factors among preterm neonates admitted to the neonatal intensive care unit at Debre Tabor Comprehensive Specialized Hospital, Ethiopia, in 2021. A hospital-based cross-sectional study was conducted from October 1 to December 30, 2021, at Debre Tabor Comprehensive Specialized Hospital in the neonatal intensive care unit ward. The data was entered in Epi-info 7 and exported to STATA version 14. A binary and multivariable logistic regression was computed at 95% confidence interval (CI). During bivariable analysis, variables having a p-value of less than 0.25 were chosen for multivariable logistic regression analysis, and variables having a p-value of less than 0.05 in multivariable analysis, were significant associations with the dependent variable. The study included 267 preterm neonates, and 23.59% (95% CI 18.9–29.1) were develop hypoglycemia. Moreover, 49 (18.35%) preterm neonates died during the study period. In this study, preterm neonates with hypothermia [Adjusted Odds Ratio (AOR = 4.5; 95 CI 3.4, 7.2)], birth asphyxia (AOR = 5.1; 95 CI 3.9, 27.1), seizure (AOR = 4.7; 95 CI 2.8, 17.8), and also preterm neonates born from diabetic mothers (AOR = 6.7; 95 CI 3.3, 27.2) were significantly associated with the occurrence of hypoglycemia in the neonatal intensive care. The proportion of hypoglycemia and associated factors among preterm neonates admitted to the neonatal intensive care unit at Debre Tabor Comprehensive Specialized Hospital was found to be high. The associated factors for the occurrence of hypoglycemia were discovered to be neonates with hypothermia, birth asphyxia, seizure, and neonates born with a diabetes mother. Thus, recognizing and treating the above associated factors is essential to preventing, and controlling hypoglycemia.