Objective To investigate the correlation between glycosylated hemoglobin A1c (HbA1c) and reactive hyperemia index (RHI) in the patients with type 2 diabetes mellitus (T2DM). Methods A total of 608 T2DM patients admitted to the Endocrinology Department of First Affiliated Hospital of Nanjing Medical University from January 2017 to February 2019 were enrolled in this study. During the hospitalization, all patients were evaluated by using Endo-PAT non-invasive vascular endothelial function detection technology to calculate RHI. The patients with RHI≥1.67 were assigned into the control group, and those with RHI＜1.67 were into the study group. According to the HbA1c level, they were divided into HbA1c <7% group, HbA1c 7%-9% group and HbA1c >9% group. Their general data, biochemical indices, blood glucose level and insulin resistance index were compared among the groups. Spearman correlation analysis was used to evaluate the correlation between HbA1c and RHI. The relationship was evaluated by multiple linear regression analysis on the factors affecting endothelial function. Results HbA1c level was significantly higher in the study group than in the control group (P=0.001), but no statistical significances were seen in age, body mass index, blood lipid, blood glucose, and homeostatic model assessment of insulin resistance (HOMA-IR) and of β-cell function (HOMA-β) between two groups. With the increase of HbA1c, RHI was decreased in the groups of different HbA1c level (P=0.004). Spearman correlation analysis showed that HbA1c level was negatively correlated with RHI in T2DM patients, and multivariate regression and logistic regression analysis indicated that HbA1c was an independent and significant predictor for vascular endothelial function assessment index (OR=0.864,95%CI 0.786-0.950, P=0.002). Conclusion For T2DM patients, HbA1c can be used to evaluate the recent endothelial function. Actively controlling the blood glucose can improve vascular endothelial dysfunction, and thus the damage to peripheral vessels and other target organs will be thereby reduced and cardiovascular events be prevented.