Abstract Background Sex hormones are strongly linked to the occurrence and development of diabetes, and influence glycated hemoglobin (HbA_1c) levels in diabetic population; but, the relationship between sex hormones and HbA_1c in non-diabetic population remains unknown. This study aimed to explore the extent of influence of sex hormones on HbA_1c levels in non-diabetic population. Methods A total of 1409 non–diabetic subjects, including 601 men and 808 postmenopausal women were recruited from Shanghai community. HbA_1c was detected using high performance liquid chromatography, and hemoglobin level was determined by sodium lauryl sulfate colorimetry. Serum estradiol (E₂), total testosterone (TT), and sex hormone binding globulin (SHBG) were measured by chemiluminescent microparticle immunoassays. Results The level of HbA_1c was 5.6 (5.4–5.9) % in all subjects, with 5.6 (5.4–5.8) % in men and 5.7 (5.5–5.9) % in postmenopausal women. After adjusting for age, body mass index (BMI), and hemoglobin, E₂ was positively correlated with HbA_1c in men (r = 0.122, P = .003), and SHBG was inversely correlated with HbA_1c (r = − 0.125, P < .001) in women. Other hormones were not correlated with HbA_1c (all P > .05). Multivariate linear regression analysis showed that, except for traditional factors, such as age, hemoglobin, and BMI, E₂ was another determinant of HbA_1c (standardized β = 0.137, P = .003) in men; besides, in women, SHBG was another determinant of HbA_1c (standardized β = − 0.178, P < .001), except for age and systolic blood pressure. Conclusion After controlling for confounding factors, two sex hormones, as E₂ and SHBG could influence HbA_1c levels in non-diabetic population.