Although many cross-section studies have assessed the determinants of glycated hemoglobin (HbA1c), there have been limited studies designed to evaluate the temporal correlates of HbA1c in non-diabetic patients. This study aimed to identify the major determinants of longitudinal change of HbA1c in non-diabetic patients. This study included subjects from the 104,451 participants enrolled between 2012 and 2018 in the Taiwan Biobank. We only included participants with complete data at baseline and follow-up (n = 27,209). Patients with diabetes at baseline or follow-up (n = 3983) were excluded. Finally, 23,226 participants without diabetes at baseline and follow-up were selected in this study. △Parameters was defined as the difference between the measurement baseline and follow-up. Multivariable linear regression analysis was used to identify the major determinants of HbA1c longitudinal change (△HbA1c). During a mean 3.8 year follow-up, after multivariable analysis, new-onset hypertension (coefficient β: 0.014, p < 0.001), high △heart rate (coefficient β: 0.020, p = 0.002), high △BMI (coefficient β: 0.171, p = 0.028), high △fasting glucose (coefficient β: 0.107, p < 0.001), low △creatinine (coefficient β: −0.042, p < 0.001), high △total cholesterol (coefficient β: 0.040, p < 0.001), high △hemoglobin (coefficient β: 0.062, p < 0.001), high △GPT (coefficient β: 0.041, p = 0.001), and low △albumin (coefficient β: −0.070, p < 0.001) were significantly associated with high △HbA1c. In non-diabetic population, strategies to decrease the development of new-onset hypertension, resting heart rate, body mass index, fasting glucose, total cholesterol, and GPT and increase serum albumin level might be helpful in slowing the longitudinal change of HbA1c. In addition, increased hemoglobin and decreased serum creatinine over time also had an impact on the HbA1c elevation over time in non-diabetic population.