Background: Antidiabetic treatment with traditional medicine simultaneously can change the pharmacokinetic (PK) parameters, effectiveness, and toxicity of the drug. The combination of glibenclamide and Moringa oleifera Lam. ethanolic leaf extract can affect the activity of lowering blood glucose in diabetic rats. Methods: Rats were induced with alloxan at a dose of 140 mg/kgBW. There were 5 groups, namely the normal group, negative control (Na-CMC 0.5%), group 1 (gibenclamide 5 mg/kgBW), group 2 (Moringa ethanolic leaf extract 300 mg/kgBW), and group 3 (combination of Moringa ethanolic leaf extract 300 mg/kgBW and glibenclamide 5 mg/kgBW). Blood sugar levels were examined using the Easy Touch GCU on days 0, 3, 7, 14 and 21. The glibenclamide bioanalysis method was carried out using RP-HPLC. Measurement of glibenclamide drug concentration was carried out at 3 hours (Tmax) after administration of the preparation on days 3, 7, 14 and 21 and pharmacokinetic parameters of glibenclamide were measured on day 22 through the relationship between concentration and time of plasma sampling using PK Solver. Results: The group of glibenclamide with moringa leaf ethanol extract did not have a significant effect compared to the treatment of glibenclamide administration or moringa leaf ethanol extract. The combination of glibenclamide and moringa leaf ethanol extract increased the concentration of glibenclamide in diabetic rats significantly in the T-test (p <0.0001) by changing the pharmacokinetic parameters Cmax and AUC0–t, and T1/2. Conclusion: Administration of moringa leaf ethanol extract with glibenclamide can increase the concentration of glibenclamide in plasma which affects the pharmacokinetic parameters, but these changes do not affect the activity of lowering blood glucose in diabetic rats.