The development of diabetes, especially type 2 diabetes, is influenced by the important role of postprandial blood glucose. One strategy for controlling postprandial hyperglycemia is inhibiting the digestion of dietary carbohydrates through inhibition of alpha-amylase and alpha-glucosidase enzymes. A natural alpha-amylase enzyme inhibition strategy that utilizes natural ingredients is an important alternative in the development of antidiabetic drugs. Peperomia pellucida herb is one of the plants which can be potentially be developed as an antidiabetic. This study aims to examine the effects of antidiabetic extract and ethyl acetate fraction of Peperomia pellucida in inhibiting the alpha-amylase enzyme. The research was conducted by determining the total flavonoid content using the quercetin standard. Antidiabetic studies were carried out by reducing sugar produced from the hydrolysis starch determined by colorimetric assay with dinitrosalicylic reagent. The activity of extract and fraction of Peperomia pellucida in inhibiting alpha-amylase enzyme were analyzed using UV-Vis spectrophotometer instrument. Total flavonoid content was calculated by entering the absorbance value into the standard quercetin curve, while the IC50 value of in vitro antidiabetic activity was determined by entering the value 50 on the curve between the sample concentration and % inhibition of the alpha-amylase enzyme. The results showed that the total flavonoid content of the ethanol extract and the ethyl acetate fraction of Peperomia pellucida were 88.24±3.07 mg QE/g extract and 80.45±2.81 mg QE/g fraction, respectively. Based on the results, the inhibition of the alpha-amylase enzyme showed that the ethanol extract and the ethyl acetate fraction of Peperomia pellucida had an inhibitory activity with the IC50 value of the ethanol extract 1066.20 μg/mL and the ethyl acetate fraction 907.19 μg/mL. Meanwhile, the control of acarbose showed an IC50 value of 499.96 μg/mL. Ethanol extract and ethyl acetate fraction of Peperomia pellucida herbs have an antidiabetic activity with the mechanism of inhibiting the activity of alpha-amylase enzyme, but the activity is lower than acarbose.