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Myristicaceae known as one of the plants that widely used as traditional medicine. Utilization of Horsfieldia spicata (Roxb.) J. Sinclair, one of the Myristicaceae family member as a sources of antidiabetic active compounds are still needed. The antidiabetic activity of methanol extract (ME) and its fractions [hexane fraction (HF), ethyl acetate fraction (EF), butanol fraction (BF) and residue fraction (RF)] of H. Spicata stem bark were in vitro tested by measuring of screening of phytochemical contents, total phenolic and flavonoid compounds, and α-glucosidase enzyme inhibitory activity. Phytochemical screening result showed that (ME) indicated the presence of flavonoid and saponin compounds. Due to the total phenolic and flavonoid compounds measurement result, indicated that ME has a highest value for total phenolic compounds (14.709 ± 0.23 µg/mL) and EF for total flavonoid compounds (55.825 ± 2.21 µg/mL). Antidiabetic activity of the ME, HF, EF, BF, and RF against α-glucosidase enzyme showed that ME has antidiabetic activity with IC50 value 49.30 µg/mL, which is it was higher than HF, BF and RF, but less active compared to EF and quercetin (positive control) with the IC50 value are 159.64, 50.60, 94.01, 22.73, 2.99 µg/mL, respectively. The result showed that (EF) of methanolic extract of H. Spicata has inhibitory activity against α-glucosidase enzyme, and potential to be develop as antidiabetic active fraction. [ABSTRACT FROM AUTHOR]