This study was conducted to evaluate the cytotoxicity and α-amylase, α-glucosidase, pancreatic lipase inhibition in vitro by different solvent fractions from the roots of Codonopsis lanceolata. The values of IC50 against Calu-6 cell showed a high effect in n-hexane fraction (10.13 μg/mL) whereas DW fraction exhibited the weakest inhibition on cell viability, having an IC50 value of over 1,000 μg/mL. The values of IC50 against HCT-116 cell showed the highest activity in n-BuOH fraction (102.01 μg/mL), followed by n-hexane fraction (145.85 μg/mL), methylene chloride fraction (332.02 μg/mL), ethyl acetate fraction (462.93 μg/mL) and DW fracion (>1,000 μg/mL). α-Amylase inhibitory activity in methylene chloride fraction and ethyl acetate fraction was found to have a higher inhibitory effect with 24.5% and 25.6% than the other fractions. The highest α-glucosidase inhibitory activity was observed from the ethyl acetate fraction extract, while the extract of DW fraction showed the lowest level of inhibitory activity at given experiment concentration. The pancreatic lipase inhibitory activity of C. lanceolata was found to have a higher the effect in ethyl acetate fraction. Inhibition of lipase activity of the ethyl acetate fraction and n-hexane fraction showed a relatively high, while the extract of DW fraction showed the lowest level at given experiment concentration. These results suggested that the roots of C. lanceolata may assist in the potential biological activity on carbohydrate, lipid Inhibitory activity and anticancer activity.