For figuring out the relationships between chemical structure and extraction properties of the soft non-hetero-cyclic N-donor ligand nitrilotriacetamide (NTAamide), eight representative ligands with different substituent groups based on NTAamide were synthesized on purpose. Their extraction behaviors toward Am3+ and Eu3+ were investigated. In the examined HNO3 concentration ranging from 0.01 to 0.50 mol/L, the distribution ratios (D) of Am3+ and Eu3+ increased with increasing the extractant concentration or decreasing the aqueous phase acidity. The highest separation factor (SFAm/E) of 12 with D-Am = 5.8 and D-Eu = 0.49 could be obtained in the case of the extraction by 0.10 mol/L HNO3 N,N',N",N"-hexadecylnitrilotriacetamide (NTAamide(n-Dec)) in MIBK as diluent from 0.10 mol/L HNO3 solution. The extractability of eight ligands decreased in the order of NTAamide(n-Dodec) > NTAamide(n-Dec) > NTAamide(n-Oct) > NTAamide(n-Bu) > NTAamide(iBu) > NTAamide(Et-p-MePh) > NTAamide(Et-Ph) > NTAamide(2-EtHex). The aliphatic substituent groups with longer straight chain are advantageous to the extraction, while the aromatic ones with larger steric hindrance and electron-withdrawing property are disadvantageous. The log beta values between NTAamides and Eu3+ were obtained by fluorescence titration for the first time. Moreover, HRMS indicated the formation of 1:1 complexes of NTAamides with Eu3+, which was in good accordance with the extraction results obtained from slope analysis.