Twelve undescribed alkaloids, including eight pyrrolo[3,2-g]isoquinoline alkaloids (+)/(-)-anacyquinoline A (1a/1b), (±)-anacyquinoline B (2), (+)/(-)-anacyquinoline C (3a/3b), (±)-anacyquinoline D (4), (±)-anacyquinoline E (5), and (±)-anacyquinoline F (6), together with four pyrrolo[2,3-g]quinoline alkaloids (+)/(-)-anacyquinoline G (7a/7b), (±)-anacyquinoline H (8), and (±)-anacyquinoline I (9), were isolated from the root of Anacyclus pyrethrum (L.) DC. Their structures were determined via spectroscopic analyses (UV, IR, NMR), HRESIMS, quantum chemical calculations of ECD and NMR data, and single-crystal X-ray diffraction analysis (Cu Kalpha). In bioassay, (+)/(-)-anacyquinoline G (7a/7b), and (±)-anacyquinoline H (8) showed inhibition on NO production with IC50 values of 41.4, 44.1, and 31.4 muM, respectively.