A combination of flash chromatography, solid phase extraction, high-performance liquid chromatography, and in vitro bioassays was used to isolate phytocomponents endowed with anticholinesterase activity in extract from Phyllanthus muellarianus. Phytocomponents responsible for the anti-cholinesterase activity of subfractions PMF1 and PMF4 were identified and re-assayed to confirm their activity. Magnoflorine was identified as an active phytocomponent from PMF1 while nitidine was isolated from PMF4. Magnoflorine was shown to be a selective inhibitor of human butyrylcholinesterase-hBChE (IC50 = 131 +/- 9 mu M and IC50 = 1120 +/- 83 mu M, for hBuChE and human acetylcholinesterase-hAChE, respectively), while nitidine showed comparable inhibitory potencies against both enzymes (IC50 = 6.68 +/- 0.13 mu M and IC50 = 5.31 +/- 0.50 mu M, for hBChE and hAChE, respectively). When compared with the commercial anti-Alzheimer drug galanthamine, nitidine was as potent as galanthamine against hAChE and one order of magnitude more potent against hBuChE. Furthermore, nitidine also showed significant, although weak, antiaggregating activity towards amyloid-beta self-aggregation.