Different plant extracts have been screened by thin-layer chromatography (TLC) bioautography in an effort to discover new acetylcholinesterase (AChE) inhibitors. The CH2 Cl-2 extract of Peucedanum ostruthium (L.) Koch roots exhibited significant inhibition of AChE-l activity. Active constituents were isolated by bioguided fractionation using almost exclusively centrifugal partition chromatography. Four coumarins (ostruthin, imperatorin, ostruthol, and oxypeucedanin hydrate) and a chromone derivative (peucenin) were found to inhibit AChE activity in this bioassay.