A series of 2,5-bis(alkyl/arylamino)-1,4-benzoquinones (1-12) were investigated in vitro for their potential to inhibit the activity of jack bean urease. Compounds 1-6, 8, 9, 11 and 12 effectively inhibited the jack bean urease activity by 90.8 % when tested at 5 μm, whereas 7 and 10 had relatively little effect. The IC50 for most compounds was in the nanomolar range (31.4 nm and 36.0 nm for 2 and 8, respectively). The mechanism of enzyme inhibition shown by 2 and 8 is typical of mixed-type inhibitors, whose affinity for the active site is over 6- and 2-fold higher (Ki =30.0 and 22.8 nm, for 2 and 8, respectively) than that of an allosteric site. Molecular docking studies revealed that both 2 and 8 establish hydrogen bonds with the amino acids residues Asp494, Met588, His593 and Ala636 in the active site of jack bean urease. These results indicate that such aminoquinones are useful leads for the development of more efficient urease inhibitors of wider utility.