In this work, the specificity of ubiquinol as inhibitor of the neutral sphingomyelinases present at the plasma membrane (Mg2+-dependent and -independent) and structural requirements for such inhibition have been studied. Our results have shown that ubiquinol specifically inhibits Mg2+-dependent neutral sphingomyelinase activity in isolated liver plasma membranes, but no significant participation of the Mg2+-independent enzyme was observed. Both the reduction state of the (hydro)quinone ring and the length of the hydrophobic side chain were important determinants in neutral sphingomyelinase inhibition. Ubiquinols inhibited the nSMase more efficiently than ubiquinones, and hydrophobic homologs with six or nine isoprene units were the most effective inhibitors. Inhibition of nSMase by ubiquinols displayed similarities with inhibition by manumycin and the hydroquinones F11334's, suggesting that these compounds could act as structural analogs of ubiquinol. Beyond its participation in mitochondrial energy metabolism, and as antioxidant, this novel role for ubiquinol as a neutral sphingomyelinase inhibitor should be considered an important factor to regulate lipid signaling at the plasma membrane that could be related to its beneficial effects on cells, tissues, and organisms. (C) 2002 Elsevier Science (USA). All rights reserved.