Ferulic acid dimer inhibits lipopolysaccharide-stimulated cyclooxygenase-2 expression in macrophages.

Phenylpropanoids may act as nonsteroidal anti- inflammatory drug (NSAID)-like compounds. 4-cis,8-cis- Bis(4-hydroxy-3- methoxyphenyl)-3 ,7-dioxabicyclo- (3.3.0)octane-2,6-dione (bis-FA, compound 2), a dimer of ferulic acid, was synthesized from ferulic acid (1), and its effect on lipopolysaccharide (LPS)-stimulated cyclooxygenase-2 (COX-2) expression in RAW 264.7 cells was compared with those of the parent ferulic acid (1) and of iso-ferulic acid (3- hydroxy-4-methoxycinnamic acid) (3). LPS-induced gene expression of COX-2 was markedly inhibited by compound 2 at a concentration of 10 IM and by compound 3 at 100 IM, but was not inhibited by compound 1 at 100 IM. This observation suggests that compound 2 may possess potent anti- inflammatory activity. These ferulic acid-related compounds were able to scavenge the stable 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical. The 50% inhibitory concentration for DPPH radicals declined in the order 3 (40.20 mM) > 2 (3.16 mM) > 1 (0.145 mM). Compound 1 possessed potent anti-radical activity, but no COX-2 inhibitory activity, which may be a result of enhancement of its conjugate properties by abstraction of an H atom from the phenolic OH group, causing loss of phenolic function. In contrast, inhibition of COX-2 expression by compounds 2 and 3 could be caused by their increased phenolic function, which is associated with decreased anti- radical activity. Compounds 2 and 3, particularly 2, may have potential as NSAID-like compounds. o-Methoxyphenols, such as ferulic acid (4-hydroxy-3- methoxycinnamic acid) and eugenol (4-allyl-2- methoxyphenol), act as antioxidants, but also act as prooxidants under certain circumstances, such as in light, oxygen-rich and alkaline conditions. The prooxidative activity of o-methoxyphenols can cause adverse effects such as allergic and inflammatory reactions (1). Dimerization of o-methoxyphenol monomers can reduce their intrinsic prooxidative properties, because the reactive moieties in the monomer are blocked by the other monomer (2). We have previously synthesized a symmetrical dimer of eugenol, bis-EUG (2,2'-dihydroxy- 3,3'-dimethoxy-5,5'-di(2-propenyl)-1,1'-biphenyl), by the ortho-ortho coupling reaction of eugenols, and an asymmetrical dimer of iso-eugenol (4-propenyl-2- methoxyphenol), namely dehydrodiisoeugenol (2-(3- methoxy-4-hydroxyphenyl)-3-methyl-5-(1-propenyl)-7- methoxy-2,3-dihydrobenzofuran), from iso-eugenol. These dimers showed inhibitory effects on lipopolysaccharide (LPS)-induced cyclooxygenase-2 (COX-2) expression in the mouse macrophage-like cell line RAW 264.7 (3, 4). Furthermore, ferulic acid derivatives, with a molecular structure in which the two ferulic acid moieties face each other, were reported to suppress COX-2 promoter activity in a concentration-dependent manner (5). These findings strongly suggest that dimers of ferulic acid would be able to inhibit COX-2 expression. With the aim of finding ortho-methoxyphenol derivatives with potent anti- inflammatory activity, a dimer of ferulic acid (bis-FA, 2) and evaluated for its effects on LPS-induced COX-2 expression in comparison with those of ferulic acid (1) and iso-ferulic acid (3). The activity of these compounds against DPPH radicals was also investigated, since COX-2 inhibitory activity has previously been reported to correlate with 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical- scavenging activity (6-8).