From the methanolic extract of the rhizome of Curcuma zedoaria, we isolated anti-inflammatory sesquiterpene furanodiene (1) and furanodienone (2) along with new sesquiterpene compound 3 and known eight sesquiterpenes, zederone (4), curzerenone (5), curzeone (6), germacrone (7), 13-hydroxygermacrone (8), dehydrocurdione (9), curcumenone (10), and zedoaronediol (11). Their structures were elucidated on the basis of spectroscopic data. The anti-inflammatory effect of isolated components on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammation of mouse ears were examined. Compounds 1 and 2 suppressed TPA-induced inflammation of mouse ears by 75% and 53%, respectively, at a dose of 1.0 µmol. Their activities are comparable to that of indomethacin, the normally used anti-inflammatory agent.Keywords: anti-inflammatory; Curcuma zadoaria; 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced edema Zedoary (Curcuma zedoaria Roscoe, Zingiberaceae) is cultivated as a vegetable, spice, and perfumery material in Southeast Asian countries. The rhizome of this plant has been used as a stimulant, stomachic, carminative, diuretic, anti-diarrheal, anti-emetic, anti-pyretic, and depurator, and also as an ointment for ulcers, wounds, and other skin disorders. 1) Recently, Yoshikawa et al have revealed that chemical constituent of C. zedoaria showed vasorelaxant, heptatoprotective, and inhibitory activity of NO production. 2)-4) The zedoary is used to the treatment of the skin disease, however the study of the anti-inflammatory effect of chemical constituents from C. zedoaria has not been studied so far except that of dehydrocurdione (9) by Yoshioka et al. 5) Therefore, we tried to isolate sesquiterpens from zedoary and subject on anti-inflammatory test. For isolating sesquiterpens like dehydrocurudione, we used the reddish spot on thin layer chromatography with spraying of anisaldehyde/sulfuric acid as index.In the present paper, we describe the structure of isolated eleven sesquiterpenes from C. zedoaria and their anti-inflammatory effect on the 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced edema of mouse ears.The zedoary rhizome (cultivated in Okinawa, Japan) was soaked in methanol, and the resulting methanol extract was concentrated in vacuo.The concentrate was successively partitioned between benzene and water, and then ethyl acetate and water.The benzene-soluble part showed anti-inflammatory activity (IE of 34% at a dose of 2 mg). The benzene-soluble part was chromatographed on silica gel to give 20% ethyl

We have synthesized ∆lac-acetogenins that are new acetogenin mimics possessing two n-alkyl tails without an R, -unsaturated γ-lactone ring and suggested that their inhibition mechanism may be different from that of common acetogenins [Hamada et al. (2004) Biochemistry 43, 3651-3658]. To elucidate the inhibition mechanism of ∆lac-acetogenins in more detail, we carried out wide structural modifications of original ∆lac-acetogenins and characterized the inhibitory action with bovine heart mitochondrial complex I. In contrast to common acetogenins, both the presence of adjacent bis-THF rings and the stereochemistry around the hydroxylated bis-THF rings are important structural factors required for potent inhibition. The inhibitory potency of a derivative possessing an n-butylphenyl ether structure (compound 7) appeared to be superior to that of the original ∆lac-acetogenins and equivalent to that of bullatacin, one of the most potent natural acetogenins. Double-inhibitor titration of steady-state complex I activity showed that the extent of inhibition of compound 7 and bullatacin is not additive, suggesting that the binding sites of the two inhibitors are not identical. Competition tests using a fluorescent ligand indicated that the binding site of compound 7 does not overlap with that of other complex I inhibitors. The effects of compound 7 on superoxide production from complex I are also different from those of other complex I inhibitors. Our results clearly demonstrate that ∆lac-acetogenins are a novel type of inhibitor acting at the terminal electron-transfer step of bovine complex I.

Studies on the inhibitory mechanism of acetogenins, the most potent inhibitors of mitochondrial complex I (NADH-ubiquinone oxidoreductase), are useful for elucidating the structural and functional features of the terminal electron transfer step of this enzyme. Previous studies of the structure-activity relationship revealed that except for the alkyl spacer linking the two toxophores (i.e., the hydroxylated THF and the gamma-lactone rings), none of the multiple functional groups of these inhibitors is essential for potent inhibition. To elucidate the function of the alkyl spacer, two sets of systematically selected analogues were synthesized. First, the length of the spacer was varied widely. Second, the local flexibility of the spacer was specifically reduced by introducing multiple bond(s) into different regions of the spacer. The optimal length of the spacer for inhibition was approximately 13 carbon atoms. The decrease in the strength of the inhibitory effect caused by elongating the spacer from 13 carbons was much more drastic than that caused by shortening. Local flexibility in a specific region of the spacer was not important for the inhibition. These observations indicate that the active conformation of the spacer is not an extended form, and is not necessarily restricted to a certain rigid shape. Moreover, an analogue in which a spacer covering 10 carbon atoms was hardened into a rodlike shape still maintained a potent inhibitory effect. Our results strongly suggest that the spacer portion is free from steric congestion arising from the putative binding site probably because there is no cavity-like binding site for the spacer portion. The manner of acetogenin binding to the enzyme may not be explained by a simple "key and keyhole" analogy.