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Certain plant species belonging to the family Annonaceae produce Annonaceous acetogenins, which are a unique class of long-chain fatty acid derivatives with potent cytotoxicity. Putative protein targets of the acetogenins are membrane-associated proteins, including complex I. Asimicin and its analogues constitute a class of Annonaceous acetogenins containing two tetrahydrofuran (THF) rings with hydrocarbon chains tethered to each ring; an alpha,beta-unsaturated gamma-lactone ring is terminal to one of the alkyl chains. The compounds examined in this study differ in the length of the alkyl chain between the THF rings and the lactone ring. The positions of both the THF and the lactone rings within liposomal membranes were determined by proton (1H) nuclear magnetic resonance spectroscopy. The depth of membrane penetration of acetogenins, coupled to membrane diffusion, controls the conformation of acetogenins as they diffuse to an active site. Based on 1H intermolecular nuclear Overhauser effects (NOEs), the THF rings of all acetogenins studied reside near the polar interfacial head group region of the DMPC. This was corroborated by 1H two-dimensional NOE spectroscopy and differential scanning calorimetry studies. The 1H difference NOE spectra indicated that the lactone rings of asimicin and parviflorin, the latter of which has two fewer carbons in its alkyl chain, are located below the glycerol backbone in the membrane. In contrast with asimicin and parviflorin, the lactone ring of longimicin B, an asimicin analogue with an alkyl chain four carbons shorter, resides close to the midplane in the membrane. This was corroborated by manganese-induced broadening studies. Since the THF rings are located near the center of the acetogenin molecules and the lactone ring is terminal to a long alkyl chain, these observations indicate that an asimicin-type acetogenin can be in either sickle-shaped or U-shaped conformations, depending on the length of the alkyl chain between the THF rings and the lactone ring. Interestingly, longimicin B does not exhibit significant cytotoxicity, but parviflorin is as cytotoxic as asimicin. The cytotoxicity of the asimicin-type of acetogenins would seem to be strongly related to the membrane conformation. This is the first report elucidating the conformation of Annonaceous acetogenins in membranes.

Two new sesquiterpenes, 10 alpha-hydroxyamorphan-4-en-3-one (1) and 4 alpha-methylcadinane-4 alpha-methyl-1 alpha,2 alpha,10 alpha-triol (2), together with four known compounds, sesquiterpenes 10 alpha-hydroxycadinan-4-en-3-one (3) and alpha-cadinol (4), diterpene ferruginol and lignan helioxanthin, were isolated from the whole plant of Taiwania cryptomerioides under bioassay-guided fractionations. The structures of 1 and 2 were elucidated mainly by the NMR spectroscopic analyses. Bioactivities of the isolated compounds against brine shrimp, yellow fever mosquito larvae, and human tumor cells are reported; compound 4 was the most bioactive, showing selectivity for the human colon tumor cell line (HT-29).

The Annonaceous, acetogenins are a class of natural products that have excellent anticancer, antiinfective, and pesticidal properties. 1 While the pace of discovering unknown acetogenins is showing an upward trend, 1 most of the newly reported acetogenins, however, belong to several classical types usually having unsubstituted tetrahydrofuran (THF) rings. 1 Mucocin, the first tetrahydropyran (THP) ring-bearing acetogenin, was recently discovered in the bioactive leaf extracts of Rollinia mucosa (Jacq.) Baill. (Annonaceae) 2 and has, as well, a nonadjacent THF ring. We have now isolated another closely related acetogenin, muconin (1, Figure 1), that bears an unsubstituted THP ring along with an adjacent THF ring. 1 is the second THP-bearing acetogenin to be reported and further indicates the existence of a novel acetogenin type with THP ring(s) as the most salient structural feature. In addition, mucoxin (2, Figure 1) is also reported here as the first acetogenin that bears a hydroxylated THF ring. 3 Both 1 and 2 were isolated 4 from the previously described leaf extract 5 by activity-directed open column fractionation using the brine shrimp lethality test 1 (BST) and, at later stages, by 1 H NMR-monitored purification using repetitive normal, and reversed-phase HPLC. The molecular formulas of 1 and 2 4 apparently suggested that they were C 37 acetogenins bearing two THF rings and three hydroxyls. The retention time of 2 on normal-phase HPLC, however, was much shorter than those typically observed for this type of acetogenins. Although the 1 H NMR spectra of 1 and 2 (Table 1) 6 showed the diagnostic peaks of the 2,4-disubstituted R, -unsaturated γ-lactone terminal (1 bears a 4-OH), which is common to most acetogenins, 1 they also presented certain salient features. For 1, the unusual δ values (3.16 and 3.31) of two protons were reminiscent of the THP oxymethines in mucocin. 2 For 2, the presence of a well-defined pseudo triplet (1 H, δ 3.74, J ) 3 Hz) was never encountered in our previous experiences with acetogenins. The above peaks were not lost to D 2 O exchanges.The presence of a THP ring in 1 was indicated by an intensive NOESY cross peak at δ 3.16/3.31 (later assigned as H-13/17) and the lack of the same cross peak in its regular, single-relayed, and double-relayed COSY spectra. The coexistence of a THF ring in 1 was required by its degree of unsaturation and was suggested by the 1 H and 13 C NMR data (Table 1). It was then deduced from the COSY spectrum that the THP ring was flanked on one side by a hydroxyl (OH-12) and, on the other, by a THF ring that was itself hydroxyl flanked. The position of the OH-12 was not proposed on the THP ring because no long-range 1 H-1 H coupling was observed between H-12 and additional oxygenated moieties, and the neighboring methylene protons of OH-12 did not show a split in their δ values. 7 The above NMR-based structural hypothesis was ambiguous in the sequential placement of the THF/THP system, and the absolute coordinates of oxygenation were determined by the EIMS o...