A sample of S. striatinux roots was harvested in Oku in the Northwest Province of Cameroon. Following gradient chromatography on silica gel and subsequent size exclusion chromatography (SEC) on Sephadex LH-20, the extract of the dried roots gave okundoperoxide (1, R_f 0.3 in 3:2 hexanes:EtOAc). Okundoperoxide was unstable to gas chromatographic analysis,³ an observation that became understandable once the peroxide subunit was identified. High resolution ESI-MS analysis gave a sodiated parent (and base peak) ion of mass 289.1402. The ¹³C NMR spectrum in CDCl₃ indicated the presence of 15 carbon atoms. Together, these data showed a molecular formula of C₁₅H₂₂O₄ and five degrees of unsaturation.

Key ¹H and ¹³C NMR data are reported in Table 1. All 15 carbon and 22 (first order) proton resonances were identified. The ¹³C NMR spectrum contained resonances for one ketone and four olefinic carbons. The ¹H NMR spectrum suggested the presence of four methyl groups (one allylic with only long-range coupling and three aliphatic singlets) and three olefinic, one oxymethine, and one pair of oxymethylene protons. The HMQC spectrum clearly showed one-bond correlations that are the primary basis for the assignments of carbon chemical shifts listed in Table 1.

The IR spectrum showed characteristic absorption bands for hydroxy (3477 cm^-1) and carbonyl (1674 cm^-1) groups. The former was consistent with a one-proton resonance at δ 1.36 ppm, which disappeared in a deuterium exchange experiment. The carbonyl absorption was suggestive of a conjugated enone, which was supported in the NMR spectrum by the chemical shifts of olefinic proton (δ 6.73 and 5.94) and carbon (δ 150.3 and 127.9) signals and of the carbonyl carbon resonance (δ 203.2). These data, together with the doublets of the olefinic proton resonances (J = 10.2 Hz), pointed to a 4,4-disubstituted Z-enone moiety.

The ¹H-¹H COSY spectrum indicated an isolated four spin system that included the proton at δ 1.96, having three large coupling constants (13.0, 13.0, 11.2 Hz). This was indicative of an axial-like methylene proton in a six-membered ring, flanked by two vicinal, trans methine protons (-CHCH_axH_eqCH-). A COSY correlation between resonances for the olefinic proton at δ 5.75 and the methylene pair centered at δ 4.26 indicated a trisubstituted olefin bearing an oxymethylene group. The connectivity pattern deduced from the HMBC spectrum integrated the above subunits, along with the four methyl groups, into a common constitution. Specifically, structure 1 was consistent with all of the COSY and HMBC correlation data.

In addition to the 1,3-diaxial nature of H-4 and H-6 deduced from the coupling constant analysis, the remaining relative configurations shown in 1 were assigned largely on the basis of NOE observations (Figure 2). The acyclic E-olefin geometry is indicated by the enhancement of H-1 by H-15.⁴ Mutual enhancements of H-4 and H-6 reaffirm their cis-relationship. The trans nature of the ring fusion was deduced from the sets of NOEs among H-5_ax/H-12/H-14 and H-4/H-5_eq/H-6/H-13.

With the intent of reducing the peroxide bond in 1 with triphenylphosphine via an intermediate like 6 (Scheme 1), we treated a sample of 1 with Ph₃P in CDCl₃ and monitored the subsequent events by ¹H NMR spectroscopy. Somewhat surprisingly, there was no observable change at ambient temperature. Moreover, when the reaction solution was heated in a 65 °C bath, the major product formed was the furan 2, which has the same overall oxidation state as 1 and is the result of a net dehydration reaction. We suspect that enone 3 is an intermediate in this transformation. Zwitterion 6, if formed, could preferentially undergo intramolecular elimination of phosphine (see arrows in 6) rather than, for example, cyclization to a fused tetrahydrofuran derivative via displacement of triphenylphosphine oxide. Alternatively, the hindered nature of the dialkylperoxide in 1 may have induced a different reaction course from the outset. Namely, the phosphine may have functioned preferentially as a base rather than as a reductant to effect an eliminative opening via loss of H-4 and cleavage of the peroxide O-O bond (see arrows in 1) to give the enone 3.⁵ E-γ-Hydroxy-α,β-enones similar to 3 are known to undergo spontaneous isomerization and dehydration reactions to give furans.⁵.⁶ Enone E- to Z-isomerization to convert 3 to 4 could involve a reversibly formed, rotatable intermediate epoxide (cf. 7a) or triphenylphosphine adduct (cf. 7b). There are many reported examples of dehydration of Z-γ-hydroxy-α,β-enones like 4 under mild conditions to give the corresponding furans,⁷ likely via hemiketals like 5. It is notable that among the many thermal decomposition products observed upon GC-MS analysis of okundoperoxide (1), the furan 2 was the most abundant.³

Finally, the R- and the S-Mosher ester (methoxytrifluoromethylphenylacetyl, MTPA) derivatives of the alcohol 1 (8R and 8S, respectively, in Figure 3) were prepared using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and the R- and S-Mosher acid (MTPA-OH), respectively. The ¹H NMR data for these esters do not allow us to deduce the absolute configuration of 1 because of the large distance between the MTPA and substrate stereogenic centers. However, the spectra of these diastereomers are distinguishable, which should be helpful for later assignment of absolute configuration upon synthesis of one enantiomer of 1, an endeavor we are pursuing.

Antiplasmodial activity for the initial crude extract and for purified okundoperoxide (1) are shown in Table 2. Although the sample of 1 used for the biological testing was only 90% pure, we attribute the observed antiplasmodial activity to 1 because the interfering material, which was also isolated from a partially overlapped chromatographic fraction, is inactive in this assay. Finally, the presence of other compounds with antiplasmodial activity in S. striatinux cannot be ruled out. Further investigation of this possibility is also ongoing.

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