Synthesis of Macrolactone Core of ent-Formosalide A via Regioselective Ether Cyclization.

Formosalide A is a cytotoxic macrolide isolated from the dinoflagellate Prorocentrum sp, whose structure is characterized by functionalized 5- and 6-membered ether rings embedded in the macrolactone and an all cis-tetraene side chain. Here, we report the synthesis of the macrolactone core of ent-formosalide A. Our approach is highlighted by the Au-mediated 6-exo-dig cyclization for the synthesis of the 6-membered ether ring, which proceeded in a highly regioselective manner. Control experiments demonstrated that the acyclic protecting group of the C9,C10-diol was crucial for the desired 6-exo-dig cyclization. Theoretical studies were performed focusing on structural component analysis, which suggested that the C8-C9-C10-C11 dihedral angle induced by the protecting group controlled the regioselectivity. An additional 6 steps including Shiina macrolactone formation from the 6-membered ether ring completed the synthesis of the macrolactone core of ent-formosalide A.

Cladoic Acid, an Anti-Trypanosoma cruzi Polyketide from Cladosporium sp. TP-F2020.

A new polyketide, cladoic acid, was isolated from a fungus of the genus Cladosporium. The structure of the highly oxygenated trans-decalin ring with an all-E triene side chain was elucidated by extensive spectroscopic analysis. The unique chair/twist-boat conformation of the trans-decalin core and the flexibility of the B-ring were demonstrated by computer-aided conformational analysis. Cladoic acid was active against Trypanosoma cruzi and inhibited the proliferation of amastigotes and epimastigotes with IC50 values of 27 and 46 µM, respectively, but it did not show any appreciable activity against P388 murine leukemia cells, bacteria, or fungi, indicating it is a potential candidate for drug development against Chagas disease.

Computational Study Focusing on a Comprehensive Conformational Analysis of Transition States for Aza-Spiro Ring Formations with N-Alkoxyamides.

An accurate understanding of conformations in transition states is a critical piece in the theoretical analysis of complex molecular reactions. In this study, we investigated conformationally diverse transition states during intramolecular aza-spiro ring formation with an allylsilane moiety and N-alkoxy iminium ion, a key reaction in the synthesis of fasicularin by Sato and Chida et al., and identified the origins of stereoselectivity of the cyclization. A large number of conformational isomers with forming C-C bonds were comprehensively analyzed using Cremer-Pople puckering parameters. It was found that the conformations of the transition states had different puckering preferences depending on the reactant's double-bond geometry and the product's stereochemical configuration. Furthermore, an asymmetric aza-spiro cyclization with a tolyl group as a chiral auxiliary was investigated, showing that conformational anchoring by both a CH-O hydrogen bond and the CH-π interaction was critical for the asymmetric induction.

Formal Total Synthesis of Batrachotoxin Enabled by Radical and Weix Coupling Reactions.

Batrachotoxin (1), originally isolated from a Columbian poison-dart frog, is a steroidal alkaloid. Its 6/6/6/5-membered carbocycle (ABCD-ring) contains two double bonds, one nitrogen, and five oxygen functionalities. We developed a radical-based convergent strategy and realized the total synthesis of 1 in 28 steps. The AB-ring and D-ring fragments were efficiently synthesized and linked by exploiting a powerful Et3B/O2-mediated radical coupling reaction. Vinyl triflate and vinyl bromide were then utilized for a Pd/Ni-promoted Weix coupling reaction to cyclize the C-ring. A hydroxy group of the C-ring was stereoselectively installed by a decarboxylative hydroxylation reaction to prepare an advanced intermediate of our previous total synthesis of 1.

Stereocontrolled Synthesis of C20S-C26 and C20R-C26 Fragments of Amphidinolide L.

Amphidinolide L is a cytotoxic macrolide isolated from marine symbiotic dinoflagellates of the genus Amphidinium. While its planar structure and the absolute stereochemistry of the C21-C26 part have been determined, six stereocenters have remained unassigned. Aiming at structure determination, we have developed a synthetic route to the C20S-C26 and C20R-C26 fragments via the Li-mediated stereocontrolled aldol reaction. Two aldehydes, 16 with the C22-hydroxy group and 19 with the C22-TES ether, were synthesized from lactone 4. The aldol reactions using the Li-enolate of 4-methyl-2-pentanone in THF provided the C20S-C26 fragment 20 from 16 and a 1:3.5 mixture of the C20-C26 fragment 22 favoring the C20R-isomer. Mechanistic studies based on an extensive search of transition states in explicit solvents indicated that the C20S-isomer would be generated via a tri-solvated transition state, while the C20R-isomer would be formed via a di-solvated transition state. The calculation emphasizes the importance of the coordination network as a higher-order complex composed of solvent molecules, aldehyde, enolate, and Li atoms in the reaction of 16 to minimize steric interactions but maximize the stabilizing effect by the coordination of solvents. The presence of the rotationally free aldehyde in the reaction of 19 results in moderate diastereoselectivity.

Collective Synthesis of Aplysiatoxin/Oscillatoxin Analogues by a Bioinspired Strategy.

Interest in the marine cyanobacteria natural products aplysiatoxin (ATX) and oscillatoxin (OTX) has been renewed recently due to the discovery of many new analogues, some exhibiting intriguing biological activities. We sought to develop a collective synthesis of these natural products, hypothesizing that ATX could serve as a common biosynthetic precursor. Herein, we reveal that the core structure of ATX has unique multiple reactivities giving access to the distinct ring structures of five of the analogues, depending upon the specific conditions used. Based on these findings, syntheses of the O-Me derivative of five analogues neo-deBr-ATX-B, OTX-H, OTX-D, neo-deBr-ATX-H, and OTX-I were achieved from the main fragment of ATX as a common intermediate in a few steps. These synthetic studies also led us to revise the relative configuration in the elucidated structures of neo-deBr-ATX-B and OTX-H, and obtain unnatural 8- and 12-membered lactones from the same intermediate.

Krasilnikolides A and B and Detalosylkrasilnikolide A, Cytotoxic 20-Membered Macrolides from the Genus Krasilnikovia: Assignment of Anomeric Configuration by J-Based Configuration Analysis.

A chemical investigation of strain RD003821, belonging to the underexplored actinomycetes genus Krasilnikovia, led to the discovery of three novel polyketides: two 20-membered glycomacrolides, krasilnikolides A (1) and B (2), and an aglycone of 1, detalosylkrasilnikolide A (3). A major challenge in the structure elucidation of 1 was to determine the anomeric configuration of the α-l-6-deoxytalose (6dTal) unit, which was achieved by J-based configuration analysis (JBCA) that incorporated anomeric carbon- and proton-specific two-bond 13C-1H spin-spin coupling constants as diagnostic parameters. The updated criteria for the conformation/configuration assignment facilitated discrimination of three out of four stereochemical variants at the anomeric and the adjacent C2 positions, which expanded the scope of the JBCA method to determination of the anomeric configuration of aldohexopyranoses. Compounds 1 and 2 are the first macrolides decorated by 6dTal. Compounds 1-3 exhibited cytotoxicity against P388 murine leukemia cells with IC50 values of 14, 8.4, and 3.9 µM, respectively. In addition, 1-3 were antibacterial against the Gram-positive bacterium Kocuria rhizophila with MIC values of 25, 50, and 100 µg/mL. 1 was inhibitory against Staphylococcus aureus with an MIC of 50 µg/mL.

Catellatolactams A-C, Plant Growth-Promoting Ansamacrolactams from a Rare Actinomycete of the Genus Catellatospora.

Catellatolactams A-C (1-3), three novel ansamacrolactams, were isolated from the culture extract of an underexplored rare actinomycete of the genus Catellatospora. Spectroscopic and spectrometric analyses by NMR and MS elucidated the structure of 1 to be a lactamized pentaketide presumably extended on a 3-amino-5-hydroxybenzoic acid starter unit. Compounds 2 and 3 further received epoxidation and intramolecular cross-linking to incorporate a 2-indolinone unit, with a 3-amino-5-hydroxybenzoic acid pendant on 3. The absolute configurations of 2 and 3 were unequivocally established to both be 2S,6R,7R by comparison of the experimental NMR chemical shifts and ECD spectra with those predicted by DFT-based quantum chemical calculation. While 1-3 showed no appreciable antimicrobial activity or cytotoxicity, root elongation of germinated lettuce seeds was promoted by 2 and 3 at 1-10 µM.

Marinoquinolones and Marinobactoic Acid: Antimicrobial and Cytotoxic ortho-Dialkylbenzene-Class Metabolites Produced by a Marine Obligate Gammaproteobacterium of the Genus Marinobacterium.

Chemical investigation of the culture extract of a marine obligate proteobacterium, Marinobacterium sp. C17-8, isolated from scleractinian coral Euphyllia sp., led to the discovery of three new o-dialkylbenzene-class metabolites, designated marinoquinolones A (1) and B (2) and marinobactoic acid (3). Spectroscopic analysis using MS and NMR revealed the structures of 1 and 2 to be 4-quinolones with an o-dialkylbenzene-containing side chain at C3 and 3 to be a fatty acid bearing an o-dialkylbenzene substructure. The 4-quinolone form of 1 and 2 was unequivocally determined by comparison of the 1H, 13C, and 15N chemical shifts of 1 with those predicted for 2-methyl-4-quinolone A and its tautomer 2-methyl-4-quinolinol B by quantum chemical calculation. Compound 1 was proven to be racemic by X-ray crystallographic analysis and chiral-phase HPLC analysis of its chemical degradation product. Compounds 1-3 exhibited antimicrobial activity against bacteria and filamentous fungi at MIC of 6.3-50 µg/mL. In addition, all compounds showed cytotoxicity against P388 murine leukemia cells at micromolar ranges.

Kumemicinones A-G, Cytotoxic Angucyclinones from a Deep Sea-Derived Actinomycete of the Genus Actinomadura.

Chemical investigation of the fermentation products of a deep sea water-derived actinomycete, Actinomadura sp. KD439, identified seven new angucyclinones, designated as kumemicinones A-G (1-7), together with the known SF2315B and miaosporone E. NMR and MS spectroscopic analyses, combined with X-ray crystallography and quantum chemical calculations of NMR chemical shifts and electronic circular dichroism (ECD) spectra, uncovered the structures of new angucyclinones as regioisomers of SF2315B at the allyl alcohol unit (1 and 2), an epoxy ring-opened γ-hydroxy enone isomer (3), a B/C-ring-rearranged product (4), or dimers with a new mode of bridging (5-7), adding new structural variation to this antibiotic group. The absolute configuration of SF2315B was also determined by comparison of ECD spectra with those of 1 and 2. All the angucyclinones exhibited cytotoxicity against P388 murine leukemia cells, with IC50 values ranging from 1.8 to 53 µM.

RCM approach to the CDE-tricyclic structure of nakiterpiosin.

A synthetic approach to the CDE-tricyclic structure of nakiterpiosin, a marine-derived antimitotic C-nor-D-homosteroid, is reported. The trans-disubstituted indanone was synthesized from a commercially available carboxylic acid in 9 steps, and the ring closing metathesis of the indanone constructed the trans-fused 5/6-membered ring system. The present approach enables the concise synthesis of the functionalized CDE-tricyclic structure, which will serve as a synthetic intermediate toward nakiterpiosin.

Structure Determination, Biosynthetic Origin, and Total Synthesis of Akazaoxime, an Enteromycin-Class Metabolite from a Marine-Derived Actinomycete of the Genus Micromonospora.

A new enteromycin-class antibiotic, akazaoxime (1), possessing an aldoxime functionality in place of O-methyl nitronic acid, was isolated from the cultured extract of a marine-derived actinomycete of the genus Micromonospora, along with known A-76356 (2). The structure of 1, including the absolute stereochemistry of three chiral centers, was established by comprehensive analysis of nuclear magnetic resonance (NMR) and mass spectrometry data coupled with magnetic anisotropy analysis of its phenylglycine methyl ester derivatives. The stereochemistry of 2, not determined previously, was proven to be the same as that of 1 on the basis of the similarity of their NMR and specific rotation data. Precursor feeding experiments using 13C-labeled compounds elucidated that the carbon skeletons of 1 and 2 are constructed from propionate (methylmalonate), leucine, and glycine. Establishment of the concise and flexible synthetic route to 1 enabled us to implement biological evaluation of 1 and its unnatural analogues, demonstrating weak to moderate antimicrobial activities of 1 against Gram-positive Kocuria rhizophila [minimum inhibitory concentration (MIC) of 50 µg/mL] and those of synthetic analogues against a plant pathogen Glomerella cingulata (MIC of 50 µg/mL) and a human pathogen Trichophyton rubrum (MIC of 25-50 µg/mL).

Cyclopeptides from the Mushroom Pathogen Fungus Cladobotryum varium.

Three new cyclopeptides with serial Phe residues were identified with the aid of HPLC-DAD analysis, from the culture broth of Cladobotryum varium, a fungal pathogen causing mushroom cobweb disease. Cladoamides A (1) and B (2) have two consecutive N-methylphenylalanine units in the destruxin class cyclic depsipentapeptide framework, while cladoamide C (3) has a three consecutive Phe motif in a cyclopentapeptide structure. Of these three cyclopeptides, 1 showed potent autophagy-inducing activity at 10 µg/mL, comparable to a positive control, rapamycin. For the determination of the absolute configurations of the Ile residues in 1 and 3, new conditions for separating Ile and allo-Ile, using a pentafluorophenyl-bonded solid phase and methanolic solvent, were established within the analytical scheme of the advanced Marfey's method, thus offering a convenient alternative to the C3 Marfey's method, which requires elution with a three-solvent mixture. The sequence of two d-Phe and one l-Phe in 3 was determined through NMR chemical shift prediction by DFT-based calculations and chemical synthesis, which demonstrated the significance of noncovalent interactions in the accurate calculation of stable conformers for peptides with multiple aromatic rings.

Nomimicins B-D, new tetronate-class polyketides from a marine-derived actinomycete of the genus Actinomadura.

Three new tetronate-class polyketides, nomimicins B, C, and D, along with nomimicin, hereafter named nomimicin A, were isolated from the culture extract of Actinomadura sp. AKA43 collected from floating particles in the deep-sea water of Sagami Bay, Japan. The structures of nomimicins B, C, and D were elucidated through the interpretation of NMR and MS analytical data, and the absolute configuration was determined by combination of NOESY/ROESY and ECD analyses. Nomimicins B, C, and D showed antimicrobial activity against Gram-positive bacteria, Kocuria rhizophila and Bacillus subtilis, with MIC values in the range of 6.5 to 12.5 µg/mL. Nomimicins B and C also displayed cytotoxicity against P388 murine leukemia cells with IC50 values of 33 and 89 µM, respectively.

Computational Analysis of the Selective Formation of the C4α-C8' Bond in the Intermolecular Coupling of Catechin Derivatives.

Procyanidin B3 is a natural flavonoid composed of two catechins connected via a C4α-C8' bond. The couplings of catechin derivatives, promoted by Lewis acids, have been widely applied to the syntheses of procyanidin B3 and related flavonoids because the reactions construct the C4α-C8' bond in a highly stereo- and regioselective manner. However, the structural complexity of the catechin derivatives has complicated the exploration of a detailed mechanism for this selectivity. Here, we report the results of a computational study to provide plausible origins for the selective C4α-C8' bond formation of catechin derivatives 1 and 2 by using models 5 and 7. Although a systematic search did not provide SN2-like transition states, we successfully identified transition states TS-A, TS-B, and TS-C for the SN1-type C4α-C8', C4ß-C8', and C4α-C6' bond formations, respectively, from a total of 233 transition states to justify the stereo- and regioselectivity of the experimental results. The analysis of these structures by NCIPLOT mapping and the distortion/interaction strain model suggests that the eclipsed interaction at the forming C-C bond between the electrophile and the nucleophile destabilizes TS-B, while the strain of the electrophile destabilizes TS-C. Consequently, the C4α-C8' bond is formed via the lowest energy transition state TS-A.

Computational and Experimental Analysis on the Conformational Preferences of Anticancer Saponin OSW-1.

The conformational properties of anticancer saponin OSW-1 were investigated by X-ray crystallography and by an integrated approach combining a conformational search and the evaluation of the computed conformational distribution by comparing the experimental and simulated spectroscopic data. Our results suggested that OSW-1 adopts two preferred conformations in solution at an approximately 2:1 ratio, of which the crystal structure is consistent with the major conformation. In the solution models, the arabinose residue of OSW-1 appears to serve as a molecular hinge by flipping from the standard 4C1 form in the major conformer to the unusual 1C4 form in the minor conformer. This results in different orientations of the biologically essential p-methoxybenzoyl group, thereby inducing a dramatic alteration of the three-dimensional shape and polarity of OSW-1.

Total Synthesis of Talatisamine.

Talatisamine (1) is a member of the C19 -diterpenoid alkaloid family, and exhibits K+ channel inhibitory and antiarrhythmic activities. The formidable synthetic challenge that 1 presents is due to its highly oxidized and intricately fused hexacyclic 6/7/5/6/6/5-membered-ring structure (ABCDEF-ring) with 12 contiguous stereocenters. Here we report an efficient synthetic route to 1 by the assembly of two structurally simple fragments, chiral 6/6-membered AE-ring 7 and aromatic 6-membered D-ring 6. AE-ring 7 was constructed from 2-cyclohexenone (8) through fusing an N-ethylpiperidine ring by a double Mannich reaction. After coupling 6 with 7, an oxidative dearomatization/Diels-Alder reaction sequence generated fused pentacycle 4 b. The newly formed 6/6-membered ring system was then stereospecifically reorganized into the 7/5-membered BC-ring of 3 via a Wagner-Meerwein rearrangement. Finally, Hg(OAc)2 induced an oxidative aza-Prins cyclization of 2, thereby forging the remaining 5-membered F-ring. The total synthesis of 1 was thus accomplished by optimizing and orchestrating 33 transformations from 8.

Nocarimidazoles C and D, antimicrobial alkanoylimidazoles from a coral-derived actinomycete Kocuria sp.: application of 1 J C,H coupling constants for the unequivocal determination of substituted imidazoles and stereochemical diversity of anteisoalkyl chains in microbial metabolites.

Chemical investigation of secondary metabolites from a marine-derived actinomycete strain of the genus Kocuria, isolated from a stony coral Mycedium sp., led to the identification of two new alkanoylimidazoles, nocarimidazoles C (1) and D (2) as well as three known congeners, nocarimidazoles A (3) and B (4) and bulbimidazole A (5). Structure analysis of 1 and 2 by NMR and MS revealed that both are 4-alkanoyl-5-aminoimidazoles with a 6-methyloctanoyl or decanoyl chain, respectively. Two possible positions of the amino group on the imidazole rings (C-2 and C-5) posed a challenge in the structure study, which was settled by the measurement of 1 J C,H coupling constants for comparison with those of synthetically prepared model imidazoles. The absolute configurations of the anteisoalkanoyl group present in 1, 4, and 5 were determined by low-temperature HPLC analysis of the degradation products labeled with a chiral anthracene reagent, which revealed that 1 is a mixture of the R- and S-enantiomers with a ratio of 73:27, 4 is the pure (S)-enantiomer, and 5 is the (S)-enantiomer with 98% ee. The present study illustrates the diversity in the stereochemistry of anteiso branching in bacterial metabolites. Compounds 1-4 were moderately antimicrobial against Gram-positive bacteria and fungi, with MIC ranges of 6.25-25 µg/mL.

A cyclopeptide and three oligomycin-class polyketides produced by an underexplored actinomycete of the genus Pseudosporangium.

Aside from the well-studied conventional actinomycetes such as Streptomyces, the less investigated genera of actinomycetes also represent a promising source of natural products. Genome mining indicated that members of the underexplored genus Pseudosporangium, from which no secondary metabolites have been reported to date, may harbor the biosynthetic machinery for the formation of novel natural products. The strain RD062863, that is available at a public culture collection, was obtained and subjected to metabolite analysis, which resulted in the discovery of a novel cyclopeptide, pseudosporamide (1), along with three new oligomycin-class polyketides, pseudosporamicins A-C (2-4). The unusual structure of compound 1, featured by a biaryl-bond bridging across a tripeptide scaffold, N-acetyl-ʟ-Tyr-ʟ-Pro-ʟ-Trp, was determined by a combination of spectroscopic analyses, chemical derivatization, ECD calculation, and DFT-based theoretical chemical shift calculation, revealing the presence of an (S a)-axial chirality around the biaryl bond. Compounds 2-4 lacked hydroxylation on the side chain of the spiroacetal rings, which showed clear contrast to other oligomycin congeners and related polyketides with ring-truncation or expansion. The new macrolides 2-4 displayed potent antimicrobial activity against the Gram-positive bacterium Kocuria rhizohpila and the plant pathogenic fungus Glomerella cingulata. All compounds showed moderate cytotoxicity against P388 murine leukemia cells with IC50 values in the micromolar to submicromolar ranges. These results exemplified the validity of phylogeny-focused strain selection combined with biosynthetic gene-directed genome mining for the efficient discovery of new natural products.

A Computational Study on the Stereo- and Regioselective Formation of the C4α-C6' Bond of Tethered Catechin Moieties by an Exhaustive Search of the Transition States.

We previously reported the total synthesis of procyanidin B6 by using the stereo- and regioselective C-C bond formation of tethered catechin moieties as the key step. The reaction afforded the product bearing a new C4α-C6' bond linkage instead of the inherently preferable C4α-C8' bond. However, the origin of this selectivity remained unclear due to the complex structure of the substrate. Here we report the results of computational exploration of this C-C bond formation to gain mechanistic insights into the selectivity. The computational study of highly flexible compounds was realized by an exhaustive search of transition states. A large library of candidate transition states was generated by a conformational search of constrained models using molecular mechanics simulations and semiempirical molecular orbital calculations. Subsequent DFT-based transition state calculations provided 367 transition states for C4-C6' and C4-C8' bond formations. Comparison of the geometries and energies showed that the C4α-C6' linkage is preferentially formed via two competing transition states, leading to a C6'-diastereomeric mixture. Interactive atomic distances and visualization of the nonbonding interactions suggest the importance of nonclassical hydrogen bonding and CH-π, π-π, and lone pair-π interactions in stabilizing the two transition states. The present study supports preferential C4α-C6' bond formation of the tethered catechins.