Bioactive Bromotyrosine Alkaloids from the Bahamian Marine Sponge Aiolochroia crassa. Dimerization and Oxidative Motifs.

Nine bromotyrosine alkaloids (BTAs), including debromoianthelline (1), pseudoceratinic acid (2a), methyl pseudoceratinate (2b), 13-oxo-ianthelline (3), aiolochroiamides A-D (4a,b and 5a,b), and 7-hydroxypurealidin J (6), were isolated from a Bahamian Aiolochroia crassa (Hyatt; previously, Pseudoceratina crassa). The structures of 1-6 were established from 1H, 13C, and 2D NMR spectra, IR, and mass spectrometry data. Compounds 2-4 comprise an O-methyl-2,6-dibromotyrosyl ketoxime (subunit A) amide linked to variable groups (subunit B). Compound 1 is debromoianthelline, and 2a and 2b are amides of 3-aminopropanoic acid and methyl 3-aminopropanoate, respectively. BTAs 3 and 4 are linked to 5-(2-aminoethyl)-2-iminoimidazolidin-4-one and a hexahydropyrrolo[2,3-d]imidazol-2(1H)-imine nucleus, respectively, whereas 5 is a self-dimerization motif of an aryl pyruvamide. Alkaloid 6 contains a spirocyclohexadienyl-isoxazoline-carboxamide amide coupled to 2-aminohistamine similar to that found in purealidin J and aerophobin-1 but with hydroxylation at C-7. The 2,4-diaminobutanoic acid residue in 3 was determined to be a 2:1 L- and D- mixture based on hydrolysis followed by derivatization with L-FDTA and LCMS. Diastereomeric pairs, 4a,b and 5a,b, were racemic. The relative configurations of 4a, 4b, 5a, and 5b were assigned by comparison of 1H and 13C chemical shifts with those calculated by DFT. Compounds 5a,b, ningalamide B (9), and ianthelline (7) moderately inhibited butyrylcholinesterase and Candida and Cryptococcus spp.

Spiroisoxazoline Inhibitors of Acetylcholinesterase from Pseudoceratina verrucosa. Quantitative Chiroptical Analysis of Configurational Heterogeneity, and Total Synthesis of (±)-Methyl Purpuroceratate C.

Examination of the MeOH extract of the sponge, Pseudoceratina cf. verrucosa, Berquist 1995 collected near Ningaloo Reef, Western Australia for selective acetylcholinesterase (AChE) inhibitors, yielded five new bromotyrosine alkaloids, methyl purpuroceratates A and B (1b and 2b), purpuroceratic acid C (3a), and ningalamides A and B (4 and 5). The structures of 1-4 share the dibromo-spirocyclohexadienyl-isoxazoline (SIO) ring system found in purealidin-R, while ketoxime 5 is analogous to ianthelline and purpurealidin I. The planar structures of all five compounds were obtained from analysis of MS, 1D and 2D NMR data, and the absolute configuration of the spiroisoxazoline (SIO) unit was assigned by electronic circular dichroism (ECD) and comparison with standards prepared by total synthesis of methyl purpuroceratate C, (±)-3b. Compound 4 is the most complex SIO described, to date. The configuration of the homoserine module (C) in 4 was ascertained, after acid hydrolysis, by derivatization of an l-tryptophanamide derivative based on Marfey's reagent. Chiral-phase HPLC, with comparison to synthetic standards, revealed that most SIOs isolated from P. cf. verrucosa were configurationally heterogeneous; some, essentially racemic. Chiral-phase HPLC, with UV-ECD detection, is demonstrated as a superlative method for configurational assignment and quantitation of the enantiomeric composition of SIOs. Two SIOs─aerophobin-1 and aplysinamisine II─emerged as selective inhibitors of AChE over butyrylcholinesterase (BuChE, IC50 ratio >10), while aplysamine-2 moderately inhibited both cholinesterases (ChEs, IC50, (AChE) 0.46 µM; IC50, (BuChE) 1.03 µM). SIO alkaloids represent a potential new structural manifold for lead-discovery of new therapeutics for treatment of Alzheimer's disease.

Bromo-spiroisoxazoline Alkaloids, Including an Isoserine Peptide, from the Caribbean Marine Sponge Aplysina lacunosa.

Three new bromotyrosine spiroisoxazoline alkaloids, lacunosins A and B (1 and 2) and desaminopurealin (3), were isolated from a MeOH extract of the marine sponge Aplysina lacunosa that showed modest α-chymotrypsin inhibitory activity. The structures of 1-3 share the spirocyclohexadienyl-isoxazoline ring system found in purealidin-R and several other Verongid sponge secondary metabolites. Compounds 1 and 2 are coupled to a glycine and an isoserine methyl ester, respectively. Alkaloid 3 is linked, contiguously, to an O-1-aminopropyl 3,5-dibromotyrosyl ether and, finally, to histamine through an amide bond. The planar structures of all three compounds were obtained from analysis of MS and 1D and 2D NMR data. The absolute configuration of the SIO unit of 1-3 was assigned by electronic circular dichroism (ECD). The isoserine amino acid residue in 2 was found to be a 1:1 mixture of epimers using a new Marfey's type reagent, derived from Trp-NH2. Allylic O-naphthoylation of the SIO subunit enhances the ECD spectrum of SIOs and improves discrimination of enantiomorphs. A unifying hypothesis is proposed that links the biosynthesis of several of the new compounds with previously reported analogues.

The Configuration of Distaminolyne A is S: Quantitative Evaluation of Exciton Coupling Circular Dichroism of N, O- Bis-arenoyl-1-amino-2-alkanols.

The 2 S configuration of the marine natural product distaminolyne A was recently disputed based upon total synthesis, yet paradoxically supported by a second independent total synthesis from a different research group. We now verify the 2 S configuration of distaminolyne A by extensive chiroptical studies and support the veracity of the EC ECD method originally used to prove it. The origin of the apparent paradox appears to lie in the limits of precision of polarimetry in the context of weakly rotatory molecules, which strikes a cautionary note on the reliability of "reassignment" of natural product configurations based solely on specific rotation.

Six Trikentrin-like Cyclopentanoindoles from Trikentrion flabelliforme. Absolute Structural Assignment by NMR and ECD.

Six new cyclopenta[g]indoles were isolated from a West Australian sponge, Trikentrion flabelliforme Hentschel, 1912, and their structures elucidated by integrated spectroscopic analysis. The compounds are analogues of previously described trikentrins, herbindoles, and trikentramides from related Axinellid sponges. The assignment of absolute configuration of the new compounds was carried out largely by comparative analysis of specific rotation, calculated and measured ECD, and exploiting van't Hoff's principle of optical superposition. Five of the new compounds were chemically interconverted to establish their stereochemical relationships, leading to a simple chiroptical mnemonic for assignment of the this family of chiral indoles. The first biosynthetic hypothesis is advanced to explain the origin of the trikentrin-herbinole family and proposes a pyrrole-carboxylic thioester-initiated polyketide synthase mechanism.

Cyclic Hexapeptide Dimers, Antatollamides A and B, from the Ascidian Didemnum molle. A Tryptophan-Derived Auxiliary for l- and d-Amino Acid Assignments.

Two dimerized cyclic hexapeptides, antatollamides A (1) and B (2), were isolated from the colonial ascidian Didemnum molle collected in Pohnpei. The amino acid compositions and sequences were determined by interpretation of MS and 1D and 2D NMR data. Raney Ni reduction of antatollamide A cleaved the dimer to the corresponding monomeric cyclic hexapeptide with replacement of Cys by Ala. The amino acid configuration of 1 was established, after total hydrolysis, by derivatization with a new chiral reagent, (5-fluoro-2,4-dinitrophenyl)-Nα-l-tryptophanamide (FDTA), prepared from l-Trp, followed by LCMS analysis; all amino acids were found to be l-configured except for d-Ala.

Small molecule in situ resin capture provides a compound first approach to natural product discovery.

Culture-based microbial natural product discovery strategies fail to realize the extraordinary biosynthetic potential detected across earth's microbiomes. Here we introduce Small Molecule In situ Resin Capture (SMIRC), a culture-independent method to obtain natural products directly from the environments in which they are produced. We use SMIRC to capture numerous compounds including two new carbon skeletons that were characterized using NMR and contain structural features that are, to the best of our knowledge, unprecedented among natural products. Applications across diverse marine habitats reveal biome-specific metabolomic signatures and levels of chemical diversity in concordance with sequence-based predictions. Expanded deployments, in situ cultivation, and metagenomics facilitate compound discovery, enhance yields, and link compounds to candidate producing organisms, although microbial community complexity creates challenges for the later. This compound-first approach to natural product discovery provides access to poorly explored chemical space and has implications for drug discovery and the detection of chemically mediated biotic interactions.

Small Molecule in situ Resin Capture - A Compound First Approach to Natural Product Discovery.

Microbial natural products remain an important resource for drug discovery. Yet, commonly employed discovery techniques are plagued by the rediscovery of known compounds, the relatively few microbes that can be cultured, and laboratory growth conditions that do not elicit biosynthetic gene expression among myriad other challenges. Here we introduce a culture independent approach to natural product discovery that we call the Small Molecule In situ Resin Capture (SMIRC) technique. SMIRC exploits in situ environmental conditions to elicit compound production and represents a new approach to access poorly explored chemical space by capturing natural products directly from the environments in which they are produced. In contrast to traditional methods, this compound-first approach can capture structurally complex small molecules across all domains of life in a single deployment while relying on Nature to provide the complex and poorly understood environmental cues needed to elicit biosynthetic gene expression. We illustrate the effectiveness of SMIRC in marine habitats with the discovery of numerous new compounds and demonstrate that sufficient compound yields can be obtained for NMR-based structure assignment. Two new compound classes are reported including one novel carbon skeleton that possesses a functional group not previously observed among natural products and a second that possesses potent biological activity. We introduce expanded deployments, in situ cultivation, and metagenomics as methods to facilitate compound discovery, enhance yields, and link compounds to producing organisms. This compound first approach can provide unprecedented access to new natural product chemotypes with broad implications for drug discovery.

Isolation of bastadin-6-O-sulfate and expedient purifications of bastadins-4, -5 and -6 from extracts of Ianthella basta.

Bastadin-6-34-O-sulfate ester (8) was isolated from methanol extracts of Ianthella basta. The structure of 8 was characterized by analysis of MS and NMR data, and conversion through acid hydrolysis, to the parent compound, bastadin-6, which was identical by HPLC, MS and NMR with an authentic sample. An improved procedure for procurement of pure samples of bastadins-4 (4), -5 (5) and -6 (6) is described.