Vitifolignans, 3,4-dibenzyltetrahydrofuran lignans from Anemone vitifolia Buch.-Ham.

Anemone vitifolia is a small herb found in Asia that is used to treat a range of diseases in Chinese traditional medicine. GNPS-based molecular networking of an Anemone vitifolia specimen revealed the presence of a network containing numerous ions indicating the presence of lignans, several of which suggested that there might be previously undescribed compounds in the extract. Fractionation of the organic extract yielded five undescribed lignans, the vitifolignans, together with one known. The structures were identified based on extensive spectroscopic data analysis (NMR, HR-ESI-MS, and UV), coupling constant calculation and comparison with reported data. Their absolute configurations were determined by comparison of experimental ECD spectra with calculated spectra. Compounds 4/5 showed weak inhibition of LPS-induced NO production in mouse mononuclear macrophages.

Precursor-Directed Biosynthesis of Antialgal Fluorinated Bacillamide Derivatives in Bacillus atrophaeus.

The bacillamides are a class of indole alkaloids produced by the Bacillus genus that possess significant antialgal activity. Incorporation of fluorine into the bacillamides was carried out using a precursor-directed biosynthesis approach, with 4-, 5-, and 6-fluorotryptophan added to growing cultures of Bacillus atrophaeus IMG-11. This yielded the corresponding fluorinated analogues of bacillamides A and C, in addition to new derivatives of the related metabolite N-acetyltryptamine, thus demonstrating a degree of plasticity in the bacillamide biosynthetic pathway. The bacillamide derivatives were tested for activity against bloom-forming algae, which revealed that fluorination could improve the antialgal activity of these compounds in a site-specific manner, with fluorination at the 6-position consistently resulting in improved activity.

Expected and Unexpected Products from the Biochemical Oxidation of Bacterial Alkylquinolones with CYP4F11.

2-Alkylquinolones are a class of microbial natural products primarily produced in the Pseudomonas and Burkholderia genera that play a key role in modulating quorum sensing. Bacterial alkylquinolones were synthesized and then subjected to oxidative biotransformation using human cytochrome P450 enzyme CYP4F11, heterologously expressed in the fission yeast Schizosaccharomyces pombe. This yielded a range of hydroxylated and carboxylic acid derivatives which had undergone ω-oxidation of the 2-alkyl chain, the structures of which were determined by analysis of NMR and MS data. Oxidation efficiency depended on chain length, with a chain length of eight or nine carbon atoms proving optimal for high yields. Homology modeling suggested that Glu233 was relevant for binding, due to the formation of a hydrogen bond from the quinolone nitrogen to Glu233, and in this position only the longer alkyl chains could come close enough to the heme moiety for effective oxidation. In addition to the direct oxidation products, a number of esters were also isolated, which was attributed to the action of endogenous yeast enzymes on the newly formed ω-hydroxy-alkylquinolones. ω-Oxidation of the alkyl chain significantly reduced the antimicrobial and antibiofilm activity of the quinolones.

Deciphering the biotransformation mechanism of dialkylresorcinols by CYP4F11.

Cytochrome P450 enzymes (CYPs) are one of the most important classes of oxidative enzymes in the human body, carrying out metabolism of various exogenous and endogenous substrates. In order to expand the knowledge of these enzymes' specificity and to obtain new natural product derivatives, CYP4F11, a cytochrome P450 monooxygenase, was used in the biotransformation of dialkylresorcinols 1 and 2, a pair of antibiotic microbial natural products. This investigation resulted in four biotransformation products including two oxidative products: a hydroxylated derivative (3) and a carboxylic acid derivative (4). In addition, acetylated (5) and esterified products (6) were isolated, formed by further metabolism by endogenous yeast enzymes. Oxidative transformations were highly regioselective, and took place exclusively at the ω-position of the C-5 alkyl chain. Homology modeling studies revealed that optimal hydrogen bonding between 2 and the enzyme can only be established with the C-5 alkyl chain pointing towards the heme. The closely-related CYP4F12 was not capable of oxidizing the dialkylresorcinol 2. Modeling experiments rationalize these differences by the different shapes of the binding pockets with respect to the non-oxidized alkyl chain. Antimicrobial testing indicated that the presence of polar groups on the side-chains reduces the antibiotic activity of the dialkylresorcinols.

Experimental and Computational Studies on the Biotransformation of Pseudopyronines with Human Cytochrome P450 CYP4F2.

The secondary metabolite pseudopyronine B, isolated from Pseudomonas mosselii P33, was biotransformed by human P450 enzymes, heterologously expressed in the fission yeast Schizosaccharomyces pombe. Small-scale studies confirmed that both CYP4F2 and CYP4F3A were capable of oxidizing the substrate, with the former achieving a higher yield. In larger-scale studies using CYP4F2, three new oxidation products were obtained, the structures of which were elucidated by UV-vis, 1D and 2D NMR, and HR-MS spectroscopy. These corresponded to hydroxylated, carboxylated, and ester derivatives (1-3) of pseudopyronine B, all of which had been oxidized exclusively at the ω-position of the C-6 alkyl chain. In silico homology modeling experiments highlighted key interactions between oxygen atoms of the pyrone ring and two serine residues and a histidine residue of CYP4F2, which hold the substrate in a suitable orientation for oxidation at the terminus of the C-6 alkyl chain. Additional modeling studies with all three pseudopyronines revealed that the seven-carbon alkyl chain of pseudopyronine B was the perfect length for oxidation, with the terminal carbon lying close to the heme iron. The antibacterial activity of the substrates and three oxidation products was also assessed, revealing that oxidation at the ω-position removes all antimicrobial activity. This study both increases the range of known substrates for human CYF4F2 and CYP4F3A enzymes and demonstrates their utility in producing additional natural product derivatives.

Antibacterial and Anti-Biofilm Activity of Pyrones from a Pseudomonas mosselii Strain.

The emergence of drug resistant microbes over recent decades represents one of the greatest threats to human health; the resilience of many of these organisms can be attributed to their ability to produce biofilms. Natural products have played a crucial role in drug discovery, with microbial natural products in particular proving a rich and diverse source of antimicrobial agents. During antimicrobial activity screening, the strain Pseudomonas mosselii P33 was found to inhibit the growth of multiple pathogens. Following chemical investigation of this strain, pseudopyronines A-C were isolated as the main active principles, with all three pseudopyronines showing outstanding activity against Staphylococcus aureus. The analogue pseudopyronine C, which has not been well-characterized previously, displayed sub-micromolar activity against S. aureus, Staphylococcus epidermidis and Pseudomonas aeruginosa. Moreover, the inhibitory abilities of the pseudopyronines against the biofilms of S. aureus were further studied. The results indicated all three pseudopyronines could directly reduce the growth of biofilm in both adhesion stage and maturation stage, displaying significant activity at micromolar concentrations.

Molecular networking-driven isolation of 8'-Glycosylated biscoumarins from Cruciata articulata.

A molecular networking-guided phytochemical investigation of Cruciata articulata led to the isolation of five unreported biscoumarins, four of which were characterized by a shared 6-methoxy-7,8'-dihydroxy-3,7'-biscoumarin aglycone. These were isolated alongside two known coumarin glycosides, daphnetin-8-O-ß-D-glucoside and 6'-acetoxy-daphnetin-8-O-ß-D-glucoside. Their structures were elucidated by extensive 1D and 2D NMR experiments, in combination with chemical transformation and MS/MS fragmentation analysis. Four of the biscoumarins were glycosylated at the 8' position: these are the first examples of this substitution pattern to be described in nature. All compounds were tested for cytotoxic, antimicrobial, anti-inflammatory, and α-glucosidase inhibitory properties, but did not display significant activity.

Cruciasides C-G, monoterpenoid glycosides from Cruciata articulata.

Cruciata articulata (L.) Ehrend. is a herbaceous species distributed in parts of Western Asia and the Mediterranean region. While research on other species in the Cruciata genus has revealed the presence of a range of flavonoids and terpenoids, few such studies have been conducted on C. articulata. Thus, in the current study, a phytochemical investigation of C. articulata was carried out. Molecular networking identified a large cluster of compounds sharing distinctive MS-MS fragmentation patterns that were targeted for isolation, leading to the isolation of five undescribed monoterpenoid glycosides, cruciasides C-G, along with two known monoterpenoid glycosides. The structures of these compounds were elucidated by using chemical and spectroscopic analyses, including 1D and 2D NMR, and MS-MS fragmentation. Structures for the ions observed in the MS-MS were proposed, and based on these fragmentation patterns, structures for several of the minor components observed in the molecular network were also proposed. All isolated compounds were tested for cytotoxic, anti-inflammatory, antimicrobial, and α-glucosidase inhibitory properties, but did not display any activity.

Brevianthrones, bianthrones from a Chinese isolate of the endophytic fungus Colletotrichum brevisporum.

Seven undescribed bianthrones, the brevianthrones, together with two known anthraquinones, were isolated from the plant-derived fungus Colletotrichum brevisporum, obtained from the plant Piper sarmentosum Roxb., collected in Guangxi, China. This is the first report of the isolation of bianthrones from the Colletotrichum genus. The structures of the compounds were elucidated by a combination of NMR and MS spectroscopic analysis, while the absolute configurations were determined by X-ray crystallography and by simulation of ECD spectra.

Feruloyl sucrose derivatives from the root of Xerophyllum tenax.

A phytochemical investigation of the roots of Xerophyllum tenax led to the isolation of three undescribed feruloyl sucrose derivatives along with two known feruloyl sucrose derivatives, heloniosides A and B. This is the first report of their occurrence in the genus Xerophyllum and the family Melanthiaceae. The structures of these compounds were elucidated on the basis of chemical and spectroscopic analysis including 1D and 2D NMR and analysis of MS-MS fragmentation.

Semi-synthesis of antibacterial dialkylresorcinol derivatives.

Dialkylresorcinols are a class of antimicrobial natural products produced by a range of bacterial species. Semi-synthetic derivatization of two microbial dialkylresorcinols isolated from a Pseudomonas aurantiaca strain has yielded 21 derivatives, which were tested for antimicrobial activity, revealing several trends in their activity. The presence of aromatic and phenolic hydrogen atoms was crucial for activity, with all derivatives lacking these features possessing greatly reduced activity. On the other hand, derivatives with shorter alkyl chains at C-5 possessed lower MIC values, while one mono-fluorosulfated derivative showed significantly improved activity against several of the test strains.

Bacterial Alkyl-4-quinolones: Discovery, Structural Diversity and Biological Properties.

The alkyl-4-quinolones (AQs) are a class of metabolites produced primarily by members of the Pseudomonas and Burkholderia genera, consisting of a 4-quinolone core substituted by a range of pendant groups, most commonly at the C-2 position. The history of this class of compounds dates back to the 1940s, when a range of alkylquinolones with notable antibiotic properties were first isolated from Pseudomonas aeruginosa. More recently, it was discovered that an alkylquinolone derivative, the Pseudomonas Quinolone Signal (PQS) plays a key role in bacterial communication and quorum sensing in Pseudomonas aeruginosa. Many of the best-studied examples contain simple hydrocarbon side-chains, but more recent studies have revealed a wide range of structurally diverse examples from multiple bacterial genera, including those with aromatic, isoprenoid, or sulfur-containing side-chains. In addition to their well-known antimicrobial properties, alkylquinolones have been reported with antimalarial, antifungal, antialgal, and antioxidant properties. Here we review the structural diversity and biological activity of these intriguing metabolites.

Synthesis of Natural and Unnatural Quinolones Inhibiting the Growth and Motility of Bacteria.

Synthesis of a recently discovered S-methylated quinolone natural product (1) was carried out, in addition to the production of a range of 2-substituted 4-quinolone derivatives (2-11). Two approaches were used: (i) the base-catalyzed cyclization of N-(ketoaryl)amides; (ii) attachment of the substituent to the quinolone core via a Suzuki-Miyaura cross-coupling. Also produced were a small suite of related 2(1H)-quinolones (12-19). The synthesized compounds were assessed for their antimicrobial properties. The alkene-substituted 4-quinolone 8 significantly inhibited the growth of a Pseudomonas aeruginosa strain, and both 4-quinolones and 2(1H)-quinolones were capable of inhibiting the swarming behavior of Bacillus subtilis.

Isolation of 2-Alkyl-4-quinolones with Unusual Side Chains from a Chinese Pseudomonas aeruginosa Isolate.

Chemical investigation of a Pseudomonas aeruginosa strain isolated from Hebei, China, led to the isolation of a suite of quinolones, quinolone-N-oxides, and phenazines, the structures of which were elucidated by detailed spectroscopic analysis. Most notable among the secondary metabolites isolated was an unprecedented 4-quinolone containing an S-methyl group in the side chain and a new derivative including a phenyl ring in the side chain, which expand significantly the variety of structural motifs found in the quinolones and raise interesting questions about their biosynthesis.

Isolation, Identification, and Decomposition of Antibacterial Dialkylresorcinols from a Chinese Pseudomonas aurantiaca Strain.

A chemical investigation of a Chinese Pseudomonas aurantiaca strain has yielded a new benzoquinone (4) and furanone (5), in addition to the known dialkylresorcinols 1 and 2. Extensive decomposition studies on the major metabolite 1 produced an additional furanone derivative (6), a hydroxyquinone (7), and two unusual resorcinol and hydroxyquinone dimers (8 and 9). Structures were elucidated by nuclear magnetic resonance spectroscopy in combination with tandem mass spectrometry analysis. These studies illustrate the potential of artifacts as a source of additional chemical diversity. Compounds 1 and 2 showed moderate antibacterial activity against a panel of Gram-positive pathogens, while the antibacterial activities of the artifacts (4-9) were reduced.

Bioactive Compounds Isolated from Marine-Derived Microbes in China: 2009-2018.

This review outlines the research that was carried out regarding the isolation of bioactive compounds from marine-derived bacteria and fungi by China-based research groups from 2009-2018, with 897 publications being surveyed. Endophytic organisms featured heavily, with endophytes from mangroves, marine invertebrates, and marine algae making up more than 60% of the microbial strains investigated. There was also a strong focus on fungi as a source of active compounds, with 80% of publications focusing on this area. The rapid increase in the number of publications in the field is perhaps most notable, which have increased more than sevenfold over the past decade, and suggests that China-based researchers will play a major role in marine microbial natural products drug discovery in years to come.

Unusual Flavonoid Glycosides from the Hawaiian Tree Metrosideros polymorpha.

Metrosideros polymorpha is a highly variable and widely-distributed tree native to the Hawaiian islands. We describe here the isolation of two new gossypetin derivatives and three new C-methylated flavonol glycosides, which are highly uncommon and may prove to be useful chemotaxonomic markers for the species. In addition, a wide range of known flavonoid glycosides, chalcones, and terpenoids were isolated alongside the new compounds.

Chemotaxonomy of Hawaiian Anthurium cultivars based on multivariate analysis of phenolic metabolites.

Thirty-six anthurium varieties, sampled from species and commercial cultivars, were extracted and profiled by liquid-chromatography-mass spectrometry (HPLC-MS). Three hundred fifteen compounds, including anthocyanins, flavonoid glycosides, and other phenolics, were detected from these extracts and used in chemotaxonomic analysis of the specimens. Hierarchical cluster analysis (HCA) revealed close chemical similarities between all the commercial standard cultivars, while tulip-shaped cultivars and species displayed much greater chemical variation. Principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) supported the results from HCA and were used to identify key metabolites characteristic of standard and tulip cultivars and to identify chemical markers indicative of a particular ancestry. Discriminating metabolites included embinin, 4, which was characteristic of standard-shaped spathes and indicated ancestry from Anthurium andraeanum, while isocytisoside 7-glucoside, 7, was found in the majority of tulip-shaped cultivars and suggested that Anthurium amnicola or Anthurium antioquiense had contributed to their pedigree.

Flavone C-glycosides from Anthurium andraeanum.

We describe here the isolation of three flavone 6-C-glycosides from the leaves of Anthurium andraeanum, The two new flavones were identified through detailed spectroscopic analysis as 4"'-(3,4-dimethoxycinnamoyl)-embinin (2) and 4"'-ferruloyl-embinin (3).

Production of anticancer polyenes through precursor-directed biosynthesis.

The biosynthesis of the pyrrolyl moiety of the fungal metabolite rumbrin originates from pyrrole-2-carboxylic acid. In an effort to produce novel derivatives with enhanced biological activity a series of substituted pyrrole-2-carboxylates were synthesised and incubated with the producing organism, Auxarthron umbrinum. Several 4-halo-pyrrole-2-carboxylic acids were incorporated into the metabolite yielding three new derivatives: 3-fluoro-, 3-chloro- and 3-bromo-isorumbrin, which were generated in milligram quantities enabling cytotoxicity assays to be conducted. The 3-chloro- and 3-bromo-isorumbrins had improved activity against HeLa cells compared with rumbrin; 3-bromoisorumbrin also showed dramatically improved activity towards a lung cancer cell line (A549).