Silver Organometallics that are Highly Potent Thioredoxin and Glutathione Reductase Inhibitors: Exploring the Correlations of Solution Chemistry with the Strong Antibacterial Effects.

The antibacterial activity of silver species is well-established; however, their mechanism of action has not been adequately explored. Furthermore, issues of low-molecular silver compounds with cytotoxicity, stability, and solubility hamper their progress to drug leads. We have investigated silver N-heterocyclic carbene (NHC) halido complexes [(NHC)AgX, X = Cl, Br, and I] as a promising new type of antibacterial silver organometallics. Spectroscopic studies and conductometry established a higher stability for the complexes with iodide ligands, and nephelometry indicated that the complexes could be administered in solutions with physiological chloride levels. The complexes showed a broad spectrum of strong activity against pathogenic Gram-negative bacteria. However, there was no significant activity against Gram-positive strains. Further studies clarified that tryptone and yeast extract, as components of the culture media, were responsible for this lack of activity. The reduction of biofilm formation and a strong inhibition of both glutathione and thioredoxin reductases with IC50 values in the nanomolar range were confirmed for selected compounds. In addition to their improved physicochemical properties, the compounds with iodide ligands did not display cytotoxic effects, unlike the other silver complexes. In summary, silver NHC complexes with iodide secondary ligands represent a useful scaffold for nontoxic silver organometallics with improved physicochemical properties and a distinct mechanism of action that is based on inhibition of thioredoxin and glutathione reductases.

Omnipolyphilins A and B: Chlorinated Cyclotetrapeptides and Naphtho-α-pyranones from the Plant Nematode-Derived Fungus Polyphilus sieberi.

Chemical exploration for two isolates of the recently described ascomycete species Polyphilus sieberi, derived from the eggs of the plant parasitic nematode Heterodera filipjevi, afforded the identification of many compounds that belong to various metabolite families: two previously undescribed chlorinated cyclotetrapeptides, omnipolyphilins A (1) and B (2), one new pyranonaphthoquinone, ventiloquinone P (3), a 6,6'-binaphto-α-pyranone dimer, talaroderxine D (4) in addition to nine known metabolites (5-13) were isolated from this biocontrol candidate. All isolated compounds were characterized by comprehensive 1D, 2D NMR, and HR-ESI-MS analyses. The absolute configurations of the cyclotetrapeptides were determined by a combination of advanced Marfey's method, ROE correlation aided by conformational analysis, and TDDFT-ECD calculations, while ECD calculations, Mosher's method, and experimental ECD spectra were used for ventiloquinone P (3) and talaroderxine D (4). Among the isolated compounds, talaroderxine D (4) showed potent antimicrobial activities against Bacillus subtilis and Staphylococcus aureus with MIC values of 2.1 and 8.3 µg mL-1, respectively. Additionally, promising inhibitory effects on talaroderxine D (4) against the formation of S. aureus biofilms were observed up to a concentration of 0.25 µg mL-1. Moreover, ophiocordylongiiside A (10) showed activity against the free-living nematode Caenorhabditis elegans.

Discovery of novel secondary metabolites from the basidiomycete Lentinus cf. sajor-caju and their inhibitory effects on Staphylococcus aureus biofilms.

Three novel derivatives of microporenic acid, microporenic acids H-J, were identified from submerged cultures of a Lentinus species obtained from a basidiome collected during a field trip in the tropical rainforest in Western Kenya. Their structures were elucidated via HR-ESIMS spectra and 1D/2D NMR spectroscopic analyses, as well as by comparison with known derivatives. Applying biofilm assays based on crystal violet staining and confocal microscopy, two of these compounds, microporenic acids H and I, demonstrated the ability to inhibit biofilm formation of the opportunistic pathogen Staphylococcus aureus. Thereby, they were effective in a concentration range that did not affect planktonic growth. Additionally, microporenic acid I enhanced the anti-biofilm activity of the antibiotics vancomycin and gentamicin when used in combination. This opens up possibilities for the use of these compounds in combination therapy to prevent the formation of S. aureus biofilms.

Lachnuoic Acids A-F: Ambuic Acid Congeners from a Saprotrophic Lachnum Species.

Chemical prospection of an extract derived from a saprotrophic fungus Lachnum sp. IW157 resulted in the isolation and characterization of six unprecedentedly reported ambuic acid analogues named lachnuoic acids A-F (1-6). Chemical structures of 1-6 were determined based on comprehensive 1D and 2D NMR spectroscopic analyses together with HR-ESI-MS spectrometry. The relative configurations of 1-3 were defined by ROESY spectroscopic analyses while their absolute configurations were unambiguously determined by Mosher's esters method. All isolated compounds were subjected to cytotoxic, antimicrobial, antibiofilm and nematicidal activity assays where only lachnuoic acid A (1) revealed potent antimicrobial activity against Staphylococcus aureus and Bacillus subtilis at MIC values of 16.6 and 8.3 µg/mL, respectively.

Bioprospection of Tenellins Produced by the Entomopathogenic Fungus Beauveria neobassiana.

Fungi are known as prolific producers of bioactive secondary metabolites with applications across various fields, including infectious diseases, as well as in biological control. However, some of the well-known species are still underexplored. Our current study evaluated the production of secondary metabolites by the entomopathogenic fungus Beauveria neobassiana from Thailand. The fermentation of this fungus in a liquid medium, followed by preparative high-performance liquid chromatography (HPLC) purification, resulted in the isolation of a new tenellin congener, namely pretenellin C (1), together with five known derivatives (2-6). Their chemical structures were elucidated by 1D and 2D nuclear magnetic resonance (NMR) spectroscopy in combination with high-resolution electrospray ionization mass spectrometry (HR-ESI-MS). We evaluated the antimicrobial and cytotoxic activities from all isolated compounds, as well as their inhibitory properties on biofilm formation by Staphylococcus aureus. Generally, tenellins displayed varying antibiofilm and cytotoxic effects, allowing us to propose preliminary structure-activity relationships (SARs). Among the tested compounds, prototenellin D (2) exhibited the most prominent antibiofilm activity, while its 2-pyridone congener, tenellin (4), demonstrated potent cytotoxic activity against all tested cell lines. Given the fact that the biological activities of the tenellins have so far been neglected in the past, our study could provide a good starting point to establish more concise structure-activity relationships in the near future.

Diaporphasines E and F: New Polyketides from the Saprotrophic Fungus Lachnum sp. IW157 Growing on the Reed Grass Phragmites communis.

Chemical investigation for the mycelial extract of a saprotrophic fungus Lachnum sp. IW157 growing on the common reed grass Phragmites communis afforded the identification of two polyketide metabolites diaporphasines E (1) and F (2). Chemical structures of isolated compounds were unambiguously elucidated based on extensive 1D and 2D NMR spectral analyses in addition to their high-resolution mass spectrometry. The isolated compounds were assessed for their cytotoxicity and antimicrobial and biofilm inhibitory activities. While compound 1 revealed potent cytotoxicity against the tested cell lines L929 and KB3.1 with IC50 values of 0.9 and 3.7 µM, respectively, compound 2 exhibited moderate effects on the formation of S. aureus biofilms at 31.25 µg/mL.

Bioactive Compounds from an Endophytic Pezicula sp. Showing Antagonistic Effects against the Ash Dieback Pathogen.

A fungal endophyte originating from the Canary Islands was identified as a potent antagonist against the fungal phytopathogen Hymenoscyphus fraxineus, which causes the devastating ash dieback disease. This endophyte was tentatively identified as Pezicula cf. ericae, using molecular barcoding. Isolation of secondary metabolites by preparative high-performance liquid chromatography (HPLC) yielded the known compounds CJ-17,572 (1), mycorrhizin A (3) and cryptosporioptides A-C (4-6), besides a new N-acetylated dihydroxyphenylalanin derivative 2, named peziculastatin. Planar structures were elucidated by NMR and HRMS data, while the relative stereochemistry of 2 was assigned by H,H and C,H coupling constants. The assignment of the unknown stereochemistry of CJ-17,572 (1) was hampered by the broadening of NMR signals. Nevertheless, after semisynthetic conversion of 1 into its methyl derivatives 7 and 8, presumably preventing tautomeric effects, the relative configuration could be assigned, whereas comparison of ECD data to those of related compounds determined the absolute configuration. Metabolites 1 and 3 showed significant antifungal effects in vitro against H. fraxineus. Furthermore, 4-6 exhibited significant dispersive effects on preformed biofilms of S. aureus at concentrations up to 2 µg/mL, while the biofilm formation of C. albicans was also inhibited. Thus, cryptosporioptides might constitute a potential source for the development of novel antibiofilm agents.

Sporothioethers: deactivated alkyl citrates from the fungus Hypomontagnella monticulosa.

Submerged cultivation of Hypomontagnella monticulosa MUCL 54604 resulted in formation of a stereoisomeric mixture of new sulfur-containing sporothriolide derivatives named sporothioethers A and B. The presence of the 2-hydroxy-3-mercaptopropanoic acid moiety attenuates the antimicrobial activity in comparison to the precursor sporothriolide suggesting a detoxification mechanism. However, moderate effects on biofilms of Candida albicans and Staphylococcus aureus were observed for sporothriolide and sporothioethers A and B at concentrations below their MICs.

Biologically active drimane derivatives isolated from submerged cultures of the wood-inhabiting basidiomycete Dentipellis fragilis.

Four previously undescribed drimane sesquiterpenoids were isolated from submerged cultures of the wood-inhabiting basidiomycete Dentipellis fragilis along with two compounds that were previously reported as synthetic or biotransformation compounds but not as natural products. The constitution and relative configuration of these compounds was determined based on high-resolution electrospray ionization mass spectrometry as well as by 1D and 2D nuclear magnetic resonance spectroscopy. The absolute configurations were established based on exemplary calculation of circular dichroism spectra and comparison with measured data as well as on biogenetic considerations. The biological activities of the isolated compounds were assessed in antimicrobial, cytotoxicity and neurotrophic assays. 10-Methoxycarbonyl-10-norisodrimenin (3) exhibited weak activity against the Gram-positive bacterium Staphylococcus aureus and the zygomycete Mucor hiemalis with minimal inhibitory concentrations of 66.7 µg mL-1. In addition, compound 3 showed weak inhibition of the mammalian cell line KB3.1 (human endocervical adenocarcinoma) with a half maximal inhibitory concentration of 21.2 µM. The neurotrophic activities of 15-hydroxyisodrimenin (1) and 10-carboxy-10-norisodrimenin (5) were assed in neurite outgrowth and real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) assays. When supplemented with 5 ng mL-1 nerve growth factor (NGF), the drimanes 1 and 5 induced neurite outgrowth in PC-12 (rat pheochromocytoma) cells compared to cells solely treated with NGF. As evaluated by RT-qPCR, compounds 1 and 5 also increased NGF and brain-derived neurotrophic factor expression levels in 1321N1 astrocytoma cells. Interestingly, the current study only represents the second report on neurotrophic activities of this widespread class of terpenoids. The only other available study deals with Cyathus africanus, another basidiomycete that can produce drimanes and cyathanes, but is only distantly related to Dentipellis and the Hericiaceae.

Inhibitory Effects of the Fungal Pigment Rubiginosin C on Hyphal and Biofilm Formation in Candida albicans and Candida auris.

The two fungal human pathogens, Candida auris and Candida albicans, possess a variety of virulence mechanisms. Among them are the formation of biofilms to protect yeast against harsh conditions through the development of (pseudo)hyphae whilst also facilitating the invasion of host tissues. In recent years, increased rates of antifungal resistance have been associated with C. albicans and C. auris, posing a significant challenge for the effective treatment of fungal infections. In the course of our ongoing search for novel anti-infectives, six selected azaphilones were tested for their cytotoxicity and antimicrobial effects as well as for their inhibitory activity against biofilm and hyphal formation. This study revealed that rubiginosin C, derived from stromata of the ascomycete Hypoxylon rubiginosum, effectively inhibited the formation of biofilms, pseudohyphae, and hyphae in both C. auris and C. albicans without lethal effects. Crystal violet staining assays were utilized to assess the inhibition of biofilm formation, while complementary microscopic techniques, such as confocal laser scanning microscopy, scanning electron microscopy, and optical microscopy, were used to investigate the underlying mechanisms. Rubiginosin C is one of the few substances known to effectively target both biofilm formation and the yeast-to-hyphae transition of C. albicans and C. auris within a concentration range not affecting host cells, making it a promising candidate for therapeutic intervention in the future.

Design of non-cytotoxic 6,7-dihydroxycoumarin-5-carboxylates with antibiofilm activity against Staphylococcus aureus and Candida albicans.

The 6,7-dihydroxycoumarin-5-carboxylates DHCou and 4-Me-DHCou have been synthesized via five-step route including a propargyl-Claisen rearrangement as key step. The compounds show antibiofilm activity against Stapylococcus aureus and Candida albicans but lack the cytotoxic activity of parent 6,7-dihydroxycoumarines such as esculetin and 4-methylesculetin.

Syntheses and Biofilm Reducing Effects of l-Dopa-Derived Analogues of the Fungal Macrocidins A and Z.

Four analogues of the fungal metabolites macrocidin A and Z, featuring [13]para- or [13]metacyclophanes, were synthesised from fully and orthogonally protected l-dopa instead of l-tyrosine. They were tested for antibiotic activities and for effects on the growth and persistence of microbial biofilms. Tentative structure-activity relationships and distinct differences when compared with the natural lead compounds were identified.

Alliacane-Type Secondary Metabolites from Submerged Cultures of the Basidiomycete Clitocybe nebularis.

Seven sesquiterpenoids, named nebucanes A-G (1-7), featuring a rare alliacane scaffold with unprecedented furan or pyrrole functions, were isolated from the fermentation broth of Clitocybe nebularis. Their structures were established on the basis of 1D/2D NMR spectroscopic analyses, HR-(+)-ESIMS spectra, and comparison of measured and calculated CD spectra for determination of the absolute configuration. Assessing the biological activities, nebucane D (4) exhibited antifungal effects against Rhodotorula glutinis, while nebucane G (7) displayed significant cytotoxicity against MCF-7 and A431 cell lines.

Antimicrobial and Cytotoxic Cyathane-Xylosides from Cultures of the Basidiomycete Dentipellis fragilis.

In our continued search for biologically active metabolites from cultures of rare Basidiomycota species, we found eight previously undescribed cyathane-xylosides from submerged cultures of Dentipellis fragilis, which were named dentifragilins A-H. In addition, the known cyathane derivatives striatal D and laxitextine A were isolated. All compounds were characterized by high-resolution electrospray ionization mass spectrometry (HR-ESIMS) as well as by 1D and 2D nuclear magnetic resonance (NMR) spectroscopy. Several of the compounds exhibited significant activities in standardized cell-based assays for the determination of antimicrobial and cytotoxic effects. The discovery of cyathanes in the genus Dentipellis has chemotaxonomic implications, as this class of diterpenoids has already been shown to be characteristic for mycelial cultures of the related genera Hericium and Laxitextum, which are classified as Dentipellis in the family Hericiaceae.

Synthetic Approach to the Natural N-Nitrosohydroxylamino Tetramic Acid JBIR-141.

An analogue of the Streptomyces metabolite JBIR-141, featuring a delicate N-nitrosohydroxylamine, a 3-acyltetramic acid, and an oxazoline, was synthesized by a convergent strategy from l-alanine, l-threonine, and l-glutamic acid. Key steps were the cyclization of an Ala-Thr derivative to give the oxazoline, a Dieckmann condensation affording the 3-acyltetramic acid, and the N-nitrosation of a hydroxylamino derivative of glutamic acid. An adequate protecting group strategy was established for coupling the three building blocks.

Syntheses and biological effects of natural Morinda lactone and derivatives.

2-Caffeoyl-3-ketohexulofuranosonic acid γ-lactone (morinda lactone; 1a), a natural constituent of Morinda citrifolia L. (Rubiaceae), and eight derivatives with variance in the aryl residue were synthesised by Tsuji-Trost allylation of vitamin C acetonide with the respective aryl allyl alcohol. They were screened for antibiotic activities and for effects on the growth and persistence of microbial biofilms. Some derivatives were active against biofilms of S. aureus or C. albicans at concentrations not toxic to these microorganisms or mammalian cells.

Synthesis and Bioactivity of Ophiofuranones A and B.

Ophiofuranones A and B, metabolites of the fungus Ophiosphaerella korrae, were synthesized in 16 steps and 12%/22% yield. The stereogenic centers were established by Sharpless dihydroxylations and epoxidation, the 1,3-dienes via Wittig or HWE olefinations. The rings were closed through Knoevenagel-type condensation and lactonization. The ophiofuranones proved nontoxic at relevant concentrations against tumor cells, fibroblasts, and various bacteria and fungi. Ophiofuranone A and the monocyclic precursors 4 were weakly active against microbial biofilms.

Synthesis and Bioactivity of Ancorinoside B, a Marine Diglycosyl Tetramic Acid.

The sponge metabolite ancorinoside B was prepared for the first time in 16 steps and 4% yield. It features a ß-d-galactopyranosyl-(1→4)-ß-d-glucuronic acid tethered to a d-aspartic acid-derived tetramic acid. Key steps were the synthesis of a fully protected d-lactose derived thioglycoside, its attachment to a C20-aldehyde spacer, functionalization of the latter with a terminal N-(ß-ketoacyl)-d-aspartate, and a basic Dieckmann cyclization to close the pyrrolidin-2,4-dione ring with concomitant global deprotection. Ancorinoside B exhibited multiple biological effects of medicinal interest. It inhibited the secretion of the cancer metastasis-relevant matrix metalloproteinases MMP-2 and MMP-9, and also the growth of Staphylococcus aureus biofilms by ca 87% when applied at concentrations as low as 0.5 µg/mL. This concentration is far below its MIC of ca 67 µg/mL and thus unlikely to induce bacterial resistance. It also led to a 67% dispersion of preformed S. aureus biofilms when applied at a concentration of ca 2 µg/mL. Ancorinoside B might thus be an interesting candidate for the control of the general hospital, catheter, or joint protheses infections.

Total Synthesis via Biomimetic Late-Stage Heterocyclization: Assignment of the Relative Configuration and Biological Evaluation of the Nitraria Alkaloid (±)-Nitrabirine.

The racemic total synthesis of nitrabirine (5) together with its previously undescribed epimer 2-epi nitrabirine (5') is accomplished via a six-step route based on a biomimetic late-stage heterocyclization. This allowed the assignment of the relative configuration of nitrabirine by the lanthanide-induced shifts (LIS) experiment, which was later on confirmed by X-ray diffraction of obtained single crystals. Furthermore, oxidation studies demonstrated that the direct N-oxidation of nitrabirine does not yield nitrabirine N-oxide as reported earlier. In contrast, the reaction of hydrogen peroxide with nitrabirine (5) yields the salt 24', whereas 2-epi nitrabirine (5') surprisingly leads to a previously uncharacterized product 22 under the same conditions. Finally, a Fischer indole reaction gave access to novel tetracyclic nitrabirine derivatives 26a-d. A comprehensive biological evaluation of nitrabirine (5), 2-epi nitrabirine (5'), and all derivatives synthesized in this study revealed general biofilm dispersal effects against Candida albicans. Moreover, specific compounds showed moderate antibacterial activities as well as potent cytotoxic activities.

Dual Agents: Fungal Macrocidins and Synthetic Analogues with Herbicidal and Antibiofilm Activities.

Eight analogues of the bioherbicides macrocidin A (1) and Z (2) with structural variance in the size of the macrocycle, its para- or meta-cyclophane character, and its functional groups were synthesized on two modular routes and tested for herbicidal, antibiotic, and antibiofilm activities. Apart from the lead compounds 1 and 2, the structurally simplified dihydromacrocidin Z (3) and normacrocidin Z (4) showed high herbicidal activity in either thistles, dandelions or in both. The derivatives 2, 3, and dibromide 9 also inhibited the growth of Staphylococcus aureus biofilms by ca 70% when applied at subtoxic concentrations as low as ca 20 µM, which are unlikely to induce bacterial resistance. They also led to the dispersion of preformed biofilms of S. aureus, exceeding a similar effect by microporenic acid A, a known biofilm inhibitor. Compounds 3 and 9 showed no noticeable cytotoxicity against human cancer and endothelial cells at concentrations below 50 µM, making them conceivable candidates for application as anti-biofilm agents in a medicinal context.