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As a part of the important species that form coral reef ecosystems, stony corals have become a potential source of pharmacologically active lead compounds for an increasing number of compounds with novel chemical structures and strong biological activity. In this study, the secondary metabolites and biological activities are reported for Aspergillus terreus C21-1, an epiphytic fungus acquired from Porites pukoensis collected from Xuwen Coral Reef Nature Reserve, China. This strain was cultured in potato dextrose broth (PDB) media and rice media with different salinities based on the OSMAC strategy. The mycelial morphology and high-performance thin layer chromatographic (HPTLC) fingerprints of the fermentation extracts together with bioautography were recorded. Furthermore, an untargeted metabolomics study was performed using principal component analysis (PCA), orthogonal projection to latent structure discriminant analysis (O-PLSDA), and feature-based molecular networking (FBMN) to analyze their secondary metabolite variations. The comprehensive results revealed that the metabolite expression in A. terreus C21-1 differed significantly between liquid and solid media. The metabolites produced in liquid medium were more diverse but less numerous compared to those in solid medium. Meanwhile, the mycelial morphology underwent significant changes with increasing salinity under PDB cultivation conditions, especially in PDB with 10% salinity. Untargeted metabolomics revealed significant differences between PDB with 10% salinity and other media, as well as between liquid and solid media. FBMN analysis indicated that alkaloids, which might be produced under high salt stress, contributed largely to the differences. The biological activities results showed that six groups of crude extracts exhibited acetylcholinesterase (AChE) inhibitory activities, along with 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging and antibacterial activities. The results of this study showed that the increase in salinity favored the production of unique alkaloid compounds by A. terreus C21-1.

Neuroinflammation is one of the main mechanisms involved in the progression of neurodegenerative diseases (NDs), and microglial activation is the main feature of neuroinflammation. Polaprezinc (Pol), a chelator of L-carnosine and zinc, is widely used as a clinical drug for gastric ulcers. However, its potential effects on NDs remain unexplored. In LPS-induced BV-2 microglia, we found that Pol reduced the generation of NO and ROS and revealed inhibited expression of iNOS, COX-2, and inflammatory factors such as IL-6, TNF-α, and 1L-1β by Pol using qRT-PCR and Western blotting. These effects were found to be associated with the suppression of the NF-κB signaling pathway. Moreover, we evaluated the potential synergistic effects of aspergillusidone G (Asp G) when combined with Pol. Remarkably, co-treatment with low doses of Asp G enhanced the NO inhibition by Pol from approximately 30% to 80% in LPS-induced BV2 microglia, indicating a synergistic anti-inflammatory effect. A bioinformatics analysis suggested that the synergistic mechanism of Asp G and Pol might be attributed to several targets, including NFκB1, NRF2, ABL1, TLR4, and PPARα. These findings highlight the anti-neuroinflammatory properties of Pol and its enhanced efficacy when combined with Asp G, proposing a novel therapeutic strategy for managing neuroinflammation in NDs.

Chemical investigation of marine fungus Nigrospora oryzae SYSU-MS0024 cultured on solid-rice medium led to the isolation of three new alkaloids, including a pair of epimers, nigrosporines A (1) and B (2), and a pair of enantiomers, (+)-nigrosporine C (+)-3, and (−)-nigrosporine C (−)-3, together with eight known compounds (4–11). Their structures were elucidated based on extensive mass spectrometry (MS) and 1D/2D nuclear magnetic resonance (NMR) spectroscopic analyses and compared with data in the literature. The absolute configurations of compounds 1–3 were determined by a combination of electronic circular dichroism (ECD) calculations, Mosher’s method, and X-ray single-crystal diffraction technique using Cu Kα radiation. In bioassays, compound 2 exhibited moderate inhibition on NO accumulation induced by lipopolysaccharide (LPS) on BV-2 cells in a dose-dependent manner at 20, 50, and 100 μmol/L and without cytotoxicity in a concentration of 100.0 μmol/L. Moreover, compound 2 also showed moderate acetylcholinesterase (AChE) inhibitory activities with IC50 values of 103.7 μmol/L. Compound 5 exhibited moderate antioxidant activity with EC50 values of 167.0 μmol/L.

BackgroundAlkaloids are the second primary class of secondary metabolites (SMs) from marine organisms, most of which have antioxidant, antitumor, antibacterial, anti-inflammatory, and other activities. However, the SMs obtained by traditional isolation strategies have drawbacks such as highly reduplication and weak bioactivity. Therefore, it is significantly important to establish an efficient strategy for screening strains and mining novel compounds.MethodsIn this study, we utilized in situ colony assay combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify the strain with high potential in alkaloids production. The strain was identified by genetic marker genes and morphological analysis. The secondary metabolites from the strain were isolated by the combine use of vacuum liquid chromatography (VLC), ODS column chromatography, and Sephadex LH-20. Their structures were elucidated by 1D/2D NMR, HR-ESI-MS, and other spectroscopic technologies. Finally, these compounds bioactivity were assay, including anti-inflammatory and anti-β aggregation.ResultsEighteen marine fungi were preliminarily screened for alkaloids production by in situ colony assay using Dragendorff reagent as dye, and nine of them turned orange, which indicated abundant alkaloids. By thin-layer chromatography (TLC), LC-MS/MS, and multiple approaches assisted Feature-Based Molecular Networking (FBMN) analysis of fermentation extracts, a strain ACD-5 (Penicillium mallochii with GenBank accession number OM368350) from sea cucumber gut was selected for its diverse alkaloids profiles especially azaphilones. In bioassays, the crude extracts of ACD-5 in Czapek–dox broth and brown rice medium showed moderate antioxidant, acetylcholinesterase inhibitory, anti-neuroinflammatory, and anti-β aggregation activities. Three chlorinated azaphilone alkaloids, compounds 1–3 (sclerotioramine, isochromophilone VI, and isochromophilone IX, respectively), were isolated from the fermentation products of ACD-5 in brown rice medium guided by bioactivities and mass spectrometry analysis. Compound 1 had shown remarkable anti-neuroinflammatory activity in liposaccharide induced BV-2 cells.ConclusionIn summary, in situ colony screening together with LC-MS/MS, multi-approach assisted FBMN can act as an efficient screening method for strains with potential in alkaloids production.

Abstract Background Mounting evidences indicate that oxidative stress, neuroinflammation, and dysregulation of gut microbiota are related to neurodegenerative disorders (NDs). Butyrolactone I (BTL-I), a marine fungal metabolite, was previously reported as an in vitro neuroprotectant and inflammation inhibitor. However, little is known regarding its in vivo effects, whereas zebrafish (Danio rerio) could be used as a convenient in vivo model of toxicology and central nervous system (CNS) diseases. Methods Here, we employed in vivo and in silico methods to investigate the anti-NDs potential of BTL-I. Specifically, we established a cognitive deficit model in zebrafish by intraperitoneal (i.p.) injection of aluminum trichloride (AlCl3) (21 μg) and assessed their behaviors in the T-maze test. The proinflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) as well as acetylcholinesterase (AChE) activity or glutathione (GSH) levels were assayed 24 h after AlCl3 injection. The intestinal flora variation of the zebrafish was investigated by 16S rDNA high-throughput analysis. The marine fungal metabolite, butyrolactone I (BTL-I), was used to modulate zebrafish cognitive deficits evoked by AlCl3 and evaluated about its effects on the above inflammatory, cholinergic, oxidative stress, and gut floral indicators. Furthermore, the absorption, distribution, metabolism, excretion, and toxicity (ADMET) and drug-likeness properties of BTL-I were studied by the in silico tool ADMETlab. Results BTL-I dose-dependently ameliorated AlCl3-induced cognitive deficits in zebrafish. While AlCl3 treatment elevated the levels of central and peripheral proinflammatory cytokines, increased AChE activity, and lowered GSH in the brains of zebrafish, these effects, except GSH reduction, were reversed by 25–100 mg/kg BTL-I administration. Besides, 16S rDNA high-throughput sequencing of the intestinal flora of zebrafish showed that AlCl3 decreased Gram-positive bacteria and increased proinflammatory Gram-negative bacteria, while BTL-I contributed to maintaining the predominance of beneficial Gram-positive bacteria. Moreover, the in silico analysis indicated that BTL-I exhibits acceptable drug-likeness and ADMET profiles. Conclusions The present findings suggest that BTL-I is a potential therapeutic agent for preventing CNS deficits caused by inflammation, neurotoxicity, and gut flora imbalance.

The prevalence of gout and the adverse effects of current synthetic anti-gout drugs call for new natural and effective xanthine oxidase (XOD) inhibitors to target this disease. Based on our previous finding that an edible seaweed Pterocladiella capillacea extract inhibits XOD, XOD-inhibitory and anti-inflammatory activities were used to evaluate the anti-gout potential of different P. capillacea extract fractions. Through affinity ultrafiltration coupled with liquid chromatography tandem mass spectrometry (LC-MS/MS), feature-based molecular networking (FBMN), and database mining of multiple natural products, the extract’s bioactive components were traced and annotated. Through molecular docking and ADMET analysis, the possibility and drug-likeness of the annotated XOD inhibitors were predicted. The results showed that fractions F4, F6, F4-2, and F4-3 exhibited strong XOD inhibition activity, among which F4-3 reached an inhibition ratio of 77.96% ± 4.91% to XOD at a concentration of 0.14 mg/mL. In addition, the P. capillacea extract and fractions also displayed anti-inflammatory activity. Affinity ultrafiltration LC-MS/MS analysis and molecular networking showed that out of the 20 annotated compounds, 8 compounds have been previously directly or indirectly reported from seaweeds, and 4 compounds have been reported to exhibit anti-gout activity. Molecular docking and ADMET showed that six seaweed-derived compounds can dock with the XOD activity pocket and follow the Lipinski drug-like rule. These results support the value of further investigating P. capillacea as part of the development of anti-gout drugs or related functional foods.

Alzheimer’s disease (AD), a neurodegenerative disease, is the most common cause of dementia in humans worldwide. Although more in-depth research has been carried out on AD, the therapeutic effect of AD is not as expected, and natural active substances are increasingly sought after by scientists. In the present study, we evaluated two benzaldehydes from a coral-derived Aspergillus terreus strain C23-3, their anti-neuroinflammatory activity in microglia (BV-2), and their neuroprotective activity and mechanisms in hippocampal neuronal cells (HT-22). These include the protein expression of iNOS, COX-2, MAPKs pathways, Tau protein-related pathways, caspases family-related signaling pathways. They also include the levels of TNF-α, IL-6, IL-18 and ROS, as well as the level of mitochondrial oxidative stress and neuronal cell apoptosis. The results showed that both benzaldehydes were effective in reducing the secretion of various inflammatory mediators, as well as pro-inflammatory factors. Among these, benzaldehyde 2 inhibited mitochondrial oxidative stress and blocked neuronal cell apoptosis through Tau protein-related pathways and caspases family-related signaling pathways, thereby inhibiting β-amyloid (Aβ)-induced neurological damage. This study reveals that benzaldehyde 2 has potential as a therapeutic agent for Alzheimer’s disease, and offers a new approach to the high-value use of marine natural products.

Co-culture is known as an efficient way to explore the metabolic potential of fungal strains for new antibiotics and other therapeutic agents that could counter emerging health issues. To study the effect of co-culture on the secondary metabolites and bioactivities of two marine strains, Aspergillus terreus C23-3 and Aspergillus. unguis DLEP2008001, they were co-cultured in live or inactivated forms successively or simultaneously. The mycelial morphology and high-performance thin layer chromatography (HPTLC) including bioautography of the fermentation extracts were recorded. Furthermore, the agar cup-plate method was used to compare the antimicrobial activity of the extracts. Based on the above, liquid chromatography-photodiode array-tandem mass spectrometry (LC-PDA-MS/MS) together with Global Natural Products Social molecular networking (GNPS) and multiple natural products database mining were used to further analyze their secondary metabolite variations. The comprehensive results showed the following trends: (1) The strain first inoculated will strongly inhibit the growth and metabolism of the latter inoculated one; (2) Autoclaved A. unguis exerted a strong inducing effect on later inoculated A. terreus, while the autoclaved A. terreus showed high stability of its metabolites and still potently suppressed the growth and metabolism of A. unguis; (3) When the two strains are inoculated simultaneously, they both grow and produce metabolites; however, the A. terreus seemed to be more strongly induced by live A. unguis and this inducing effect surpassed that of the autoclaved A. unguis. Under some of the conditions, the extracts showed higher antimicrobial activity than the axenic cultures. Totally, A. unguis was negative in response but potent in stimulating its rival while A. terreus had the opposite effect. Fifteen MS detectable and/or UV active peaks showed different yields in co-cultures vs. the corresponding axenic culture. GNPS analysis assisted by multiple natural products databases mining (PubChem, Dictionary of Natural Products, NPASS, etc.) gave reasonable annotations for some of these peaks, including antimicrobial compounds such as unguisin A, lovastatin, and nidulin. However, some of the peaks were correlated with antagonistic properties and remain as possible novel compounds without mass or UV matching hits from any database. It is intriguing that the two strains both synthesize chemical ‘weapons’ for antagonism, and that these are upregulated when needed in competitive co-culture environment. At the same time, compounds not useful in this antagonistic setting are downregulated in their expression. Some of the natural products produced during antagonism are unknown chlorinated metabolites and deserve further study for their antimicrobial properties. In summary, this study disclosed the different responses of two Aspergillus strains in co-culture, revealed their metabolic variation, and displayed new opportunities for antibiotic discovery.