Retrobisabolane A, a Novel Bisabolane-Derived Sesquiterpenoid Isolated from Deep-Sea-Derived Fungus Retroconis fusiformis MCCC 3A00792.

One novel bisabolane-derived sesquiterpenoid retrobisabolane A (1), featuring a methyl group location at the C-4 position instead of C-3 in the bisabolanes, and a known ester-substituted eremophilane-type sesquiterpenoid cryptosphaerolide (2), along with three known indole alkaloids (3-5) were discovered from the fermented cultures of a deep-sea-derived fungus Retroconis fusiformis MCCC 3A00792. The planar structure of new compound 1 was determined by extensive analysis of the NMR and HRESIMS spectra. The relative and absolute configurations of 1 were resolved by the coupling constant (J), calculation of ECD and NMR spectra, and the DP4+ probability analysis of the 1H and 13C NMR data. Interestingly, retrobisabolane A was the new subclass of bisabolanes bearing a methyl group linkage at C-4 instead of C-3 position. Three human cancer cell lines (Hela, AGS, and BIU-87) were subjected to evaluate the cytotoxic activities of compounds 1-5. As a result, compound 2 exhibited significant inhibitory activities against three cell lines with IC50 values ranging from 9.95 to 18.77 µM.

Phaeosphamides A and B, Cytotoxic Cyclodecadepsipeptides from the Mangrove-Derived Fungus Phaeosphaeriopsis sp. S296.

Chemical examination of the fermented broth of the mangrove-derived fungus Phaeosphaeriopsis sp. S296 resulted in the isolation of two new cyclodecadepsipeptides, namely phaeosphamides A (1) and B (2), as well as one known congener Sch 217048 (3). The structures of new metabolites, including absolute configurations, were established on the basis of extensive spectroscopic data analyses, chemical conversion, and Marfey's method. The 2-hydroxy-3-methylpentanoic acid (Hmp) moiety and pipecolic acid (Pip) unit in structures were rarely discovered in nature. Interestingly, compounds 1-3 are examples of peptides discovered from the fungal genus Phaeosphaeriopsis for the first time. All identified compounds were evaluated for their cytotoxicity against five tumor cell lines of AGS, BEL-7402, HepG2, B16, and BIU87. Among them, compound 1 showed inhibitory activities against these tumor cell lines with IC50 values ranging from 5.14 to 66.38 µM. A further mechanistic investigation found that 1 arrested AGS cells in the G2 phase and induced their apoptosis in a dose-dependent manner.

Antiviral Cyclopropane Acids from Deep-Sea-Derived Fungus Aspergillus sydowii.

Four novel monocyclic cyclopropane acids, namely, sydocyclopropanes A-D (1-4), along with one known congener hamavellone B (5), were isolated from the Aspergillus sydowii MCCC 3A00324 fungus, which was isolated from the deep-sea sediment. The gross structures of novel compounds were established by detailed analyses of the spectroscopic data (HRESIMS and NMR spectra), and their absolute configurations were resolved on the basis of the quantum chemical calculations of ECD and NMR data, in association with DP4+ probability analyses. Sydocyclopropanes A-D, featuring the 1,1,2,3-tetrasubstituted cyclopropane nucleus with different lengthy alkyl side chains, were discovered in nature for the first time. All compounds exhibited antiviral activities against A/WSN/33 (H1N1), with IC50 values ranging from 26.7 to 77.2 µM, of which compound 1 exhibited a moderate inhibitory effect (IC50 = 26.7 µM).

Ion Mobility-Derived Collision Cross-Sections Add Extra Capability in Distinguishing Isomers and Compounds with Similar Retention Times: The Case of Aphidicolanes.

The hyphenation of ion mobility spectrometry with high-resolution mass spectrometry has been widely used in the characterization of various metabolites. Nevertheless, such a powerful tool remains largely unexplored in natural products research, possibly mainly due to the lack of available compounds. To evaluate the ability of collision cross-sections (CCSs) in characterizing compounds, especially isomeric natural products, here we measured and compared the traveling-wave IMS-derived nitrogen CCS values for 75 marine-derived aphidicolanes. We established a CCS database for these compounds which contained 227 CCS values of different adducts. When comparing the CCS differences, 36 of 57 pairs (over 60%) of chromatographically neighboring compounds showed a ΔCCS over 2%. What is more, 64 of 104 isomeric pairs (over 60%) of aphidicolanes can be distinguished by their CCS values, and 13 of 18 pairs (over 70%) of chromatographically indistinguishable isomers can be differentiated from the mobility dimension. Our results strongly supported CCS as an important parameter with good orthogonality and complementarity with retention time. CCS is expected to play an important role in distinguishing complex and diverse marine natural products.

Phenolic Metabolites from a Deep-Sea-Derived Fungus Aspergillus puniceus A2 and Their Nrf2-Dependent Anti-Inflammatory Effects.

Four undescribed phenolic compounds, namely asperpropanols A-D (1-4), along with two known congeners 5 and 6, were isolated from Aspergillus puniceus A2, a deep-sea-derived fungus. The gross structures of the compounds were established by detailed analyses of the HRESIMS and NMR data, and their absolute configurations were resolved by modified Mosher's method and calculations of ECD data. Compounds 1-6 were found to have excellent anti-inflammatory effect on lipopolysaccharide (LPS)-induced RAW264.7 cells at 20 µM, evidenced by the reduced nitric oxide (NO), tumor necrosis factor α, and interleukin 6 production. Among them, 5 and 6 showed inhibitory effects on NO production comparable with the positive control (BAY11-7083 at 10 µM). Additionally, the LPS-induced mRNA expressions of inducible nitric oxide synthase and cyclooxygenase-2 were also decreased. Interestingly, mRNA expression of nuclear factor erythroid 2-related factor 2 (Nrf2) was downregulated by LPS and recovered by 1-6, suggesting a vital role of Nrf2 in their effect. We further found that pharmacological inhibition of Nrf2 by ML385 largely abrogated the effects of 1-6 on RAW264.7 cells. Therefore, 1-6 may share a common anti-inflammatory mechanism via Nrf2 upregulation and activation.

Bisabolane Sesquiterpenes with Anti-Inflammatory Activities from the Endophytic Fungus Penicillium citrinum DF47.

Seven new bisabolane-type sesquiterpenes (1-7), namely penicibisabolanes A-G, together with eight known analogs (8-15) were obtained from the AcOEt extract of the millet fermentation broth of the endophytic fungus Penicillium citrinum DF47, which was isolated from the fresh root of Codonopsis pilosula (Franch.) Nannf. The gross structures of new metabolites were determined on the basis of the spectroscopic data (HR-ESI-MS, 1D and 2D NMR spectra), while their absolute configurations were resolved by comparison of the experimental and calculated ECD spectra, in association with specific rotation data. Compound 1 is a rare seco-trinor-bisabolane sesquiterpene found in nature, while 3 is the first example of phenolic bisabolanes bearing a methoxy group at C-1. All the isolates were evaluated their inhibitory effects against NO production in lipopolysaccharides (LPS) stimulated RAW264.7 cells. Among them, compounds 7 and 13 showed moderately anti-inflammatory effects with the inhibitory rate more than 50 % at the concentration of 20 µM.

Complete genome sequence of Streptomyces sp. HSG2 from rhizosphere soil of mangrove in Qingmei Gang, Sanya.

Streptomyces sp. HSG2 was isolated from rhizosphere soil of a mangrove forest sample at Qingmei Gang, Sanya. The complete genome sequence of the strain HSG2 was obtained using PacBio Sequel HGAP.4 and comprised of 5,282,528 base pairs with a 71.9 mol% G + C content, 4504 protein-coding genes, and 71 RNAs. An in-silico analysis confirmed that genes associated with polysaccharide hydrolyzation, hydrocarbon degradation, and aerobic denitrification were presented in the genome. We also identified 24 natural product biosynthetic gene clusters for secondary metabolites, including those for streptobactin and nystatin A1. The complete genome sequence indicated that Streptomyces sp. HSG2 will provide insight into the biosynthesis and regulatory mechanisms for its secondary metabolites, and propose a potential use in biotechnological and novel bioactive natural product biosynthetic applications.

Epigenetic Manipulation to Trigger Production of Guaiane-Type Sesquiterpenes from a Marine-Derived Spiromastix sp. Fungus with Antineuroinflammatory Effects.

Epigenetic manipulation of a deep-sea sediment-derived Spiromastix sp. fungus using suberoylanilide hydroxamic acid (SAHA) induction resulted in the activation of a terpene-related biosynthetic gene cluster, and nine new guaiane-type sesquiterpenes, spiromaterpenes A-I (1-9), were isolated. Their structures were determined using various spectroscopic techniques, in association with the modified Mosher's method, computed electronic circular dichroism (ECD) spectra, and chemical conversion for configurational assignments. Compounds 4-6 exhibited significant effects against the NO production on lipopolysaccharide (LPS)-induced microglia cells BV2, and the preliminary SAR analyses demonstrated that a 2(R),11-diol unit is favorable. The most active 5 abolished LPS-induced NF-κB translocation from the cytosol to the nucleus in BV-2 microglial cells, accompanied by the marked reduction of the transcription levels of pro-inflammatory cytokines, including IL-1ß, IL-6, and TNF-α dose-dependently in both LPS-induced BV-2 and BV-2 cells, as well as the protein and mRNA levels of iNOS and COX-2. This study complements the gap in knowledge regarding the anti-neuroinflammatory activity of guaiane-type sesquiterpenoids at the cellular level and suggests that 5 is promising for further optimization as a multifunctional agent for antineuroinflammation.

Brevibacterium profundi sp. nov., isolated from deep-sea sediment of the Western Pacific Ocean.

A new Gram-stain-positive, aerobic, non-motile and rod-shaped actinobacterium, designated O1T, was isolated from a deep-sea sediment of the Western Pacific Ocean. Strain O1T showed optimal growth at 30 °C, between pH 6.0 and 8.0, and in the presence of 1-5 % (w/v) NaCl. The predominant menaquinone was MK-8 (H2), and anteiso-C15 : 0 and anteiso-C17 : 0 were the major fatty acids. The diagnostic diamino acid in the cell-wall peptidoglycan was meso-diaminopimelic acid. The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol and one unknown glycolipid. The DNA G+C content of strain O1T was 64.9 mol% and the genome size was 4.17 Mb. Based on a similarity search and phylogenetic analysis of the 16S rRNA gene sequence, strain O1T belonged to the genus Brevibacterium. The values of average nucleotide identity and in silico DNA-DNA hybridization between strain O1T and its close relatives were well below the thresholds used for the delineation of a new species. On the basis of the morphological and chemotaxonomic characteristics, as well as the genotypic data, it is proposed that strain O1T represents a novel species of the genus Brevibacterium, for which the name Brevibacterium profundi sp. nov. is proposed. The type strain is O1T (=JCM 33845T=MCCC 1A16744T).

Phenolic bisabolane and cuparene sesquiterpenoids with anti-inflammatory activities from the deep-sea-derived Aspergillus sydowii MCCC 3A00324 fungus.

Seventeen undescribed sesquiterpenoids including 14 phenolic bisabolanes, namely asperbisabolanes A-N (1-14), and 3 cuparenes (aspercuparenes A-C, 15-17), together with 10 known bisabolane analogues (18-27) were isolated from the EtOAc extract of fermented cultures of the deep sea sediment-derived fungus Aspergillus sydowii MCCC 3A00324. The new structures were established on the basis of extensive NMR and HRESIMS spectroscopic data analyses, while their absolute configurations were assigned by comparison of the experimental ECD spectra with those of the TDDFT-ECD calculated spectra or reported data in literature. Asperbisabolanes A (1) and B (2) are the first examples of bisabolane sesquiterpenoids featuring a 6/6/6 tricyclic nucleus. Compound 3 possessed a novel seco-bisabolane skeleton with a rare dioxolane ring moiety, while asperbisabolane K (11) represents the first case of bisabolanes bearing a rare methylsulfonyl group. All the isolated compounds (1-27) were evaluated their activities against NO secretion in LPS-activated BV-2 microglia cells. As a result, 6, 12, 16, and 25-27 exhibited the inhibition rate over 45% at a concentration of 10 µM. Moreover, 12 exerted the anti-inflammatory activity by inhibiting the NF-κB-activated pathway in dose-dependent manner.

New Monoterpenoids and Polyketides from the Deep-Sea Sediment-Derived Fungus Aspergillus sydowii MCCC 3A00324.

Chemical study of the secondary metabolites of a deep-sea-derived fungus Aspergillus sydowii MCCC 3A00324 led to the isolation of eleven compounds (1-11), including one novel (1) and one new (2) osmane-related monoterpenoids and two undescribed polyketides (3 and 4). The structures of the metabolites were determined by comprehensive analyses of the NMR and HRESIMS spectra, in association with quantum chemical calculations of the 13C NMR, ECD, and specific rotation data for the configurational assignment. Compound 1 possessed a novel monoterpenoid skeleton, biogenetically probably derived from the osmane-type monoperpenoid after the cyclopentane ring cleavage and oxidation reactions. Additionally, compound 3 was the first example of the α-pyrone derivatives bearing two phenyl units at C-3 and C-5, respectively. The anti-inflammatory activities of 1-11 were tested. As a result, compound 6 showed potent inhibitory nitric oxide production in lipopolysaccharide (LPS)-activated BV-2 microglia cells with an inhibition rate of 94.4% at the concentration of 10 µM. In addition, a plausible biosynthetic pathway for 1 and 2 was also proposed.

Synthesis and Biological Evaluation of Analogues of Butyrolactone I as PTP1B Inhibitors.

In recent years, a large number of pharmacologically active compounds containing a butenolide functional group have been isolated from secondary metabolites of marine microorganisms. Butyrolactone I was found to be produced by Aspergillus terreus isolated from several marine-derived samples. The hypoglycemic activity of butyrolactone I has aroused our great interest. In this study, we synthesized six racemic butenolide derivatives (namely BL-1-BL-6) by modifying the C-4 side chain of butyrolactone I. Among them, BL-3 and BL-5 improved the insulin resistance of HepG2 cells and did not affect the proliferation of RIN-m5f cell line, which indicated the efficacy and safety of BL-3 and BL-5. Furthermore, BL-3, BL-4, BL-5, and BL-6 displayed a significant protein tyrosine phosphatase 1B (PTP1B) inhibitory effect, while the enantiomers of BL-3 displayed different 50% percentage inhibition concentration (IC50) values against PTP1B. The results of molecular docking simulation of the BLs and PTP1B explained the differences of biological consequences observed between the enantiomers of BL-3, which supported BLs as PTP1B inhibitors, and also indicated that the chirality of C-4 might influence the inhibitory effect of the BLs. Our findings provide a novel strategy for the development of butyrolactone derivatives as potential PTP1B inhibitors for the treatment of type 2 diabetes mellitus.

Aphidicolin Chemistry of the Deep-Sea-Derived Fungus Botryotinia fuckeliana MCCC 3A00494.

Aphidicolin, a potent DNA polymerase α inhibitor, has been explored in clinical trials for the treatment of cancer. So far, about 300 modified aphidicolins have been discovered. However, none have shown a stronger effect. Herein, we report 71 new (aphidicolins A1-A71, 1-71) and eight known (72-79) aphidicolin congeners from Botryotinia fuckeliana MCCC 3A00494, a fungus isolated from the western Pacific Ocean (-5572 m). The structures of 1-71 were determined through extensive spectroscopic analysis, X-ray crystallography, chemical derivatization, modified Mosher's method, and the ECD exciton chirality method. Compounds 54-57 and 58-64 are novel 6/6/5/6/5 pentacyclic aphidicolins featuring tetrahydrofuran and dihydrofuran rings, respectively, while compounds 65-71 are rare noraphidicolins. Aphidicolin A8 (8) significantly induced apoptosis in T24 (IC50 = 2.5 µM) and HL-60 (IC50 = 6.1 µM) cancer cells by causing DNA damage. By docking its structure to the human DNA polymerase α binding pocket, 8 was found to form tight intermolecular contacts, elaborating aphidicolin A8 as a potently cytotoxic lead compound.

Fusarisolins A⁻E, Polyketides from the Marine-Derived Fungus Fusarium solani H918.

Five new (fusarisolins A⁻E, 1 to 5) and three known (6 to 8) polyketides were isolated from the marine-derived fungus Fusarium solani H918, along with six known phenolics (9 to 14). Their structures were established by comprehensive spectroscopic data analyses, methoxyphenylacetic acid (MPA) method, chemical conversion, and by comparison with data reported in the literature. Compounds 1 and 2 are the first two naturally occurring 21 carbons polyketides featuring a rare ß- and γ-lactone unit, respectively. All isolates (1 to 14) were evaluated for their inhibitory effects against tea pathogenic fungus Pestalotiopsis theae and 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) synthase gene expression. Compound 8 showed potent antifungal activity with an ED50 value of 55 µM, while 1, 8, 13, and 14 significantly inhibited HMG-CoA synthase gene expression.

Cytotoxic Polyketides Isolated from the Deep-Sea-Derived Fungus Penicillium chrysogenum MCCC 3A00292.

The chemical examination of the solid cultures of the deep-sea-derived fungus Penicillium chrysogenum MCCC 3A00292 resulted in the isolation of three new versiol-type analogues, namely peniciversiols A-C (1-3), and two novel lactone derivatives, namely penicilactones A and B (6 and 7), along with 11 known polyketides. The planar structures of the new compounds were determined by the comprehensive analyses of the high-resolution electrospray ionization mass spectroscopy (HRESIMS) and nuclear magnetic resonance (NMR) data, while their absolute configurations were resolved on the basis of comparisons of the experimental electronic circular dichroism (ECD) spectra with the calculated ECD data. Compound 1 is the second example of versiols featuring a 2,3-dihydropyran-4-one ring. Additionally, compounds 6 and 7 are the first representatives of γ-lactone derivatives constructed by a 1,3-dihydroxy-5-methylbenzene unit esterifying with the α-methyl-γ-hydroxy-γ-acetic acid α,ß-unsaturated-γ-lactone moiety and α-hydroxy-γ-methyl-γ-acetic acid α,ß-unsaturated-γ-lactone unit, respectively. All of the isolated compounds were evaluated for their cytotoxic activities against five human cancer cell lines of BIU-87, ECA109, BEL-7402, PANC-1, and Hela-S3. Compound 1 exhibited a selective inhibitory effect against the BIU-87 cell line (IC50 = 10.21 µM), while compounds 4, 5, 8, and 12-16 showed inhibitory activities against the ECA109, BIU-87, and BEL-7402 cell lines with the IC50 values ranging from 7.70 to > 20 µM.

A New Pimarane Diterpenoid from the Botryotinia fuckeliana Fungus Isolated from Deep-Sea Water.

A new pimarane diterpenoid, named botryopimarene A (1), was discovered from the fungus Botryotinia fuckeliana MCCC 3 A00494 isolated from the deep-sea water, together with ten known compounds. The planar structure of 1 was established based on the extensive spectroscopic analyses. The absolute configurations of tricyclic system in 1 were resolved by the theoretical ECD calculation, while the 15,16-diol moiety in the side chain was resolved by the Mo2 (OAc)4 -induced ECD spectrum. Compound 1, featuring a Δ9(11) double bond, was rarely discovered in pimarane family. Compounds 1-11 were tested for their cytotoxic activities using six human cancer cell lines by the MTT method. However, none of the compounds exhibited detectable cytotoxicities (IC50 >20 µm).

Graphostromols A-K, Eleven New Chained Polyketides from the Deep-Sea-Derived Graphostroma sp.

Chromatographic separation of the AcOEt extract of the fermented cultures of the deep-sea-derived fungus Graphostroma sp. resulted in the isolation of 11 new chained polyketides, namely graphostromols A-K (1-11). The structures of the new compounds were established by the extensive analyses of their NMR and HR-ESI-MS data. The absolute configuration at C-11 in 1 was determined on the basis of the modified Mosher's method. Compounds 1-5, bearing a 2,2,10,10-tetramethyldodecane skeleton, were discovered for the first time from nature. All isolates were evaluated for their cytotoxicities against five different cancer cell lines (HeLa, Eca-109, Biu-87, Bel-7402, and PANC-1). However, none showed positive effects.

Spirograterpene A, a Tetracyclic Spiro-Diterpene with a Fused 5/5/5/5 Ring System from the Deep-Sea-Derived Fungus Penicillium granulatum MCCC 3A00475.

A novel spiro-tetracyclic diterpene, spirograterpene A (1), was isolated from the deep-sea-derived fungus Penicillium granulatum MCCC 3A00475, together with two biosynthetically related cyclopianes, conidiogenone I (2) and conidiogenone C (3). The relative configuration of 1 was elucidated by extensive spectroscopic analyses, and the absolute structure was established by the modified Mosher's method. Compound 1 is the second example of a diterpene featuring a 5/5/5/5 spirocyclic carbon skeleton. It showed modest antiallergic activity.

Microindolinone A, a Novel 4,5,6,7-Tetrahydroindole, from the Deep-Sea-Derived Actinomycete Microbacterium sp. MCCC 1A11207.

A novel indole, microindolinone A (1), was isolated from a deep-sea-derived actinomycete Microbacterium sp. MCCC 1A11207, together with 18 known compounds (2-19). By detailed analysis of the ¹H, 13C, HSQC, COSY, HMBC, high resolution electron spray ionization mass spectrum (HRESIMS), and circular dichroism (CD) data, the absolute configuration of 1 was elucidated as 5R-hydroxy-4,5,6,7-tetrahydroindole-4-one. It is noteworthy that 1 is the second example of a saturated indole isolated from nature.

New polyphenols from a deep sea Spiromastix sp. Fungus, and their antibacterial activities.

Eleven new polyphenols namely spiromastols A-K (1-11) were isolated from the fermentation broth of a deep sea-derived fungus Spiromastix sp. MCCC 3A00308. Their structures were determined by extensive NMR data and mass spectroscopic analysis in association with chemical conversion. The structures are classified as diphenyl ethers, diphenyl esters and isocoumarin derivatives, while the n-propyl group in the analogues is rarely found in natural products. Compounds 1-3 exhibited potent inhibitory effects against a panel of bacterial strains, including Xanthomanes vesicatoria, Pseudomonas lachrymans, Agrobacterium tumefaciens, Ralstonia solanacearum, Bacillus thuringensis, Staphylococcus aureus and Bacillus subtilis, with minimal inhibitory concentration (MIC) values ranging from 0.25 to 4 µg/mL. The structure-activity relationships are discussed, while the polychlorinated analogues 1-3 are assumed to be a promising structural model for further development as antibacterial agents.