Cytotoxic and Apoptosis-Inducing Activity of Triterpene Glycosides from Holothuria scabra and Cucumaria frondosa against HepG2 Cells

The cytotoxic effects of thirteen triterpene glycosides from Holothuria scabra Jaeger and Cucumaria frondosa Gunnerus (Holothuroidea) against four human cell lines were detected and their cytotoxicity-structure relationships were established. The apoptosis-inducing activity of a more potent glycoside echinoside A (1) in HepG2 cells was further investigated by determining its effect on the morphology, mitochondrial transmembrane potential (Δψ_m) and mRNA expression levels of the apoptosis-related genes. The results showed that the number of glycosyl residues in sugar chains and the side chain of aglycone could affect their cytotoxicity towards tumor cells and selective cytotoxicity. 1 significantly inhibited cell viability and induced apoptosis in HepG2 cells. 1 also markedly decreased the Δψ_m and Bcl-2/Bax mRNA express ratio, and up-regulated the mRNA expression levels of Caspase-3, Caspase-8 and Caspase-9 in HepG2 cells. Therefore, 1 induced apoptosis in HepG2 cells through both intrinsic and extrinsic pathway. These findings could potentially promote the usage of these glycosides as leading compounds for developing new antitumor drugs.     

Alterporriol-Type Dimers from the Mangrove Endophytic Fungus,Alternaria sp. (SK11), and Their MptpB Inhibitions

A new alterporriol-type anthranoid dimer, alterporriol S (1), along with seven known anthraquinone derivatives, (+)-aS-alterporriol C (2), hydroxybostrycin (3), halorosellinia A (4), tetrahydrobostrycin (5), 9α-hydroxydihydrodesoxybostrycin (6), austrocortinin (7) and 6-methylquinizarin (8), were isolated from the culture broth of the mangrove fungus, Alternaria sp. (SK11), from the South China Sea. Their structures and the relative configurations were elucidated using comprehensive spectroscopic methods, including 1D and 2D NMR spectra. The absolute configurations of 1 and the axial configuration of 2 were defined by experimental and theoretical ECD spectroscopy. 1 was identified as the first member of alterporriols consisting of a unique C-10−C-2′ linkage. Atropisomer 2 exhibited strong inhibitory activity against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) with an IC₅₀ value 8.70 μM.     

Phallusiasterols A and B: Two New Sulfated Sterols from the Mediterranean Tunicate Phallusia fumigata and Their Effects as Modulators of the PXR Receptor

Purification of the apolar extracts of the marine ascidian Phallusia fumigata, afforded two new sulfated sterols, phallusiasterols A (1) and B (2). The structures of the new compounds have been elucidated using mass spectrometry and NMR experiments. The effects of phallusiasterols A and B as modulators of pregnane-X-receptor (PXR) have been investigated. These studies revealed that phallusiasterol A induces PXR transactivation in HepG2 cells and stimulates the expression of the PXR target genes CYP3A4 and MDR1 in the same cell line. Molecular docking calculations suggested the theoretical binding mode of phallusiasterol A with hPXR and revealed that phallusiasterol A fitted well in the LBD of PXR.     

Psammaceratin A: A Cytotoxic Psammaplysin Dimer Featuring an Unprecedented (2Z,3Z)-2,3-Bis(aminomethylene)succinamide Backbone from the Red Sea Sponge Pseudoceratina arabica

Bioassay-guided partition of the extract of the Red Sea sponge Pseudoceratina arabica and HPLC purification of the active fraction gave a psammaplysin dimer, psammaceratin A (1), along with psammaplysin A (2). The dimer comprises two units of psammaplysin A (2) connected via the terminal amines with an unprecedented (2Z,3Z)-2,3-bis(aminomethylene)succinamide moiety, and it represents the first dimer to be identified among the psammaplysin family. Data from 1D- and 2D-NMR and HRMS supported the chemical structures of the compounds. Psammaceratin A (1) and psammaplysin A (2) exhibited significant growth inhibition of HCT 116, HeLa, and MBA-MB-231 cells down to 3.1 μM.     

Neritinaceramides A–E,New Ceramides from the Marine Bryozoan Bugula neritina Inhabiting South China Sea and Their Cytotoxicity

Five new ceramides, neritinaceramides A (1), B (2), C (3), D (4) and E (5), together with six known ceramides (6–11), two known alkyl glycerylethers (12 and 13) and a known nucleoside (14), were isolated from marine bryozoan Bugula neritina, which inhabits the South China Sea. The structures of the new compounds were elucidated as (2S,3R,3′S,4E,8E,10E)-2-(hexadecanoylamino)-4,8,10-octadecatriene-l,3,3′-triol (1), (2S,3R,2′R,4E,8E,10E)-2-(hexadecanoylamino)-4,8,10-octadecatriene-l,3,2′-triol (2), (2S,3R,2′R,4E,8E,10E)-2-(octadecanoylamino)-4,8,10-octadecatriene-l,3,2′-triol (3), (2S,3R,3′S,4E,8E)-2-(hexadecanoylamino)-4,8-octadecadiene-l,3,3′-triol (4) and (2S,3R,3′S,4E)-2-(hexadecanoylamino)-4-octadecene-l,3,3′-triol (5) on the basis of extensive spectral analysis and chemical evidences. The characteristic C-3′S hydroxyl group in the fatty acid moiety in compounds 1, 4 and 5, was a novel structural feature of ceramides. The rare 4E,8E,10E-triene structure in the sphingoid base of compounds 1–3, was found from marine bryozoans for the first time. The new ceramides 1–5 were evaluated for their cytotoxicity against HepG2, NCI-H460 and SGC7901 tumor cell lines, and all of them exhibited selective cytotoxicity against HepG2 and SGC7901 cells with a range of IC₅₀ values from 47.3 μM to 58.1 μM. These chemical and cytotoxic studies on the new neritinaceramides A–E (1–5) added to the chemical diversity of B. neritina and expanded our knowledge of the chemical modifications and biological activity of ceramides.     

A New in Vitro Anti-Tumor Polypeptide Isolated from Arca inflata

A new in vitro anti-tumor polypeptide, coded as J2-C3, was isolated from Arca inflata Reeve and purified by diethyl-aminoethanol (DEAE)-sepharose Fast Flow anion exchange and phenyl sepharose CL-4B hydrophobic chromatography. J2-C3 was identified to be a homogeneous compound by native polyacrylamide gel electrophoresis (Native-PAGE). The purity of J2-C3 was over 99% in reversed phase-high performance liquid chromatography (RP-HPLC). The molecular weight was determined as 20,538.0 Da by electrospray-ionization mass spectrometry (ESI-MS/MS). J2-C3 was rich in Glx (Gln + Glu), Lys, and Asx (Asp + Asn) according to amino acid analysis. Four partial amino acid sequences of this peptide were determined as L/ISMEDVEESR, KNGMHSI/LDVNHDGR, AMKI/LI/LNPKKGI/LVPR and AMGAHKPPKGNEL/IGHR via MALDI-TOF/TOF-MS and de novo sequencing. Secondary structural analysis by CD spectroscopy revealed that J2-C3 had the α-helix (45.2%), β-sheet (2.9%), β-turn (26.0%) and random coil (25.9%). The anti-tumor effect of J2-C3 against human tumor cells was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and the IC₅₀ values of J2-C3 were 65.57, 93.33 and 122.95 µg/mL against A549, HT-29 and HepG2 cell lines, respectively. Therefore, J2-C3 might be developed as a potential anti-tumor agent.     

Sporiolides A and B,New Cytotoxic Twelve-Membered Macrolides from a Marine-Derived Fungus Cladosporium Species

Two new cytotoxic twelve-membered macrolides, sporiolides A (1) and B (2), were isolated from the cultured broth of a fungus Cladosporium sp., which was separated from an Okinawan marine brown alga Actinotrichia fragilis, and the structures were elucidated by spectroscopic data. Sporiolides A (1) and B (2) exhibited cytotoxicity against murine lymphoma L1210 cells. Spoliolide A (1) showed antifungal activity against Cryptococcus neoformans and Neurospora crassa.     

Preparative Separation of Sulfur-Containing Diketopiperazines from Marine Fungus Cladosporium sp. Using High-Speed Counter-Current Chromatography in Stepwise Elution Mode

High-speed counter-current chromatography (HSCCC) was successively applied to the separation of three sulfur-containing diketopiperazines (DKPs) (including two new compounds cladosporin A (1) and cladosporin B (3), and a known compound haematocin (2)) from a marine fungus Cladosporium sp. The two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water at (1:1:1:1, v/v) and (2:1:2:1, v/v), in stepwise elution mode, was used for HSCCC. The preparative HSCCC separation was performed on 300 mg of crude sample yielding 26.7 mg of compound 3 at a purity of over 95%, 53.6 mg of a mixture of compounds 1 and 2, which was further separated by preparative-HPLC yielding 14.3 mg of compound 1 and 25.4 mg of compound 2 each at a purity of over 95%. Their structures were established by spectroscopic methods. The sulfur-containing DKPs suppressed the proliferation of hepatocellular carcinoma cell line HepG2. The present work represents the first application of HSCCC in the efficient preparation of marine fungal natural products.     

Indole Diterpenoids and Isocoumarin from the Fungus,Aspergillus flavus,Isolated from the Prawn,Penaeus vannamei

Two new indole-diterpenoids (1 and 2) and a new isocoumarin (3), along with the known β-aflatrem (4), paspalinine (5), leporin B (6), α-cyclopiazonic acid (7), iso-α-cyclopiazonic acid (8), ditryptophenaline (9), aflatoxin B₁ (10), 7-O-acetylkojic acid (11) and kojic acid (12), were isolated from the fermentation broth of the marine-derived fungus, Aspergillus flavus OUCMDZ-2205. The structures of Compounds 1–12 were elucidated by spectroscopic analyses, quantum ECD calculations and the chemical method. New Compound 1 exhibited antibacterial activity against Staphylococcus aureus with a MIC value of 20.5 μM. Both new Compounds 1 and 2 could arrest the A549 cell cycle in the S phase at a concentration of 10 μM. Compound 1 showed PKC-beta inhibition with an IC₅₀ value of 15.6 μM. In addition, the absolute configurations of the known compounds, 4–6 and leporin A (6a), were also determined for the first time.     

Cytotoxic and Antibacterial Angucycline- and Prodigiosin- Analogues from the Deep-Sea Derived Streptomyces sp. SCSIO 11594

Two new C-glycoside angucyclines, marangucycline A (1) and marangucycline B (2), along with three known compounds, dehydroxyaquayamycin (3), undecylprodigiosin (4) and metacycloprodigiosin (5), have been identified as products of the deep-sea sediment strain Streptomyces sp. SCSIO 11594. New structures were elucidated on the basis of HRESIMS, 1D and 2D NMR analyses and comparisons to previously reported datasets. Compounds 2 and 4 displayed in vitro cytotoxicity against four cancer cell lines A594, CNE2, HepG2, MCF-7 superior to those obtained with cisplatin, the positive control. Notably, compound 2 bearing a keto-sugar displayed significant cytotoxicity against cancer cell lines with IC₅₀ values ranging from 0.24 to 0.56 μM; An IC₅₀ value of 3.67 μM was found when using non-cancerous hepatic cell line HL7702, demonstrating the cancer cell selectivity of 2. Compounds 1–3 were proved to have weak antibacterial activities against Enterococcus faecalis ATCC29212 with an MIC value of 64.0 μg/mL. Moreover, 3 displayed selective antibacterial activity against methicillin-resistant Staphylococcus epidermidis shhs-E1 with an MIC value of 16.0 μg/mL.     

Oxygenated Ylangene-Derived Sesquiterpenoids from the Soft Coral Lemnalia philippinensis

Chemical examination of a Taiwanese soft coral Lemnalia philippinensis led to the isolation of three oxygenated ylangene-derived sesquiterpenoids 1–3, including two new metabolites, philippinlins A and B (1 and 2). The structures of these compounds were elucidated on the basis of detailed spectroscopic data. Compound 1 was shown to exhibit cytotoxicity against HepG2, MDA-MB231 and A549 cancer cell lines.     

Tanjungides A and B: New Antitumoral Bromoindole Derived Compounds from Diazona cf formosa. Isolation and Total Synthesis

Tanjungides A (1) (Z isomer) and B (2) (E isomer), two novel dibrominated indole enamides, have been isolated from the tunicate Diazona cf formosa. Their structures were determined by spectroscopic methods including HRMS, and extensive 1D and 2D NMR. The stereochemistry of the cyclised cystine present in both compounds was determined by Marfey’s analysis after chemical degradation and hydrolysis. We also report the first total synthesis of these compounds using methyl 1H-indole-3-carboxylate as starting material and a linear sequence of 11 chemical steps. Tanjungides A and B exhibit significant cytotoxicity against human tumor cell lines.     

Identification and Expression of Two Novel Cytochrome P450 Genes,CYP6CV1 and CYP9A38, in Cnaphalocrocis medinalis (Lepidoptera: Pyralidae)

Cnaphalocrocis medinalis Güenée can cause severe losses in rice. Cytochrome P450s play crucial roles in the metabolism of allelochemicals in herbivorous insects. Two novel P450 cDNAs, CYP6CV1 and CYP9A38, were cloned from the midgut of C. medinalis. CYP6CV1 encodes a protein of 500 amino acid residues, while CYP9A38-predicted protein has 531 amino acid residues. Both cDNA-predicted proteins contain the conserved functional domains for all P450s. Phylogenetic analyses showed that CYP6CV1 is grouped in the cluster containing CYP6B members, while CYP9A38 is in the cluster including CYP9 members. However, both clusters are contained in the same higher lineage. Homologous analysis revealed that CYP6CV1 is most similar to CYP6B8, CYP6B7, CYP6B6, CYP6B2, and CYP6B4 with the highest amino acid identity of 41%. CYP9A38 is closest to CYP9A17, CYP9A21, CYP9A20, and CYP9A19 with the highest amino acid identity of 66%. Studies of temporal expression profiles revealed that CYP9A38 showed a steady increase in mRNA level during the five instar stages, but a low-expression level in pupae, and then presented at a high-expression level again in adults. Similar expression patterns were obtained with CYP6CV1. In the fifth instar larvae, CYP6CV1 was mainly expressed in midgut and fat bodies, whereas CYP9A38 was mainly expressed in midgut. Expression studies also revealed a 3.20-fold over-expression of CYP6CV1 and 3.54-fold over-expression of CYP9A38 after larval exposure to host rice resistance. Our results suggest that both CYP6CV1 and CYP9A38 may be involved in detoxification of rice phytochemicals.     

Anti-Inflammatory Potential of Monogalactosyl Diacylglycerols and a Monoacylglycerol from the Edible Brown Seaweed Fucus spiralis Linnaeus

A monoacylglycerol (1) and a 1:1 mixture of two monogalactosyl diacylglycerols (MGDGs) (2 and 3) were isolated from the brown seaweed Fucus spiralis Linnaeus. The structures were elucidated by spectroscopic means (NMR and MS) and by comparison with the literature. Compound 1 was composed of a glycerol moiety linked to oleic acid (C18:1 Ω9). Compounds 2 and 3 contained a glycerol moiety linked to a galactose unit and eicosapentaenoic acid (C20:5 Ω3) combined with octadecatetraenoic acid (C18:4 Ω3) or linolenic acid (C18:3 Ω3), respectively. The isolated compounds were tested for their cytotoxic and anti-inflammatory activity in RAW 264.7 macrophage cells. All of them inhibited NO production at non-cytotoxic concentrations. The fraction consisting of compounds 2 and 3, in a ratio of 1:1, was slightly more effective than compound 1 (IC₅₀ of 60.06 and 65.70 µg/mL, respectively). To our knowledge, this is the first report of these compounds from F. spiralis and on their anti-inflammatory capacity.     

New Anti-Inflammatory Cembranes from the Cultured Soft Coral Nephthea columnaris

Two new cembranes, columnariols A (1) and B (2), were isolated from the cultured soft coral Nephthea columnaris. The structures of cembranes 1 and 2 were elucidated by spectroscopic methods. In the anti-inflammatory effects test, cembranes 1 and 2 were found to significantly inhibit the accumulation of the pro-inflammatory iNOS and COX-2 protein of the lipopolysaccharide (LPS)-stimulated RAW264.7 macrophage cells. Compound 1 exhibited moderate cytotoxicity toward LNCaP cells with an IC₅₀ value of 9.80 μg/mL.     

Zn-Driven Discovery of a Hydrothermal Vent Fungal Metabolite Clavatustide C,and an Experimental Study of the Anti-Cancer Mechanism of Clavatustide B

A naturally new cyclopeptide, clavatustide C, was produced as a stress metabolite in response to abiotic stress elicitation by one of the hydrothermal vent fluid components Zn in the cultured mycelia of Aspergillus clavatus C2WU, which were isolated from Xenograpsus testudinatus. X. testudinatus lives at extreme, toxic habitat around the sulphur-rich hydrothermal vents in Taiwan Kueishantao. The known compound clavatustide B was also isolated and purified. This is the first example of a new hydrothermal vent microbial secondary metabolite produced in response to abiotic Zn treatment. The structures were established by spectroscopic means. The regulation of G1-S transition in hepatocellular carcinoma cell lines by clavatustide B was observed in our previous study. The purpose of the present study was to verify these results in other types of cancer cell lines and elucidate the possible molecular mechanism for the anti-cancer activities of clavatustide B. In different human cancer cell lines, including pancreatic cancer (Panc-1), gastric cancer (MGC-803), colorectal cancer (SW-480), retinoblastoma (WERI-Rb-1) and prostate cancer (PC3), clavatustide B efficiently suppressed cell proliferations in a dose-dependent manner. Although different cancer cell lines presented variety in Max effect dose and IC₅₀ dose, all cancer cell lines showed a lower Max effect dose and IC₅₀ dose compared with human fibroblasts (hFB) (p < 0.05). Moreover, significant accumulations in G1 phases and a reduction in S phases (p < 0.05) were observed under clavatustide B treatment. The expression levels of 2622 genes including 39 cell cycle-associated genes in HepG2 cells were significantly altered by the treatment with 15 μg/mL clavatustide B after 48 h. CCNE2 (cyclin E2) was proved to be the key regulator of clavatustide B-induced G1-S transition blocking in several cancer cell lines by using real-time PCR.     

Bioactive Hydantoin Alkaloids from the Red Sea Marine Sponge Hemimycale arabica

In the course of our continuing efforts to identify bioactive secondary metabolites from Red Sea marine invertebrates, we have investigated the sponge Hemimycale arabica. The antimicrobial fraction of an organic extract of the sponge afforded two new hydantoin alkaloids, hemimycalins A and B (2 and 3), together with the previously reported compound (Z)-5-(4-hydroxybenzylidene)imidazolidine-2,4-dione (1). The structures of the compounds were determined by extensive 1D and 2D NMR (COSY, HSQC and HMBC) studies and high-resolution mass spectral determinations. Hemimycalins A (2) and B (3) represent the first examples of the natural N-alkylated hydantoins from the sponge Hemimycale arabica. Compounds 1–3 displayed variable antimicrobial activities against E. coli, S. aureus, and C. albicans. In addition, compound 1 displayed moderate antiproliferative activity against the human cervical carcinoma (HeLa) cell line. These findings provide further insight into the chemical diversity as well as the biological activity of this class of compounds.     

Three bianthraquinone derivatives from the mangrove endophytic fungus Alternaria sp. ZJ9-6B from the South China Sea

Three new bianthraquinone derivatives, alterporriol K (1), L (2) and M (3), along with six known compounds were obtained from extracts of the endophytic fungus Alternaria sp. ZJ9-6B, isolated from the mangrove Aegiceras corniculatum collected in the South China Sea. Their structures were elucidated by one- and two-dimensional NMR spectroscopy, MS data analysis and circular dichroism measurements. Compounds 1, 2 and 3 were first isolated alterporriols with a C-2–C-2′ linkage. The crystallographic data of tetrahydroaltersolanol B (7) was reported for the first time. In the primary bioassays, alterporriol K and L exhibited moderate cytotoxic activity towards MDA-MB-435 and MCF-7 cells with IC₅₀ values ranging from 13.1 to 29.1 μM.     

Lobocrassins A-E: new cembrane-type diterpenoids from the soft coral Lobophytum crassum

Five new cembrane-type diterpenoids, lobocrassins A-E (1-5), were isolated from the soft coral Lobophytum crassum. The structures of cembranes 1-5 were established by spectroscopic and chemical methods and by comparison of the spectral data with those of known cembrane analogues. Lobocrassin A (1) is the first cembranoid possessing an α-chloromethyl-α-hydroxy-γ-lactone functionality and is the first chlorinated cembranoid from soft corals belonging to the genus Lobophytum. Lobocrassins B (2) and C (3) were found to be the stereoisomers of the known cembranes, 14-deoxycrassin (6) and pseudoplexaurol (7), respectively. Lobocrassin B (2) exhibited modest cytotoxicity toward K562, CCRF-CEM, Molt4, and HepG2 tumor cells and displayed significant inhibitory effects on the generation of superoxide anion and the release of elastase by human neutrophils.