Antitumor Effects of a Sesquiterpene Derivative from Marine Sponge in Human Breast Cancer Cells

In this study, the anti-proliferative effect of ilimaquinone, a sesquiterpene derivative from the marine sponge, in breast cancer cells was investigated. Ilimaquinone inhibited the proliferation of MCF-7 and MDA-MB-231 breast cancer cells with IC₅₀ values of 10.6 μM and 13.5 μM, respectively. Non-tumorigenic human breast epithelial cells were less sensitive to ilimaquinone than breast cancer cells. Flow cytometric and Western blot analysis showed that ilimaquinone induced S-phase arrest by modulating the expression of p-CDC-2 and p21. Ilimaquinone induces apoptosis, which is accompanied by multiple biological biomarkers, including the downregulation of Akt, ERK, and Bax, upregulation of p38, loss of mitochondrial membrane potential, increased reactive oxygen species generation, and induced autophagy. Collectively, these findings suggest that ilimaquinone causes cell cycle arrest as well as induces apoptosis and autophagy in breast cancer cells.     

Insights into Cytotoxic Behavior of Lepadins and Structure Elucidation of the New Alkaloid Lepadin L from the Mediterranean Ascidian Clavelina lepadiformis

The chemical investigation of the Mediterranean ascidian Clavelina lepadiformis has led to the isolation of a new lepadin, named lepadin L, and two known metabolites belonging to the same family, lepadins A and B. The planar structure and relative configuration of the decahydroquinoline ring of lepadin L were established both by means of HR-ESIMS and by a detailed as extensive analysis of 1D and 2D NMR spectra. Moreover, microscale derivatization of the new alkaloid lepadin L was performed to assess the relative configuration of the functionalized alkyl side chain. Lepadins A, B, and L were tested for their cytotoxic activity on a panel of cancer cell lines (human melanoma [A375], human breast [MDA-MB-468], human colon adenocarcinoma [HT29], human colorectal carcinoma [HCT116], and mouse myoblast [C2C12]). Interestingly, a deeper investigation into the mechanism of action of the most cytotoxic metabolite, lepadin A, on the A375 cells has highlighted its ability to induce a strongly inhibition of cell migration, G2/M phase cell cycle arrest and a dose-dependent decrease of cell clonogenity, suggesting that it is able to impair self-renewing capacity of A375 cells.     

Anticancer activity evaluation of kuanoniamines A and C isolated from the marine sponge Oceanapia sagittaria, collected from the Gulf of Thailand

The pyridoacridine alkaloids kuanoniamines A and C were isolated together with 24 α-methylcholestanol, p-hydroxybenzaldehyde, p-hydroxybenzoic acid, phenylacetic acid and 3-formylindole from the ethyl acetate extract of the marine sponge Oceanapia sagittaria (Sollas), collected from the Gulf of Thailand. Kuanoniamines A and C were evaluated for their effect on the growth of five human tumour and a non-tumour cell lines, as well as on the proliferation of human lymphocytes. Kuanoniamine A was found to be a potent growth inhibitor of all the tumour and a non-tumour cell lines while kuanoniamine C was less potent but showed high selectivity toward the estrogen dependent (ER+) breast cancer cell line. Kuanoniamine A has shown to be a more potent inhibitor of DNA synthesis than kuanoniamine C. Kuanoniamine A was also found to cause an extensive reduction of the MCF-7 cells in G2/M phase as well as an increase in the apoptotic cells.     

Structural characterization and anti-HSV-1 and HSV-2 activity of glycolipids from the marine algae Osmundaria obtusiloba isolated from Southeastern Brazilian coast

Glycolipids were extracted from the red alga Osmundaria obtusiloba from Southeastern Brazilian coast. The acetone insoluble material was extracted with chloroform/methanol and the lipids, enriched in glycolipids, were fractionated on a silica gel column eluted with chloroform, acetone and then methanol. Three major orcinol-positive bands were found in the acetone and methanol fractions, being detected by thin layer chromatography. The structures of the corresponding glycolipids were elucidated by ESI-MS and (1)H/(13)C NMR analysis, on the basis of their tandem-MS behavior and HSQC, TOCSY fingerprints. For the first time, the structure of sulfoquinovosyldiacylglycerol from the red alga Osmundaria obtusiloba was characterized. This molecule exhibited potent antiviral activity against HSV-1 and HSV-2 with EC(50) values of 42 μg/mL to HSV-1 and 12 μg/mL to HSV-2, respectively. Two other glycolipids, mono- and digalactosyldiacylglycerol, were also found in the alga, being characterized by ESI-MS/MS. The structural elucidation of algae glycolipids is a first step for a better understanding of the relation between these structures and their biological activities.     

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.     

Characterization of the Dual Functions of LvCrustinVII from Litopenaeus vannamei as Antimicrobial Peptide and Opsonin

Crustin are a family of antimicrobial peptides that play an important role in protecting against pathogens infection in the innate immune system of crustaceans. Previously, we identified several novel types of crustins, including type VI and type VII crustins. However, their immune functions were still unclear. In the present study, the immune function of type VII crustin LvCrustinVII were investigated in Litopenaeus vannamei. LvCrustinVII was wildly expressed in all tested tissues, with relatively high expression levels in hepatopancreas, epidermis and lymphoid organ. Upon Vibrio parahaemolyticus infection, LvCrustinVII was significantly upregulated in hepatopancreas. Recombinant LvCrustinVII (rLvCrustinVII) showed strong inhibitory activities against Gram-negative bacteria Vibrio harveyi and V. parahaemolyticus, while weak activities against the Gram-positive bacteria Staphylococcus aureus. Binding assay showed that rLvCrustinVII could bind strongly to V. harveyi and V. parahaemolyticus, as well as the cell wall components Glu, LPS and PGN. In the presence of Ca²⁺, rLvCrustinVII could agglutinate V. parahaemolyticus and enhance hemocyte phagocytosis. The present data partially illustrate the immune function of LvCrustinVII, which enrich our understanding on the functional mechanisms of crustins and provide useful information for application of this kind of antimicrobial peptides.     

香菇胞壁多糖的分子量测定及结构分析

采用高效液相色谱、气相色谱和化学分析等方法对香菇胞壁多糖（Lwp）的纯度、分子量及基本结构进行研究。结果表明：Lwp分子量为245 742 u,单糖组成为葡萄糖和极少量的甘露糖;Lwp中1→4糖苷键残基比例为42.7％,1→6糖苷键残基比例为38.1％,1→3糖苷键残基比例为11.1％,1→2糖苷键残基比例为8.1％。     

Dihydroaustrasulfone Alcohol Inhibits PDGF-Induced Proliferation and Migration of Human Aortic Smooth Muscle Cells through Inhibition of the Cell Cycle

Dihydroaustrasulfone alcohol is the synthetic precursor of austrasulfone, which is a marine natural product, isolated from the Taiwanese soft coral Cladiella australis. Dihydroaustrasulfone alcohol has anti-inflammatory, neuroprotective, antitumor and anti-atherogenic properties. Although dihydroaustrasulfone alcohol has been shown to inhibit neointima formation, its effect on human vascular smooth muscle cells (VSMCs) has not been elucidated. We examined the effects and the mechanisms of action of dihydroaustrasulfone alcohol on proliferation, migration and phenotypic modulation of human aortic smooth muscle cells (HASMCs). Dihydroaustrasulfone alcohol significantly inhibited proliferation, DNA synthesis and migration of HASMCs, without inducing cell death. Dihydroaustrasulfone alcohol also inhibited platelet-derived growth factor (PDGF)-induced expression of cyclin-dependent kinases (CDK) 2, CDK4, cyclin D1 and cyclin E. In addition, dihydroaustrasulfone alcohol inhibited PDGF-induced phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2), whereas it had no effect on the phosphorylation of phosphatidylinositol 3-kinase (PI3K)/(Akt). Moreover, treatment with PD98059, a highly selective ERK inhibitor, blocked PDGF-induced upregulation of cyclin D1 and cyclin E and downregulation of p27^kip1. Furthermore, dihydroaustrasulfone alcohol also inhibits VSMC synthetic phenotype formation induced by PDGF. For in vivo studies, dihydroaustrasulfone alcohol decreased smooth muscle cell proliferation in a rat model of restenosis induced by balloon injury. Immunohistochemical staining showed that dihydroaustrasulfone alcohol noticeably decreased the expression of proliferating cell nuclear antigen (PCNA) and altered VSMC phenotype from a synthetic to contractile state. Our findings provide important insights into the mechanisms underlying the vasoprotective actions of dihydroaustrasulfone alcohol and suggest that it may be a useful therapeutic agent for the treatment of vascular occlusive disease.     

Antinociceptive and anti-inflammatory activity from algae of the genus Caulerpa

Marine natural products have been the focus of discovery for new products of chemical and pharmacological interest. The aim of this study was to evaluate the antinociceptive activity of the methanolic (ME), acetate (AE), hexanic (HE) and chloroform (CE) extracts obtained from Caulerpa mexicana, and ME, CE and HE obtained from Caulerpa sertularioides. These marine algae are found all over the world, mainly in tropical regions. Models such as the writhing test, the hot plate test and formalin-induced nociception test were used to evaluate antinociceptive activity in laboratory mice. In the writhing test, all the extracts were administered orally at a concentration of 100 mg/kg, and induced high peripheral antinociceptive activity, with a reduction in the nociception induced by acetic acid above 65%. In the hot plate test, treatment with extracts from C. sertularioides (100 mg/kg, p.o.) did not significantly increase the latency of response, although the ME, AE and HE from C. mexicana showed activity in this model. This result suggests that these extracts exhibit antinociceptive activity. In the formalin test, it was observed that ME, AE and HE obtained from C. mexicana reduced the effects of formalin in both phases. On the other hand only CE from C. sertularioides induced significant inhibition of the nociceptive response in the first phase. To better assess the potential anti-inflammatory activity of the extracts, the carrageenan-induced peritonitis test was used to test Caulerpa spp. extracts on cell migration into the peritoneal cavity. In this assay, all extracts evaluated were able to significantly inhibit leukocyte migration into the peritoneal cavity in comparison with carrageenan. These data demonstrate that extracts from Caulerpa species elicit pronounced antinociceptive and anti-inflamatory activity against several nociception models. However, pharmacological and chemical studies are continuing in order to characterize the mechanism(s) responsible for the antinociceptive action and also to identify the active principles present in the Caulerpa species.     

依兰香细胞离体培养及其精油成分分析

以依兰香（Ｃａｎａｎｇａ ｏｄｏｒａｔａ）的花瓣及雄蕊群为外植体，在１／２ＭＳ＋２，４－Ｄ１－２＋ＫＴ０．２的培养基上分别诱导出白色和红色的两种愈伤组织，建立了依兰香细胞的大量培养（包括固体培养，悬浮培养及发酵罐培养）体系和单细胞克隆，通过化学方法进行高产同细胞系的初步筛选，从培养细胞中分离出依兰油，对油成分作了详细的分析。     

Evaluation of micropropagation for the introduction into cultivation and conservation of Lippia junelliana, an endemic aromatic plant from Argentina

The aims of this work were to introduce Lippia junelliana into cultivation, to compare the essential oil accumulation between cultivated and wild plants, and to reintroduce micropropagated plants in the location of the original population. The leaves and inflorescences of cultivated plants accumulated, on a dry weight basis, higher amounts of essential oil than their wild counterparts. Thus, total essential oil accumulation of cultivated plant parts was also significantly higher than that of wild counterparts. The cultivated plants showed the same essential oil profile than the wild plants. This work demonstrates that cultivation can be a more efficient vehicle to both preserve and exploit L. junelliana, than collection from the wild, because higher yields of biomass and oil accumulation can be achieved, while essential oil composition is less affected by the different treatments. The reintroduction of new plants into the species’ original location has proved to be a viable alternative for their in situ preservation or enrichment planting. This model of introduction of aromatic plants into cultivation through micropropagation could be a useful technique to recover valuable chemotypes from the wild in the search for new alternatives in the agriculture and for the preservation of natural resources for future generations.     

Cytotoxic Alkylynols of the Sponge Cribrochalina vasculum: Structure,Synthetic Analogs and SAR Studies

A series of twenty-three linear and branched chain mono acetylene lipids were isolated from the Caribbean Sea sponge Cribrochalina vasculum. Seventeen of the compounds, 1–17, are new, while six, 18–23, were previously characterized from the same sponge. Some of the new acetylene-3-hydroxy alkanes 1, 6, 7, 8, 10 were tested for selective cytotoxicity in non-small cell lung carcinoma (NSCLC) cells over WI-38 normal diploid lung fibroblasts. Compound 7, presented clear tumor selective activity while, 1 and 8, showed selectivity at lower doses and 6 and 10, were not active towards NSCLC cells at all. The earlier reported selective cytotoxicity of some acetylene-3-hydroxy alkanes (scal-18 and 23), in NSCLC cells and/or other tumor cell types were also confirmed for 19, 20 and 22. To further study the structure activity relationships (SAR) of this group of compounds, we synthesized several derivatives of acetylene-3-hydroxy alkanes, rac-18, scal-S-18, R-18, rac-27, rac-32, R-32, S-32, rac-33, rac-41, rac-42, rac-43, rac-45, rac-48 and rac-49, along with other 3-substituted derivatives, rac-35, rac-36, rac-37, rac-38, rac-39 and rac-40, and assessed their cytotoxic activity against NSCLC cells and diploid fibroblasts. SAR studies revealed that the alcohol moiety at position 3 and its absolute R configuration both were essential for the tumor cell line selective activity while for its cytotoxic magnitude the alkyl chain length and branching were of less significance.     

正红505玉米花丝多糖结构及其降血糖活性分析

采用水提醇沉法提取正红505玉米花丝中的多糖成分,通过核磁共振波谱法(NMR)、高效液相色谱法(HPLC)、傅里叶变换红外吸收光谱法(FTIR)及气相色谱-质谱联用法(GC-MS)研究该多糖的单糖组成及结构,借助噻唑蓝比色(MTT)细胞试验,研究多糖CSP505对大鼠胰岛细胞RINm5F和人体肝脏细胞HepG2的降血糖活性。结果表明,多糖CSP505可能是一种由D-葡萄糖和D-半乳糖组成的β构型的多糖,其中葡萄糖和半乳糖的比约为3∶1。随着多糖浓度的增加,RINm5F细胞存活力随着增强,加药组中多糖以200、500、1000μg/mL浓度与四氧嘧啶共同作用时,细胞存活率分别提高了16.1%、16.1%、21.1%,达到极显著差异(P0.01);HepG2细胞存活能力也增强,加药组中多糖分别以200、500、1 000μg/mL浓度作用于细胞时,细胞存活力分别为100.4%、101.4%、103.3%。与阴性对照组相比较,加药组中在3个浓度的多糖作用下,腺苷酸环化酶(ADCY4)、糖原磷酸化酶(GPBB)和糖原合酶(GYS2)浓度表现出显著或极显著差异,而cAMP在浓度为1 000μg/mL时也表现出极显著差异。结果表明,该多糖可通过促进ADCY4与cAMP的合成来使胰岛素分泌增加,且可通过抑制糖原分解并促进糖原合成等过程起到降低血糖的作用。     

Structure and Absolute Configuration of Phenanthro-perylene Quinone Pigments from the Deep-Sea Crinoid Hypalocrinus naresianus

Two new water-soluble phenanthroperylene quinones, gymnochrome H (2) and monosulfated gymnochrome A (3), as well as the known compounds gymnochrome A (4) and monosulfated gymnochrome D (5) were isolated from the deep-sea crinoid Hypalocrinus naresianus, which had been collected in the deep sea of Japan. The structures of the compounds were elucidated by spectroscopic analysis including HRMS, 1D ¹H and ¹³C NMR, and 2D NMR. The absolute configuration was determined by ECD spectroscopy, analysis of J-couplings and ROE contacts, and DFT calculations. The configuration of the axial chirality of all isolated phenanthroperylene quinones (2–5) was determined to be (P). For gymnochrome H (2) and monosulfated gymnochrome A (3), a (2′S,2″R) configuration was determined, whereas for monosulfated gymnochrome D (5) a (2′R,2″R), configuration was determined. Acetylated quinones are unusual among natural products from an echinoderm and gymnochrome H (2) together with the recently reported gymnochrome G (1) represent the first isolated acetylated phenanthroperylene quinones.     

Isolation and Structure Elucidation of Three New Dolastanes from the Brown Alga Dilophus spiralis

Three new dolastane diterpenes (1–3) and five previously reported perhydroazulenes were isolated from the organic extracts of the brown alga Dilophus spiralis. The structure elucidation and the assignment of the relative configurations of the isolated natural products were based on extensive analyses of their spectroscopic data, whereas the absolute configuration of metabolite 2 was determined through its chemical conversion to a previously isolated compound of known configuration.     

Isolation and Structure Elucidation of GM4-Type Gangliosides from the Okinawan Starfish Protoreaster nodosus

Three new ganglioside molecular species, termed PNG-1, PNG-2A, and PNG-2B were isolated from pyloric caeca of the starfish Protoreaster nodosus. Their structures were elucidated using a combination of spectroscopic and chemical methods, and characterized as 1-O-[8-O-methyl-N-acetyl-α-neuraminosyl-(2→3)-β-galactopyranosyl]-ceramide for PNG-1, 1-O-[β-galactofuranosyl-(1→3)-α-galactopyranosyl-(1→4)-8-O-methyl-N-acetyl-α-neuraminosyl-(2→3)-β-galactopyranosyl]-ceramide for PNG-2A, and 1-O-[β-galactofuranosyl-(1→3)-α-galactopyranosyl-(1→9)-N-acetyl-α-neuraminosyl-(2→3)-β-galactopyranosyl]-ceramide for PNG-2B. PNG-2A and PNG-2B represent the first GM4 elongation products in nature.     

Cerium binding activity of pectins isolated from the seagrasses Zostera marina and Phyllospadix iwatensis

Cerium binding activity of three different water soluble pectin compounds of different origin was studied in a batch sorption system. The Langmuir, Freundlich and BET sorption models were adopted to describe the binding reactions between metal ions and pectin molecules. The Langmuir model provided the best fit. Within the pH range from 4.0 to 6.0, the largest amount of the cerium ions was bound by pectin isolated from the seagrass Phylospadix iwatensis in comparison to pectin extracted from the seagrass Zostera marina and pectin obtained from citrus peel (commercial grade). The Langmuir constants were also highest for the pectin samples isolated from the seagrass P. iwatensis. The results obtained from this study suggest that pectin is a prospective source for the development of radioisotope-removing pharmaceuticals.     

Preparation and Characterization of Thermally Stable Collagens from the Scales of Lizardfish (Synodus macrops)

Marine collagen is gaining vast interest because of its high biocompatibility and lack of religious and social restrictions compared with collagen from terrestrial sources. In this study, lizardfish (Synodus macrops) scales were used to isolate acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC). Both ASC and PSC were identified as type I collagen with intact triple-helix structures by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and spectroscopy. The ASC and PSC had high amino acids of 237 residues/1000 residues and 236 residues/1000 residues, respectively. Thus, the maximum transition temperature (T_max) of ASC (43.2 °C) was higher than that of PSC (42.5 °C). Interestingly, the T_max of both ASC and PSC was higher than that of rat tail collagen (39.4 °C) and calf skin collagen (35.0 °C), the terrestrial collagen. Solubility tests showed that both ASC and PSC exhibited high solubility in the acidic pH ranges. ASC was less susceptible to the “salting out” effect compared with PSC. Both collagen types were nontoxic to HaCaT and MC3T3-E1 cells, and ASC was associated with a higher cell viability than PSC. These results indicated that ASC from lizardfish scales could be an alternative to terrestrial sources of collagen, with potential for biomedical applications.     

橡胶树树皮中ACC氧化酶基因(HbACO7)的表达特性及功能探究

天然橡胶是重要的工业原材料,主要来自橡胶树树皮中的乳管。乙烯能促进橡胶树乳管产胶和排胶,在树皮上施用乙烯利（一种乙烯释放剂）或乙烯可显著提高胶乳产量。基于本课题组前期获得的橡胶树基因组和转录组数据,克隆橡胶树中乙烯合成通路关键酶——1-氨基环丙烷基-1-羧酸（ACC）氧化酶基因家族（HbACOs）的全部8个家族基因,研究其组织表达模式。结果表明：在分析的7种组织中,尽管HbACO基因存在不同程度的冗余表达,但每种基因的组织表达模式具有明显的特异性,其中HbACO7是树皮中表达量最高的家族基因。HbACO7基因的表达在割胶季节的不同月份发生明显变化,其中在8月的表达量最高。成功构建了HbACO7基因的原核表达载体,实现其在大肠杆菌BL21中的诱导表达。纯化后的HbACO7重组蛋白具有明显的ACO酶催化活性,成功地催化了乙烯的体外合成。该结果将为后续橡胶树树皮中的乙烯生物合成调控机制研究提供参考。