Changes at small intestine induced by food-fish contaminated with ciguatoxins.

Ciguateric syndrome is a food poisoning associated with the consumption of some species of fish that have accumulated ciguatoxins (CTXs) in their tissues. The effects of the syndrome occur with nervous imbalances which have been described for quite some time, and mentioned in sailing literature for centuries. In the last decade, research has been focused on the implementation of analytical methods for toxin identification and the study of action modes of CTXs to design effective treatments. However, an important aspect is to determine the damage that CTXs caused in the organs of affected individuals. In this work, the damages observed in tissues of mice, mainly in the small intestine, were analyzed. The animals were fed with CTX-contaminated fish muscle at concentrations 10-times below the median lethal dose (LD50) for 10 weeks. The analysis of tissues derived from the oral treatment resulted in an increased occurrence of Paneth cells, presence of lymphoid tissue infiltrating the mucosa and fibrous lesions in the mucosal layer of the small intestine. A decreasing weight in animals fed with toxic muscle was observed.

Laurequinone, a Lead Compound against Leishmania.

Among neglected tropical diseases, leishmaniasis is one of the leading causes, not only of deaths but also of disability-adjusted life years. This disease, caused by protozoan parasites of the genus Leishmania, triggers different clinical manifestations, with cutaneous, mucocutaneous, and visceral forms. As existing treatments for this parasitosis are not sufficiently effective or safe for the patient, in this work, different sesquiterpenes isolated from the red alga Laurencia johnstonii have been studied for this purpose. The different compounds were tested in vitro against the promastigote and amastigote forms of Leishmania amazonensis. Different assays were also performed, including the measurement of mitochondrial potential, determination of ROS accumulation, and chromatin condensation, among others, focused on the detection of the cell death process known in this type of organism as apoptosis-like. Five compounds were identified that displayed leishmanicidal activity: laurequinone, laurinterol, debromolaurinterol, isolaurinterol, and aplysin, showing IC50 values against promastigotes of 1.87, 34.45, 12.48, 10.09, and 54.13 µM, respectively. Laurequinone was the most potent compound tested and was shown to be more effective than the reference drug miltefosine against promastigotes. Different death mechanism studies carried out showed that laurequinone appears to induce programmed cell death or apoptosis in the parasite studied. The obtained results underline the potential of this sesquiterpene as a novel anti-kinetoplastid therapeutic agent.

Advanced Methods for Natural Products Discovery: Bioactivity Screening, Dereplication, Metabolomics Profiling, Genomic Sequencing, Databases and Informatic Tools, and Structure Elucidation.

Natural Products (NP) are essential for the discovery of novel drugs and products for numerous biotechnological applications. The NP discovery process is expensive and time-consuming, having as major hurdles dereplication (early identification of known compounds) and structure elucidation, particularly the determination of the absolute configuration of metabolites with stereogenic centers. This review comprehensively focuses on recent technological and instrumental advances, highlighting the development of methods that alleviate these obstacles, paving the way for accelerating NP discovery towards biotechnological applications. Herein, we emphasize the most innovative high-throughput tools and methods for advancing bioactivity screening, NP chemical analysis, dereplication, metabolite profiling, metabolomics, genome sequencing and/or genomics approaches, databases, bioinformatics, chemoinformatics, and three-dimensional NP structure elucidation.

Chamigrane-Type Sesquiterpenes from Laurencia dendroidea as Lead Compounds against Naegleria fowleri.

Naegleria fowleri is an opportunistic protozoon that can be found in warm water bodies. It is the causative agent of the primary amoebic meningoencephalitis. Focused on our interest to develop promising lead structures for the development of antiparasitic agents, this study was aimed at identifying new anti-Naegleria marine natural products from a collection of chamigrane-type sesquiterpenes with structural variety in the levels of saturation, halogenation and oxygenation isolated from Laurencia dendroidea. (+)-Elatol (1) was the most active compound against Naegleria fowleri trophozoites with IC50 values of 1.08 µM against the ATCC 30808™ strain and 1.14 µM against the ATCC 30215™ strain. Furthermore, the activity of (+)-elatol (1) against the resistant stage of N. fowleri was also assessed, showing great cysticidal properties with a very similar IC50 value (1.14 µM) to the one obtained for the trophozoite stage. Moreover, at low concentrations (+)-elatol (1) showed no toxic effect towards murine macrophages and could induce the appearance of different cellular events related to the programmed cell death, such as an increase of the plasma membrane permeability, reactive oxygen species overproduction, mitochondrial malfunction or chromatin condensation. Its enantiomer (-)-elatol (2) was shown to be 34-fold less potent with an IC50 of 36.77 µM and 38.03 µM. An analysis of the structure-activity relationship suggests that dehalogenation leads to a significant decrease of activity. The lipophilic character of these compounds is an essential property to cross the blood-brain barrier, therefore they represent interesting chemical scaffolds to develop new drugs.

Antiamoebic effects of sesquiterpene lactones isolated from the zoanthid Palythoa aff. clavata.

Opportunistic parasitic protozoa of genus Acanthamoeba are responsible to cause severe infections in humans such as Acanthamoeba Keratitis or Amoebic Granulomatous Encephalitis. Current treatments are usually toxic and inefficient and there is a need to access new therapeutic agents. The antiamoebic effects of nephthediol (1) and fourteen germacranolide and eudesmanolide sesquiterpene lactones (2-5, 7-12) isolated from the indigenous zoanthid Palythoa aff. clavata collected at the coast of Lanzarote, Canary Islands were studied against Acanthamoeba castellanii Neff, and the clinical strains A. polyphaga and A. griffini. 4-epi-arbusculin A (11) presented the lowest IC50 value (26,47 ± 1,69 µM) against A. castellanii Neff and low cytotoxicity against murine macrophages, followed by isobadgerin (2), which also showed to be active against A. castellanii Neff cysts. The studies on the mode of action of compounds 2 and 11 revealed these sesquiterpene lactones induce mechanisms of PDC on A. castellanii Neff.

Antimycobacterial Activity of Laurinterol and Aplysin from Laurencia johnstonii.

Marine environments represent a great opportunity for the discovery of compounds with a wide spectrum of bioactive properties. Due to their large variety and functions derived from natural selection, marine natural products may allow the identification of novel drugs based not only on newly discovered bioactive metabolites but also on already known compounds not yet thoroughly investigated. Since drug resistance has caused an increase in infections by Mycobacterium tuberculosis and nontuberculous mycobacteria, the re-evaluation of known bioactive metabolites has been suggested as a good approach to addressing this problem. In this sense, this study presents an evaluation of the in vitro effect of laurinterol and aplysin, two brominated sesquiterpenes isolated from Laurencia johnstonii, against nine M. tuberculosis strains and six nontuberculous mycobacteria (NTM). Laurinterol exhibited good antimycobacterial activity, especially against nontuberculous mycobacteria, being remarkable its effect against Mycobacterium abscessus, with minimum inhibitory concentration (MIC) values lower than those of the reference drug imipenem. This study provides further evidence for the antimycobacterial activity of some sesquiterpenes from L. johnstonii, which can be considered interesting lead compounds for the discovery of novel molecules to treat NTM infections.

Modulation of Glial Responses by Furanocembranolides: Leptolide Diminishes Microglial Inflammation in Vitro and Ameliorates Gliosis In Vivo in a Mouse Model of Obesity and Insulin Resistance.

Neurodegenerative diseases are age-related disorders caused by progressive neuronal death in different regions of the nervous system. Neuroinflammation, modulated by glial cells, is a crucial event during the neurodegenerative process; consequently, there is an urgency to find new therapeutic products with anti-glioinflammatory properties. Five new furanocembranolides (1-5), along with leptolide, were isolated from two different extracts of Leptogorgia sp., and compound 6 was obtained from chemical transformation of leptolide. Their structures were determined based on spectroscopic evidence. These seven furanocembranolides were screened in vitro by measuring their ability to modulate interleukin-1ß (IL-1ß) production by microglial BV2 cells after LPS (lipopolysaccharide) stimulation. Leptolide and compounds 3, 4 and 6 exhibited clear anti-inflammatory effects on microglial cells, while compound 2 presented a pro-inflammatory outcome. The in vitro results prompted us to assess anti-glioinflammatory effects of leptolide in vivo in a high-fat diet-induced obese mouse model. Interestingly, leptolide treatment ameliorated both microgliosis and astrogliosis in this animal model. Taken together, our results reveal a promising direct biological effect of furanocembranolides on microglial cells as bioactive anti-inflammatory molecules. Among them, leptolide provides us a feasible therapeutic approach to treat neuroinflammation concomitant with metabolic impairment.

Marine Anticancer Agents: An Overview with a Particular Focus on Their Chemical Classes.

The marine environment is a rich source of biologically active molecules for the treatment of human diseases, especially cancer. The adaptation to unique environmental conditions led marine organisms to evolve different pathways than their terrestrial counterparts, thus producing unique chemicals with a broad diversity and complexity. So far, more than 36,000 compounds have been isolated from marine micro- and macro-organisms including but not limited to fungi, bacteria, microalgae, macroalgae, sponges, corals, mollusks and tunicates, with hundreds of new marine natural products (MNPs) being discovered every year. Marine-based pharmaceuticals have started to impact modern pharmacology and different anti-cancer drugs derived from marine compounds have been approved for clinical use, such as: cytarabine, vidarabine, nelarabine (prodrug of ara-G), fludarabine phosphate (pro-drug of ara-A), trabectedin, eribulin mesylate, brentuximab vedotin, polatuzumab vedotin, enfortumab vedotin, belantamab mafodotin, plitidepsin, and lurbinectedin. This review focuses on the bioactive molecules derived from the marine environment with anticancer activity, discussing their families, origin, structural features and therapeutic use.

Antiprotozoal activities of marine polyether triterpenoids.

Chagas disease and leishmaniasis are tropical neglected diseases caused by kinetoplastids protozoan parasites of Trypanosoma and Leishmania genera, and a public health burden with high morbidity and mortality rates in developing countries. Among difficulties with their epidemiological control, a major problem is their limited and toxic treatments to attend the affected populations; therefore, new therapies are needed in order to find new active molecules. In this work, sixteen Laurencia oxasqualenoid metabolites, natural compounds 1-11 and semisynthetic derivatives 12-16, were tested against Leishmania amazonensis, Leishmania donovani and Trypanosoma cruzi. The results obtained point out that eight substances possess potent activities, with IC50 values in the range of 5.40-46.45 µM. The antikinetoplastid action mode of the main metabolite dehydrothyrsiferol (1) was developed, also supported by AFM images. The semi-synthetic active compound 28-iodosaiyacenol B (15) showed an IC50 5.40 µM against Leishmania amazonensis, turned to be non-toxic against the murine macrophage cell line J774A.1 (CC50 > 100). These values are comparable with the reference compound miltefosine IC50 6.48 ±â€¯0.24 and CC50 72.19 ±â€¯3.06 µM, suggesting that this substance could be scaffold for development of new antikinetoplastid drugs.

Spiralyde A, an Antikinetoplastid Dolabellane from the Brown Alga Dictyota spiralis.

Bioassay-guided fractionation of the antikinetoplastid extract of the brown alga Dictyota spiralis has led to the isolation of spiralyde A (1), a new dolabellane aldehyde, along with other five known related diterpenes (2⁻6). Their structures were determined by HRESIMS, 1D and 2D NMR spectroscopy, and comparison with data reported in the literature. The antiparasitic activity of all compounds was evaluated. Spiralyde A (1) and the known compound 3,4-epoxy-7,18-dolabelladiene (2) were the most active compounds against Leishmania amazonensis and Trypanosoma cruzi. Spiralyde A (1) was the most potent compound, comparable to benznidazole, the reference drug for trypanocidal activity.

Evaluation of Oxasqualenoids from the Red Alga Laurencia viridis against Acanthamoeba.

Acanthamoeba genus is a widely distributed and opportunistic parasite with increasing importance worldwide as an emerging pathogen in the past decades. This protozoan has an active trophozoite stage, a cyst stage, and is dormant and very resistant. It can cause Acanthamoeba keratitis, an ocular sight-threatening disease, and granulomatous amoebic encephalitis, a chronic, very fatal brain pathology. In this study, the amoebicidal activity of sixteen Laurencia oxasqualenoid metabolites and semisynthetic derivatives were tested against Acanthamoeba castellanii Neff. The results obtained point out that iubol (3) and dehydrothyrsiferol (1) possess potent activities, with IC50 values of 5.30 and 12.83 µM, respectively. The hydroxylated congeners thyrsiferol (2) and 22-hydroxydehydrothyrsiferol (4), active in the same value range at IC50 13.97 and 17.00 µM, are not toxic against murine macrophages; thus, they are solid candidates for the development of new amoebicidal therapies.

Antitumoral Effect of Laurinterol on 3D Culture of Breast Cancer Explants.

Macroalgae represent an important source of bioactive compounds with a wide range of biotechnological applications. Overall, the discovery of effective cytotoxic compounds with pharmaceutical potential is a significant challenge, mostly because they are scarce in nature or their total synthesis is not efficient, while the bioprospecting models currently used do not predict clinical responses. Given this context, we used three-dimensional (3D) cultures of human breast cancer explants to evaluate the antitumoral effect of laurinterol, the major compound of an ethanolic extract of Laurencia johnstonii. To this end, we evaluated the metabolic and histopathological effects of the crude extract of L. johnstonii and laurinterol on Vero and MCF-7 cells, in addition to breast cancer explants. We observed a dose-dependent inhibition of the metabolic activity, as well as morphologic and nuclear changes characteristic of apoptosis. On the other hand, a reduced metabolic viability and marked necrosis areas were observed in breast cancer explants incubated with the crude extract, while explants treated with laurinterol exhibited a heterogeneous response which was associated with the individual response of each human tumor sample. This study supports the cytotoxic and antitumoral effects of laurinterol in in vitro cell cultures and in ex vivo organotypic cultures of human breast cancer explants.

Antiamoebic Activities of Indolocarbazole Metabolites Isolated from Streptomyces sanyensis Cultures.

Indolocarbazoles are a family of natural alkaloids characterized by their potent protein kinase and topoisomerase I inhibitory activity. Among them, staurosporine (1) has exhibited promising inhibitory activity against parasites. Based on new insights on the activity and mechanism of action of STS in Acanthamoeba parasites, this work reports the isolation, identification, and the anti-Acanthamoeba activity of the minor metabolites 7-oxostaurosporine (2), 4'-demethylamino-4'-oxostaurosporine (3), and streptocarbazole B (4), isolated from cultures of the mangrove strain Streptomyces sanyensis. A clear correlation between the antiparasitic activities and the structural elements and conformations of the indolocarbazoles 1-4 was observed. Also, the study reveals that 7-oxostaurosporine (2) affects membrane permeability and causes mitochondrial damages on trophozoites of A. castellanii Neff.

Anti-Acanthamoeba Activity of Brominated Sesquiterpenes from Laurencia johnstonii.

Focused on our interest to develop novel antiparasistic agents, the present study was aimed to evaluate the biological activity of an extract of Laurencia johnstonii collected in Baja California Sur, Mexico, against an Acantamoeba castellanii Neff strain. Bioassay-guided fractionation allowed us to identify the amoebicidal diastereoisomers α-bromocuparane (4) and α-isobromocuparane (5). Furthermore, bromination of the inactive laurinterol (1) and isolaurinterol (2) yielded four halogenated derivatives, (6)⁻(9), which improved the activity of the natural sesquiterpenes. Among them, the most active compound was 3α-bromojohnstane (7), a sesquiterpene derivative which possesses a novel carbon skeleton johnstane.

Chloro-Furanocembranolides from Leptogorgia sp. Improve Pancreatic Beta-Cell Proliferation.

Two new chloro-furanocembranolides (1, 2) and two new 1,4-diketo cembranolides (3, 4) were isolated from the crude extract of Leptogorgia sp. together with a new seco-furanocembranolide (5) and the known Z-deoxypukalide (6), rubifolide (7), scabrolide D (8) and epoxylophodione (9). Their structures were determined based on spectroscopic evidence. Four compounds: 1, 2, 7 and 8 were found to activate the proliferation of pancreatic insulin-producing (beta) cells.

Leptolide Improves Insulin Resistance in Diet-Induced Obese Mice.

Type 2 diabetes (T2DM) is a complex disease linked to pancreatic beta-cell failure and insulin resistance. Current antidiabetic treatment regimens for T2DM include insulin sensitizers and insulin secretagogues. We have previously demonstrated that leptolide, a member of the furanocembranolides family, promotes pancreatic beta-cell proliferation in mice. Considering the beneficial effects of leptolide in diabetic mice, in this study, we aimed to address the capability of leptolide to improve insulin resistance associated with the pathology of obesity. To this end, we tested the hypothesis that leptolide should protect against fatty acid-induced insulin resistance in hepatocytes. In a time-dependent manner, leptolide (0.1 µM) augmented insulin-stimulated phosphorylation of protein kinase B (PKB) by two-fold above vehicle-treated HepG2 cells. In addition, leptolide (0.1 µM) counteracted palmitate-induced insulin resistance by augmenting by four-fold insulin-stimulated phosphorylation of PKB in HepG2 cells. In vivo, acute intraperitoneal administration of leptolide (0.1 mg/kg and 1 mg/kg) improved glucose tolerance and insulin sensitivity in lean mice. Likewise, prolonged leptolide treatment (0.1 mg/kg) in diet-induced obese mice improved insulin sensitivity. These effects were paralleled with an ~50% increased of insulin-stimulated phosphorylation of PKB in liver and skeletal muscle and reduced circulating pro-inflammatory cytokines in obese mice. We concluded that leptolide significantly improves insulin sensitivity in vitro and in obese mice, suggesting that leptolide may be another potential treatment for T2DM.

Tanzawaic acids isolated from a marine-derived fungus of the genus Penicillium with cytotoxic activities.

Tanzawaic acids M (1), N (2), O (3) and P (4) and the known tanzawaic acids B (5) and E (6), have been isolated from an extract of a cultured marine-derived fungus (strain CF07370) identified as a member of the genus Penicillium. The structures of 1-4 were determined based on spectroscopic evidence. The antimicrobial and cytotoxic activities of compounds 1-6 were evaluated.

Recirculating packed-bed biofilm photobioreactor combined with membrane ultrafiltration as advanced wastewater treatment.

Packed-bed biofilm photobioreactor combined with ultrafiltration membrane was investigated for intensifying the process for secondary wastewater effluent treatment. Cylindrical glass carriers were used as supporting material for the microalgal-bacterial biofilm, which developed from indigenous microbial consortium. Glass carriers allowed adequate growth of the biofilm with limited suspended biomass. Stable operation was achieved after a start-up period of 1000 h, where supernatant biopolymer clusters were minimized and complete nitrification was observed. After that time, biomass productivity was 54 ± 18 mg·L-1·day-1. Green microalgae Tetradesmus obliquus and several strains of heterotrophic nitrification-aerobic denitrification bacteria and fungi were identified. Combined process exhibited COD, nitrogen and phosphorus removal rates of 56 ± 5%, 12 ± 2% and 20 ± 6%, respectively. Membrane fouling was mainly caused by biofilm formation, which was not effectively mitigated by air-scouring aided backwashing.

Gongolarones as antiamoeboid chemical scaffold.

Free Living Amoeba (FLA) infections caused by Acanthamoeba genus include chronic nervous system diseases such as Granulomatous Amoebic Encephalitis (GAE), or a severe eye infection known as Acanthamoeba keratitis (AK). Current studies focused on therapy against these diseases are aiming to find novel compounds with amoebicidal activity and low toxicity to human tissues. Brown algae, such as Gongolaria abies-marina (previously known as Cystoseira abies-marina, S.G. Gmelin), presents bioactive molecules of interest, including some with antiprotozoal activity. In this study, six meroterpenoids were isolated and purified from the species Gongolaria abies-marina. Gongolarones A (1), B (2) and C (3) were identified as new compounds. Additionally, cystomexicone B (4), 1'-methoxyamentadione (5) and 6Z-1'-methoxyamentadione (6) were isolated. All compounds exhibited amoebicidal activity against Acanthamoeba castellanii Neff, A. polyphaga and A. griffini strains. Gongolarones A (1) and C (3) showed the lowest IC50 values against the two stages of these amoebae (trophozoite and cyst). Structure-activity relationship revealed that the cyclization by ether formation from C-12 to C-15 of 1, and the isomerization Δ2 t to Δ3 t of 3, increased the antiamoeboid activity of both compounds. Furthermore, gongolarones A (1) and C (3) triggered chromatin condensation, mitochondrial malfunction, oxidative stress, and disorganization of the tubulin-actin cytoskeleton in treated trophozoites. Moreover, transmission electron microscopy (TEM) images analysis revealed that compounds 1 and 3 induced autophagy process and inhibited the encystation process. All those results suggest that both compounds could induce programmed cell death (PCD) in Acanthamoeba.

Meroterpenoids from Gongolaria abies-marina against Kinetoplastids: In Vitro Activity and Programmed Cell Death Study.

Leishmaniasis and Chagas disease affect millions of people worldwide. The available treatments against these parasitic diseases are limited and display multiple undesired effects. The brown alga belonging to the genus Gongolaria has been previously reported as a source of compounds with different biological activities. In a recent study from our group, Gongolaria abies-marine was proven to present antiamebic activity. Hence, this brown alga could be a promising source of interesting molecules for the development of new antiprotozoal drugs. In this study, four meroterpenoids were isolated and purified from a dichloromethane/ethyl acetate crude extract through a bioguided fractionation process targeting kinetoplastids. Moreover, the in vitro activity and toxicity were evaluated, and the induction of programmed cell death was checked in the most active and less toxic compounds, namely gongolarone B (2), 6Z-1'-methoxyamentadione (3) and 1'-methoxyamentadione (4). These meroterpenoids triggered mitochondrial malfunction, oxidative stress, chromatin condensation and alterations of the tubulin network. Furthermore, a transmission electron microscopy (TEM) image analysis showed that meroterpenoids (2-4) induced the formation of autophagy vacuoles and ER and Golgi complex disorganization. The obtained results demonstrated that the mechanisms of action at the cellular level of these compounds were able to induce autophagy as well as an apoptosis-like process in the treated parasites.