Bioactive lichen metabolites: alpine habitats as an untapped source

Lichens are fungal and algal/cyanobacterial symbioses resulting in the production of specific metabolites. Some of these are forming an available biomass for phytochemical investigations, including the assessment of biological activities of the isolated compounds. The alpine or polar region are characterised by highly stressful environmental conditions for many organisms, but lichens are among the dominating organisms in these habitats. In the performant mutual protective system, lichen fungi often accumulate high amounts of metabolites with specific physicochemical properties (UV absorbents, hydrophobicity) which help the lichens to survive. Unique secondary metabolites and polysaccharides have been isolated and tested from these organisms. Even though this has been tested until now only with a low number of compounds so far, interesting activities have been recorded. We review here some of the antimicrobial, anti-inflammatory, antiproliferative and antioxidant activities properties described. Solutions with axenic biotechnological cultivation of each symbiotic partner and particularly the mycobiont to obtain the lichen secondary metabolites are challenging to overcome the limitations for the supply of these rare compounds. Additionally, these lichens appear to harbour a diversity of culturable microorganisms from which active compounds have also been isolated recently.     

Lichens as a potential natural source of bioactive compounds: a review

Biological activity of material whether known in folk medicine or observed in planned screening program has been the starting point in the drug research. The general pattern is the isolation of active principles, elucidation their structures, followed by attempts for modulation of its activity potential by chemical modification. Lichens are valuable plant resources and are used as medicine, food, fodder, perfume, spice, dyes and for miscellaneous purposes throughout the world. Lichens are well known for the diversity of secondary metabolites that they produce. Compounds isolated from various lichen species have been reported to display diverse biological activities. Here we review the medicinal efficacy of lichen substances, which intends to explore the pharmaceutical potential of lichen substances.     

Host use of a specialist lichen-feeder: dealing with lichen secondary metabolites

Host use by herbivores is largely determined by host properties such as nutrient content and chemical defence against foragers. The impacts of these attributes on a herbivore may largely depend on its life cycle stage. Lichen species are known to differ in nutritional quality and level of chemical defence and, consequently, vary as fodder for herbivores. The aim of this study was to explore the impact of several lichen species and the presence of their secondary metabolites on their use as hosts by a specialist lichen-feeder, Cleorodes lichenaria. This study also addressed, for the first time, how a specialist lichen-feeder deals with different lichen secondary metabolites. In the beginning of their development, larvae grew better on Xanthoria parietina than on the other host lichens, whereas older larvae grew best on Ramalina fraxinea. Lichen secondary chemicals in R. fraxinea and Parmelia sulcata hindered larval growth in the beginning but after 75 days lichen secondary chemicals had no impact on the mass of larvae. Physodic acids in Hypogymnia physodes were lethal to larvae. In general, larvae metabolized 70–95% of ingested lichen secondary chemicals and the rest of these were excreted in frass. Lichen secondary metabolites in P. sulcata restrict and in H. physodes prevent their use as a host for C. lichenaria larvae. Our main finding, the ability of larvae to metabolize several lichen secondary metabolites, indicates digestive adaptation to these chemicals. No signs of sequestration of these chemicals were found.     

Tetraoxygenated naturally occurring xanthones

This review, with 350 references, gives information on the chemical study of 234 naturally occurring tetraoxygenated xanthones in 12 families, 53 genus and 182 species of higher plants, and two which are described as fungal and lichen metabolites. The value of these groups of substances in connection with pharmacological activity and therapeutic use of some species is described. The structural formulas of 135 isolated compounds, and their distribution, are also given.     

Lichen secondary metabolites affect growth of <Emphasis Type="Italic">Physcomitrella patens</Emphasis> by allelopathy

Lichen secondary metabolites can function as allelochemicals and affect the development and growth of neighboring bryophytes, fungi, vascular plants, microorganisms, and even other lichens. Lichen overgrowth on bryophytes is frequently observed in nature even though mosses grow faster than lichens, but there is still little information on the interactions between lichens and bryophytes.In the present study, we used extracts from six lichen thalli containing secondary metabolites like usnic acid, protocetraric acid, atranorin, lecanoric acid, nortistic acid, and thamnolic acid. To observe the influence of these metabolites on bryophytes, the moss Physcomitrella patens was cultivated for 5 weeks under laboratory conditions and treated with lichen extracts. Toxicity of natural mixtures of secondary metabolites was tested at three selected doses (0.001, 0.01, and 0.1 %). When the mixture contained substantial amounts of usnic acid, we observed growth inhibition of protonemata and reduced development of gametophores. Significant differences in cell lengths and widths were also noticed. Furthermore, usnic acid had a strong effect on cell division in protonemata suggesting a strong impact on the early stages of bryophyte development by allelochemicals contained in the lichen secondary metabolites.Biological activities of lichen secondary metabolites were confirmed in several studies such as antiviral, antibacterial, antitumor, antiherbivore, antioxidant, antipyretic, and analgetic action or photoprotection. This work aimed to expand the knowledge on allelopathic effects on bryophyte growth.     

Anticancer evaluation of structurally diverse Amaryllidaceae alkaloids and their synthetic derivatives

Plants of the Amaryllidaceae family have been under intense scrutiny for the presence of the specific metabolites responsible for the medicinal properties associated with them. The study began in 1877 with the isolation of alkaloid lycorine from Narcissus pseudonarcissus and since then more than 100 alkaloids, exhibiting diverse biological activities, have been isolated from the Amaryllidaceae plants. Based on the present scientific evidence, it is likely that isocarbostyril constituents of the Amaryllidaceae, such as narciclasine, pancratistatin and their congeners, are the most important metabolites responsible for the therapeutic benefits of these plant species in the folk medical treatment of cancer. Notably, Narcissus poeticus L., used by the ancient Greek physicians, is now known to contain about 0.12 g of narciclasine per kg of fresh bulbs. The focus of the present research work is the chemistry and biology of these compounds as specifically relevant to their potential use in medicine. In particular, the anticancer evaluation of lycorine, narciclasine as well as of other Amaryllidaceae alkaloids and their synthetic derivatives are presented in this paper. The structure–activity relationships among some groups of Amaryllidaceae alkaloids will be discussed.     

Phytochemical and antioxidant studies of Laurera benguelensis growing in Thailand

The aim of this study was to investigate metabolites of the lichen Laurera benguelensis. A high-performance liquid chromatographic (HPLC) method has been developed for the characterization of xanthones and anthraquinones in extracts of this lichen. Lichexanthone, secalonic acid D, norlichexanthon, parietin, emodin, teloschistin and citreorosein were detected in the lichen samples, which were collected from two places in Thailand. Components of the lichen were identified by relative retention time and spectral data. This is the first time that a detailed phytochemical analysis of the lichen L. benguelensis was reported and this paper has chemotaxonomic significance because very little has been published on the secondary metabolites present in Laurera species. Some of the metabolites were detected for the first time in the family Trypetheliaceae. The results of preliminary testing of benzene extract and its chloroform and methanol fractions showed that all samples showed a weak radical scavenging activity. The chloroform extract showed the highest antioxidant activity.     

中国地衣学现状综述

本文对中国地衣学在多样性、系统性及其物种与基因资源生物学研究方面进行了综述。文中强调了多样性和系统性研究在自然界地衣资源与研究开发之间的桥梁作用。论述了地衣结皮固沙、固碳及其基因资源在沙漠生物地毯工程中的意义。揭开了不产次生代谢产物的地衣秘密及其破解途径。     

Intraspecific chemical variation within the crustose lichen genus Haematomma: anthraquinone production in selected cultured mycobionts as a response to stress and nutrient supply

The mycobionts isolated from selected species of Haematomma (Haematomma africanum, Haematomma fenzlianum, Haematomma flourescens, Haematomma persoonii, Haematomma stevensiae) have been successfully cultured. The chemical profile of the mycobionts could be effectively influenced and modulated by varying the composition of the nutrient medium using alternative carbohydrates (glucose, sucrose, and polyols). Under artifical laboratory conditions and simulated environmental stress (exposure to UV light, desiccation, and lower temperatures) the mycobionts began producing typical secondary lichen metabolites after an incubation time of 5–6 months. Modified Lilly and Barnett medium (LBM) and Murashige Skoog Medium favoured the production of depsides such as sphaerophorin and isosphaeric acid. Surprisingly, the mycobiont from H. stevensiae in modified MS medium produced two anthraquinones in the mycelia, haematommone at the base and russulone in the upper parts of the mycelium. By contrast, the natural lichen only produced these anthraquinones in the reddish orange apothecia. The mycobiont from H. flourescens only produced the expected lichexanthone in LBM, enriched with the polyols, sorbitol and mannitol. Once the media requirements and environmental stress factors that trigger polyketide production in lichen mycobionts have been determined, it is possible to obtain a particular lichen product by a completely defined procedure. Using such knowledge, we should be able to study polyketide expression in mycobionts under optimized culture conditions for various genetic applications.     

Biocidal effect of lichen secondary metabolites against rock-dwelling microcolonial fungi,cyanobacteria and green algae

The use of commercial biocides in outdoor environments is increasingly discouraged because of their ecotoxicity, new methods being thus invoked to control patinas of biological origin on the stone cultural heritage. The effects of secondary metabolites (usnic acid, norstictic acid, parietin) produced by saxicolous lichens, natural competitors of rock dwelling microorganisms, were investigated in vitro against microcolonial fungi (MCF: Coniosporium apollinis, Coniosporium perforans, Coniosporium uncinatum, Phaeococcomyces-like sp.), coccoid cyanobacteria (Chroococcus minutus) and green algae (Scenedesmus ecornis) which commonly occur on stonework. An acetone/water 10/90 vol/vol mixture was screened as suitable to solubilise the lichen metabolites and to not affect the bioassay results. Benzalkonium chloride 1% was used as positive control.All the three metabolites (approx. 10⁻² mM) inhibited the growth of the assayed MCF species, displaying the same effect of benzalkonium chloride. Chroococcus and Scenedesmus exhibited sensibility to the lichen metabolites when exposed to high incubation temperatures (35 °C), chemicals and temperature synergically yielding percentage decreases of intact cells with red chlorophyll epifluorescence. These findings suggest lichen secondary metabolites as allelopathic agents against rock dwelling microorganisms and as potential natural sources for their control on stone materials in restoration and conservation programmes. In this perspective, the detection of a negligible chromatic alteration (ΔE<0.5) caused by LSM to the white Carrara marble is reported as the first step of the necessary extensive evaluation of the LSM-stone material interactions.     

Advanced methods for the study of the chemistry and the metabolism of lichens

Lichens are compound entities of a fungal partner (“mycobiont”) and one or more photosynthetically active algae or cyanobacteria (“photobionts”). The organisms live in an intimate, symbiotic association which has been classified as a mutualistic or controlled parasitic relationship. Several metabolites from lichens display unique structures with unknown functions, and only a few model species have been analysed comprehensively. The complex metabolic interplay between the organisms in lichens is also incompletely understood. Earlier experiments with ¹⁴C-labelled precursors indicated that the photobionts produce from CO₂ glucose or sugar alcohols (e.g. ribitol and arabitol) which are then transferred to the mycobionts. In the fungi, these compounds are believed to be converted into mannitol serving as the carbon and energy source in the downstream metabolic processes. Recent methodological developments in spectroscopy and “systems biology” now enable a concise analysis of the metabolite profiles, networks and fluxes by non-targeted quantitative approaches. In this review, we summarize the current knowledge about lichen metabolism and report on the potential of the advanced methods to reinvestigate lichen chemistry and metabolism on a quantitative basis.     

地衣型真菌次级代谢产物抗菌活性初步研究

地衣是一种传统的民族药物,能产生多种具有活性的物质。该研究对地衣型真菌(Xanthoria elegans,Myelochroa indica,Ramalina peruviana,Cladonia macilenta,Nephromopsis pallescens,Cladonia coccifera)进行液体培养,2个月后,培养液用乙酸乙酯萃取后获得初提物。该研究采用抑菌圈法评价地衣型真菌初提物对7种致病细菌(Bacillus subtilis,Bacillus cereus,Vibrio parahaemolyticus,Straphylococcus haemolyticus,Pseudomonas aeruginosa,Staphylococcus aureus,Micrococcus luteus)的抗菌活性,并测定最低抑菌浓度(MIC)。结果表明:6种地衣型真菌的初提物均具有一定的抗菌活性,且不同培养基对地衣型真菌产生抗菌物质有显著影响。其中,R.peruviana在MY液体培养基中所产生的次级代谢产物对金黄色葡萄球菌、藤黄微球菌、溶血性葡萄球菌、铜尿假单胞菌具有抑制效果,但在YMG培养基中所得初提物对供试7种致病细菌不具有抑菌效果。X.elegans在YMG培养基中所得初提物对枯草芽孢杆菌具有明显抗菌活性,其抑菌圈直径可达17.77 mm。该研究证实不同地衣型真菌液体培养初提物具有抗菌活性,不同的培养基也直接影响地衣型真菌抗菌效果。该研究结果为地衣型真菌的进一步研究及民族药的开发利用奠定了基础。     

Lichens: a promising source of antibiotic and anticancer drugs

Lichens are symbiotic associations between fungi and a photosynthetic alga and/or cyanobacteria. Lichenized fungi have been found to produce a wide array of secondary metabolites, most of which are unique to the lichenized condition. These secondary metabolites have shown an impressive range of biological activities including antibiotics, antifungal, anti-HIV, anticancer, anti-protozoan, etc. This review focuses primarily on the antibiotic and anticancer properties of lichen secondary chemicals. We have reviewed various publications related to antibiotic and anticancer drug therapies emphasizing results about specific lichens and/or lichen compounds, which microbes or cancer cells were involved and the main findings of each study. We found that crude lichen extracts and various isolated lichen compounds often demonstrate significant inhibitory activity against various pathogenic bacteria and cancer cell lines at very low concentrations. There were no studies examining the specific mechanism of action against pathogenic bacteria; however, we did find a limited number of studies where the mechanism of action against cancer cell lines had been explored. The molecular mechanism of cell death by lichen compounds includes cell cycle arrest, apoptosis, necrosis, and inhibition of angiogenesis. Although lichens are a reservoir for various biologically active compounds, only a limited number have been tested for their biological significance. There is clearly an urgent need for expanding research in this area of study, including in depth studies of those compounds which have shown promising results as well as a strong focus on identifying specific mechanisms of action and extensive clinical trials using the most promising lichen based drug therapies followed by large scale production of the best of those compounds.     

Enzyme cytochemical techniques for metabolic mapping in living cells, with special reference to proteolysis.

Specific enzymes play key roles in many pathophysiological processes and therefore are targets for therapeutic strategies. The activity of most enzymes is largely determined by many factors at the post-translational level. Therefore, it is essential to study the activity of target enzymes in living cells and tissues in a quantitative manner in relation to pathophysiological processes to understand its relevance and the potential impact of its targeting by drugs. Proteases, in particular, are crucial in every aspect of life and death of an organism and are therefore important targets. Enzyme activity in living cells can be studied with various tools. These can be endogenous fluorescent metabolites or synthetic chromogenic or fluorogenic substrates. The use of endogenous metabolites is rather limited and nonspecific because they are involved in many biological processes, but novel chromogenic and fluorogenic substrates have been developed to monitor activity of enzymes, and particularly proteases, in living cells and tissues. This review discusses these substrates and the methods in which they are applied, as well as their advantages and disadvantages for metabolic mapping in living cells.     

Anti-protease and Immunomodulatory Activities of Bacteria Associated with Caribbean Sponges

Marine sponges and their associated bacteria have been proven to be a rich source of novel secondary metabolites with therapeutic usefulness in cancer, infection, and autoimmunity. In this study, 79 strains belonging to 20 genera of the order Actinomycetales and seven strains belonging to two genera of the order Sphingomonadales were cultivated from 18 different Caribbean sponges and identified by 16S rRNA gene sequencing. Seven of these strains are likely to represent novel species. Crude extracts from selected strains were found to exhibit protease inhibition against cathepsins B and L, rhodesain, and falcipain-2 as well as immunomodulatory activities such as induction of cytokine release by human peripheral blood mononuclear cells. These results highlight the significance of marine sponge-associated bacteria to produce bioactive secondary metabolites with therapeutic potential in the treatment of infectious diseases and disorders of the immune system.     

Secondary metabolites in plants: transport and self-tolerance mechanisms

Plants produce a host of secondary metabolites with a wide range of biological activities, including potential toxicity to eukaryotic cells. Plants generally manage these compounds by transport to the apoplast or specific organelles such as the vacuole, or other self-tolerance mechanisms. For efficient production of such bioactive compounds in plants or microbes, transport and self-tolerance mechanisms should function cooperatively with the corresponding biosynthetic enzymes. Intensive studies have identified and characterized the proteins responsible for transport and self-tolerance. In particular, many transporters have been isolated and their physiological functions have been proposed. This review describes recent progress in studies of transport and self-tolerance and provides an updated inventory of transporters according to their substrates. Application of such knowledge to synthetic biology might enable efficient production of valuable secondary metabolites in the future.     

Exosomes: A New Weapon to Treat the Central Nervous System

The potential of exosomes to treat central nervous system (CNS) pathologies has been recently demonstrated. These studies make way for a complete new field that aims to exploit the natural characteristics of these vesicles, considered for a long time as side products of physiological cellular pathways. Recently, however, the biological significance of exosomes has been evaluated and exosomes can now be viewed upon as new relevant functional entities for development of novel therapeutic strategies. In this review, we aim to summarize the state-of-the-art role of exosomes in the CNS and to speculate about possible future therapeutic applications of exosomes. In particular, we will speculate about the use of these vesicles as a substitute of cell-based therapies for the treatment of brain damage and review the potential of exosomes as drug delivery vehicles for the CNS.     

Some advances in the study of lichens since the time of E. M. Holmes

Some of the principal advances in our knowledge of lichens made since the time of E. M. Holmes are briefly reviewed. Floristic studies on British lichens have continued and distribution maps of particular species are now being produced. These studies have shown that many species have disappeared from areas affected by air pollution by sulphur dioxide. Lichen taxonomy has seen the acceptance of chemical characters as valid taxonomic criteria, and the realization that ascus types, ascocarp structure and ascocarp ontogeny are more important at the generic level than ascospore characters alone. Lichens are not now thought of as a single taxonomic group but as fungi related to various orders of non-lichenized Ascomycotina, united only in their common method of nutrition (symbiosis with algae); some lichenized and non-lichenized species with identical types of ascocarps are consequently being treated as members of the same genus. Considerable progress has been made in describing lichen communities characteristic of particular substrates and situations and in understanding the factors which may affect the occurrence of particular species. Lichens have proved to be particularly valuable indicators of mean air pollution levels and scales for the estimation of these levels based solely on the lichen vegetation have been devised. Physiological and ultrastructural studies on lichens have progressed rapidly in the last decade. Carbohydrates have been demonstrated to move from the algal to the fungal component and the mobile carbohydrates found to differ in lichens with different genera of algae. The algal and fungal partners have been isolated and studied in pure culture. Most attempts to synthesize lichens have met with little success but in 1970 a thallus was formed from its components in culture. It is now possible to grow some lichens under controlled conditions in the laboratory and these techniques, together with transplant experiments, will clearly assume considerable importance in the investigation of taxonomic and ecological problems in the future. Little progress has been made in the study of lichen cytology or the factors affecting the establishment of lichen thalli. Lichens constitute an important part of some ecosystems and their decline due to anthropogenic influences is consequently a matter of concern for all biologists.     

Biophysical and biochemical characterization of active secondary metabolites from Aspergillus allahabadii

Fungal secondary metabolites are a diverse group of natural chemical products with physiological relevance. We aimed to identify bioactive secondary metabolites from Aspergillus allahabadii. We used “activity-guided fractionation” strategy for the isolation of secondary metabolites. Crude extracts showed good antibacterial activity. Two antibacterial secondary metabolites have been isolated from the crude extract. Chemical characterization of these compounds was performed using biophysical techniques (FT-IR, NMR, and mass spectrometry). Structural characterization confirmed these to be pyrone derivatives: 3-hydroxy 2-methyl 4-pyrone and 5-hydroxy-2-(hydroxymethyl)-4H-pyrone. These bioactive pyrone derivatives have been identified as maltol and kojic acid. From our initial observations, we infer that these pyrone derivatives have potent antimicrobial, antioxidant, antidiabetic, and mosquito larvicidal activities and no cytotoxicity. These compounds could have potential therapeutic and biomedical applications, but further mechanistic studies using animal models are very much necessary.     

Antiinflammatory and immunomodulating properties of fungal metabolites

We discuss current information on the ability of extracts and isolated metabolites from mushrooms to modulate immune responses. This can result in a more enhanced innate and acquired disease resistance. The major immunomodulating effects of these active substances derived from mushrooms include mitogenicity and activation of immune effector cells, such as lymphocytes, macrophages, and natural killer cells, resulting in the production of cytokines, including interleukins (ILs), tumor necrosis factor alpha (TNF)-alpha, and interferon gamma (INF)-gamma. In particular, the ability of selective mushroom extracts to modulate the differentiation capacity of CD4(+) T cells to mature into T(H)1 and/or T(H)2 subsets will be discussed. As a consequence these extracts will have profound effects in particular diseases, like chronic autoimmune T(H)1-mediated or allergic T(H)2-mediated diseases. Immunosuppressive effects by mushroom components have also been observed. The therapeutic effects of mushrooms, such as anticancer activity, suppression of autoimmune diseases, and allergy have been associated with their immunomodulating effects. However, further studies are needed to determine the molecular mechanisms of the immunomodulating effects of mushrooms metabolites both individually and in complex mixtures, for example, extracts.