Fumarprotocetraric Acid Research Articles

The algal partnership is associated with quantitative variation of lichen specific metabolites in Cladonia foliacea from Central and Southern Europe

AbstractLichen specific metabolites (LSMs) have interesting biological activities and quantitative variations may be present intraspecifically. For example, variations in medullary fumarprotocetraric acid (FA) and cortical usnic acid (UA) were observed in the lichen Cladonia foliacea, but the mechanism of variation is not well understood. The current study aimed to characterise the quantitative variation of FA and UA and to investigate the association between lichen metabolite content and ecological / biological variables. Fungal and algal trees were constructed using fungal (nrITS, RPB2) and algal (nrITS) loci, respectively. Using a chiral chromatographic method, the contents of (-)-UA were determined in 29 C. foliacea specimens and range from 6.88 to 34.27 mg/g dry wt. The FA contents were lower and varied from 1.44 to 9.87 mg/g dry wt. Although the fungal tree showed two well resolved clades, no significant differences of UA or FA contents were found between the two fungal clades. However, a significantly higher UA/FA ratio as well as a unique habitat were found to be associated with specimens which contained the alga Asterochloris lobophora than those specimens associated other Asterochloris algae. Taking all predictive variables into account (i.e. substrate type, elevation, collection season, photobiont identity), the multivariate data analysis indicated that photobiont identity explained most of the variance of LSM contents in C. foliacea. Thus future LSM biosynthetic studies should take the photobiont into consideration when dealing with intraspecific quantitative variation.

Characteristics, Chemical Analysis and Biological Activities of Methanol Extracts of Lichens Pleurosticta Acetabulum and Cladonia Subulata

Abstract The aim of this study is to investigate the chemical composition of methanol extracts of the lichens Pleurosticta acetabulum and Cladonia subulata and their antioxidant, and anticancer activities. The phytochemical analysis of the extracts of lichens was determined by HPLC-UV method. The predominant phenolic compounds in these extracts were norstictic acid and salazinic acids in P. acetabulum, while hypoprotocetraric acid and fumarprotocetraric acid were the major metabolites detected in C. subulata. Total phenolics and flavonoids in the extracts were determined spectrophotometrically, with the varied amount from 21.31 to 73.45 mg GA/g and from 8.48 to 15.42 mg RU/g, respectively. The lichen extracts showed comparable and strong antioxidant activity, exhibited higher DPPH and hydroxyl radical scavengings, inhibitory activity towards lipid peroxidation and reducing power. Cytotoxic effects of lichens were tested against HeLa S3 and LS174 cell lines using MTT method.The cytotoxic effects of P. acetabulum and C. subulata extracts toward two cancer cell lines were in the range from 39.17 to >200 μg/mL IC50 value. The present study showed that the tested extracts of lichens demonstrated important antioxidant and anticancer effects. That suggests that these lichens can be used as new sources of the natural antioxidants and anti-cancer compounds.

HS-SPEM-GC-MS Analyses of Volatiles and Transcriptome Analysis of Camellia tetracocca in Response to Feeding by Empoasca onukii Matsuda

The tea leafhopper (Empoasca (Matsumurasca) onukii Matsuda, 1952) is the predominant pest infesting tea and causes significant losses in yield. Plants have evolved multiple defense responses to protect themselves from insect herbivores. Camellia tetracocca, a kind of ancient tea with local popularity, is unique to southwest China. This ancient tea is often attacked by the tea leafhopper. However, there are no relevant reports on changes in volatile components and defense signaling pathways after Camellia tetracocca has been attacked by E. onukii. To reveal changes in the responses of the volatile components of C. tetracocca to E. onukii, the headspace sampling technique and GC-MS were adopted to determine and analyze the volatiles. Furthermore, to unveil the defense mechanisms of C. tetracocca in response to E. onukii, transcriptome analysis was applied to damaged tea leaves and healthy leaves. The GC-MS analysis showed significant increases in the linalool and (E)-2-hexenal contents. The transcriptome analysis revealed a set of core pest-responsive genes (3074 upregulated and 5098 downregulated). Most of the enriched pest-responsive genes are mainly involved in plant hormone signal transduction photosynthesis, terpenoid backbone biosynthesis, alpha-linolenic acid metabolism, and the NF-κB signaling pathway. A conjoint analysis of the transcriptome and metabolome indicated that N2-acetylornithine, D-malic acid, theogallin, fucosyllactose, 5-caffeoylquinic acid, fumarprotocetraric acid, and arabinogalactose have a significant correlation with LOC114279735, LOC114317534, LOC114282078, LOC114317768, and LOC114263181. This study lays a foundation for further research on the breeding and cultivation of insect-resistant germplasm resources for C. tetracocca and on the promotion and development of the plant.

Effects of Exhaustion Gases from Biodiesel Blends on the Lichen Cladonia verticillaris: A Complimentary Evaluation

The objective of this study was to evaluate the effects of gases emitted by blends of biodiesel with petroleum diesel at intermediate concentrations (between 10 % and 40 %) on the biochemistry of the lichen Cladonia verticillaris (Raddi) Fr. Samples of the respective species were exposed to the gases during a period of sixty minutes, using a stationary engine, with blends of 20 % (B20), 25 % (B25) and 30 % (B30) of biodiesel, from the oil from cotton, to petroleum diesel. In the period of 30, 60 and 90 days, the concentrations of primary metabolites (chlorophylls and pheophytins) and secondary (protocetraric acid and fumarprotocetraric acid) were determined. A control group was used for comparison purposes. The gases emitted during the combustion process were also evaluated at three times (0 minutes, 30 minutes and 60 minutes). The results did not show significant differences for chlorophylls, when all treatments and control materials were compared, however, variations were observed for the other metabolites. The concentration of protocetraric acid was higher compared to fumarprotocetraric acid, indicating that there were blocks in the synthesis of the metabolic pathway. Cluster analysis showed that for groups B20 and B25, pheophytins are related to CO2, while for B30 their chlorophylls are related to NO2. The results demonstrate that the respective mixtures do not affect the integrity of the species, confirming the recommendation to use mixtures of biodiesel with petroleum diesel up to 40 %. Keywords: biofuels, environmental impact, living organisms, lichen substances, biomonitoring.

The Key Features of Catechols and α,β Unsaturated Carbonyl Moieties: Interaction With α‐syn Hydrophobic peptide and Activation of Catecholamines Pathway in Cells

AbstractParkinson's disease (PD) is the second most prevalent neurodegenerative disorder worldwide, and the treatment focuses on delivering L‐DOPA. In this work, we isolated and tested several compounds against α‐synuclein and the hydrophobic peptide 71VTGVTAVAQKTV82 including flavonols (kaempferol, quercetin, and isorhamnetin), isoflavone (genistein) and flavone (luteolin), and compounds with α, β unsaturated carbonyl moieties such as chlorogenic acid and the depsidone fumarprotocetraric acid. Most compounds inhibit both α‐synuclein and hydrophobic peptide fibrillization. Moreover, ITC experiments showed a Kd varying from 9 to 20 μM, and ΔH values vary from −1.94 to −10.5 among the compounds. Docking experiments showed the intermolecular interactions within the sites 2, 9, and 3/13 of α‐synuclein, and with the hydrophobic peptide. In cultured cells, the presence of the compounds showed that most of them can promote cell proliferation and differentiation. Considering that treatments for neurodegenerative disorders, including PD, is only palliative the evaluation of these compounds that can prevent the fibrillization of α‐synuclein and stimulate the catecholamines pathway is promising.

Larvicidal activity and docking study of Ramalina complanata and Cladonia verticillaris extracts and secondary metabolites against Aedes aegypti

The Aedes aegypti mosquito is the main vector of the arboviruses that cause Zika, Yellow Fever, Dengue, and Chikungunya, incapacitating diseases that may even be fatal. There are still no vaccines for any of these viruses. Therefore, the only way to contain outbreaks is to control the vector that transmits them. In the present study, we investigated the larvicidal activity of lichen extracts (Ramalina complanata (Sw.) Ach., 1810 and Cladonia verticillaris (Raddi) Fr.), isolating components that were used against larvae of the mosquito A. aegypti. The ether extract from R. complanata exhibited an LC50 = 5.9 µg.ml−1, while a compound isolated from this species, divaricatic acid, had an LC50 = 27.1 µg.ml−1. The acetone extract of C. verticilaris exhibited an LC50 = 17.4 µg.ml−1 and the isolated compound fumarprotocetraric acid (FUM) had an LC50 = 13.6 µg.ml−1. These results demonstrate that extracts and isolated compounds from the two lichen species have strong larvicidal activities. FUM is the most active compound in our investigation. The molecular docking studies of divaricatic acid and FUM demonstrated that they bind to the active site of AaAChE1 in an energetically favorable manner. The TRP286 residue may have contributed to the more pronounced activity of fumarprotocetraric acid compared to divaricatic acid. Other aspects related to the more pronounced activity of fumarprotocetraric acid are its molecular weight and higher solubility. Our results provide evidence of the importance of studying lichen secondary metabolites as natural sources of insecticides and investigating possible interactions with AaAChE1.

Antioxidant activity and enzymatic of lichen substances: A study based on cyclic voltammetry and theoretical

The antioxidant activity of nine lichen substances, including methylatrarate (1), methyl haematommate (2), lobaric acid (3), fumarprotocetraric acid (4), sphaerophorin (5), subsphaeric acid (6), diffractaic acid (7), barbatolic acid (8) and salazinic acid (9) has been determined through cyclic voltammetry. The compounds 1–4 presented slopes close to the Nernst constant of 0.059 V, indicating a 2H+/2e− relation between protons and electrons, as long as the compounds 5, 6, 7, 8, and 9 present slopes between 0.037 V and 0.032 V, indicating a 1H+/2e− relation between protons and electrons. These results show a high free radical scavenging activity by means of the release of H+, suggesting an important antioxidant capacity of these molecules. Theoretical calculations of hydrogen bond dissociation enthalpies (BDE), proton affinities (PA), and Proton Transfer (PT) mechanisms, at M06-2x/6-311+G(d,p) level complement the experimental results. Computations support that the best antioxidant activity is obtained for the molecules (3, 4, 5, 6, 7 and 8), that have a carboxylic acid group close to a phenolic hydroxyl group, through hydrogen atomic transfer (HAT) and sequential proton loss electron transfer (SPLET) mechanisms. Additional computations were performed for modelling binding affinity of the lichen substances with CYPs enzymes, mainly CYP1A2, CYP51, and CYP2C9*2 isoforms, showing strong affinity for all the compounds described in this study.

Microcrystallization method use for assessing the content of secondary metabolites of lichens

The paper presents photographs of crystals of fumarprotocetraric, evernic, usninic, solorinic, olivetoric, perlatolinic, physodalic, salazinic, tamnolic acids and atranorin obtained from their acetone extracts of lichens. The shape of the crystals is specific to each substance, and their number, size and degree of aggregation depend on the concentration in the solution. Stellate crystals have atranorine, salazinic and evernic acids, with atranorine having the largest and salacic acid having the smallest; rodshaped branched crystals form solorinic and fumarprotocetraric acids, wider in the first, but longer in the second; the oval shape of crystals in the larger tamnolic and small and narrow perlatolinic acid; usnic, physodalic and olivetoric acids form unique crystal forms prismatic, needle branched and pinnate, respectively. It is possible to observe crystals of secondary metabolites of lichens with a conventional light microscope under a 40 lens without immersion oil. For the subsequent comparative assessment of the amount of substances, it is necessary to standardize the suspension of the studied lichen thallus, use a strictly fixed volume of acetone, view at least four sides of the dried drop on the slide and identify their total number in points, noting the presence, degree of branching and aggregation of crystals into conglomerates.

Phytochemical and biological characterization of acetone extracts from the lichen Cladonia coniocraea and Cladonia ramulosa growing in Serbia

A comparative study of aromatic secondary metabolites and antioxidant, antimicrobial, and anticancer potential of two lichen species Cladonia coniocraea and Cladonia ramulosa is presented in this paper. HPLC-UV analysis revealed the presence of depsidone fumarprotocetraric acid and depside sekikaic acid as major aromatic metabolites in both tested species but in different amounts. The antioxidant activity was evaluated by free radical scavenging and reducing power assays. In both assays the extracts showed weak activity (IC50>1000 μg/ml, while absorbances for reducing power were from 0.0118 to 0.1675). The total content of phenol and flavonoid in extracts was examined using Folin–Ciocalteu reagent and aluminium chloride method and the obtained values were expressed as pyrocatechol equivalents, and as rutin equivalents, respectively. Further, the antimicrobial activity was estimated by determination of the minimal inhibitory concentration by the broth microdilution method against five bacterial and 10 fungal species. Tested extracts showed similar antimicrobial activity with minimum inhibitory concentration values ranging from 0.156 to 20 mg/ml. Finally, the cytotoxic activity was tested using MTT method on the human epithelial carcinoma (Hela), human lung carcinoma (A549) and human colon carcinoma (LS174) cells. The strongest cytotoxic activity with IC50 value of 185.59 μg/ml expressed extract of C. ramulosa toward Hela cells. Obtained results indicate that these lichens showed the potential for further investigation and possible biopharmaceutical application.

The Role of Secondary Metabolites and Bark Chemistry in Shaping Diversity and Abundance of Epiphytic Lichens

Diversity of secondary lichen metabolites was studied in epiphytic lichens on six phorophytes—spruce, pine, birch, alder, aspen and poplar in the Middle Urals of Russia. Atranorin, usnic, fumarprotocetraric acid, zeorin, and gyrophoric acid were found in 31, 24, 23, 18, and 14 species, respectively, of 237 taxa collected. Seventy-seven species (i.e., 32% of total species documented) contained no secondary metabolites. Spectra of secondary metabolites of fruticose and foliose lichens varied on different phorophytes, while in crustose species the strong dependence on the tree species was not detected. This is different to the pH dependence of saxicolous lichens where crustose lichens were more susceptible to the rock chemistry. The results of Canonical Correspondence Analysis reveal the affinity of species containing depsides, depsidones or usnic acid to acidic substrata and those lacking secondary metabolites or containing terpenes and antraquinones to the pH-neutral bark. We suppose that phenolic compounds and flavonoids, as chemical constituents of bark, may interact with lichen symbioses and elements in phellem, and similarly to the lichen acids shape the affinity of species to the substrata.

Two New Fumarprotocetraric Acid Lactones Identified and Characterized by UHPLC-PDA/ESI/ORBITRAP/MS/MS from the Antarctic Lichen Cladonia metacorallifera

Lichens are symbiotic organisms between algae and fungi, which are makers of secondary compounds named as lichen substances. Hyphenated techniques have significantly helped natural product chemistry, especially UHPLC/ESI/MS/MS in the identification, separation, and tentative characterization of secondary metabolites from natural sources. Twenty-five compounds were detected from the Antarctic lichen Cladonia metacorallifera for the first time using UHPLC-PDA/ESI/Orbitrap/MS/MS. Compounds 5 and 7 are reported as new compounds, based on their MS/MS fragmentation routes, and considered as fumarprotocetraric acid derivatives. Besides, ten known phenolic identified as orsellinic acid, ethyl 4-carboxyorsellinate, psoromic acid isomer, succinprotocetraric acid, siphulellic acid, connorstictic acid, cryptostictic acid, lecanoric acid, lobaric acid and gyrophoric acid are noticed for the first time in the Cladonia genus.

Comparative analyses of antioxidant, cytotoxic, and anti-inflammatory activities of different Cladonia species and determination of fumarprotocetraric acid amounts

In traditional medicine, Cladonia lichens are used for several disorders like hypertension, headaches, dizziness, cough, and menstrual disorders. The study aimed to determine the biological activities of 70% methanol extracts of nine Cladonia species from Turkey. The chemical composition was analyzed via spectrophotometric and chromatographic (HPLC) techniques. DPPH● (1,1-diphenyl-2-picrylhydrazyl) and ABTS+● (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) radical scavenging activities, inhibition of β-carotene/linoleic acid co-oxidation, and cytotoxicity on A549 and COLO205 cell lines were studied. Anti-inflammatory activity was assessed on the Raw 264.7 mouse macrophage cell line by determining nitric oxide (NO) levels. Among the studied Cladonia species Cladonia firma was found the most active extract in ABTS+● test system (2.58±0.07 mmol/L/Trolox) and also in the cytotoxicity and anti-inflammatory experiments. Cladonia pocillum, Cladonia rangiformis, and Cladonia foliaceae were also found to be active. Fumarprotocetraric acid amounts are between 1.89 -23.82 mg/gextract and fumarprotocetraric acid content did not show linearity with biological activities. It has been proved with this study that lichens traditionally used in treatment have as much biological activity as plants and can be evaluated in pharmaceuticals and cosmetics.

Cladonia verticillaris (lichen) indicates negative impacts derived from the combustion of biodiesel blends: an alert for the environmental management for biofuels use.

The use of biodiesel blends with petroleum diesel in vehicular engines demands the evaluation of the possible impacts and effects of the gases emitted from their combustion on the environment. Among studies on these questions, biomonitoring using lichens is a viable alternative, given their interactions with the elements dispersed in the atmosphere, as well as its sensitivity and capacity to retain contaminants. In this study, we analyzed the effects of gas emissions from the combustion of biodiesel mixture with petroleum diesel on Cladonia verticillaris thalli. Samples of the lichen (10g) were exposed to the gases emitted by the exhaust of the generator engine during the combustion process of biodiesel mixtures to petroleum diesel (7% (B7), 10% (B10), 40% (B40), 50% (B50), and 70% (B70)). At 90days after exposure, samples were analyzed for n-alkane profiles, thallus morphology, photosynthetic pigment contents, and secondary lichen metabolites (protocetraric and fumarprotocetraric acids). Sets B7 and B10 showed better resistance of the lichen to pollutants. Set B40 showed a high stress evidenced by the chain elongation of n-alkanes structure and high chlorophyll production, presenting high morphological damages when compared to the control sets, B7 and B10. The results showed significant reductions of n-alkanes profiles for mixtures with high concentrations of biodiesel (B50 and B70), as well as decreases in the chlorophyll content. These groups showed an increase in the synthesis of secondary metabolites, corroborating the hypothesis that high concentrations of biodiesel in the mixture with petroleum diesel have greater impacts on the lichen. Schematic model for demonstration of using the lichen Cladonia verticillaris as biomonitor of effects from gas emissions from the combustion of biodiesel blends with petroleum diesel by a stationary engine.

INFLUENCE OF TEMPERATURE CONDITIONS ON THE ACCUMULATION OF PRIMARY AND SECONDARY METABOLITES BY LICHENS FLAVOCETRARIA CUCULLATA AND CETRARIA LAEVIGATA

The study of the effect of temperature on the accumulation of primary and secondary metabolites by lichens Flavocetraria cucullata (Bellardi) Kärnefelt & Thell and Cetraria laevigata Rass was carried out. Lichen samples were taken out from under the snow (-20 °C) together with the soil substrate and transferred to the climatic chamber. Then the temperature in the climatic chamber was sequentially increased to +10 and +20 °C. The lichen exposure was carried out for 30 days for each temperature regime. The analysis of primary metabolites was performed by gas chromatography-mass spectrometry. It was shown that in lichens F. cucullata and C. laevigata at temperatures of +10 and +20 °C, an increase in the content of mannitol, ribitol, sucrose, and hydroxyproline was observed, as well as a decrease in the content of unsaturated fatty acids as compared to the initial samples. At the same time, the content of glycerol and arabitol in the thalli of the initial lichens (-20 °C) was higher than after exposure at +10 and +20 °C, which, apparently, is associated with the cryoprotective properties of these compounds. The content of secondary metabolites in lichens was determined by high performance liquid chromatography. It was shown that the content of usnic, allo-protolichesterinic, and protolichesterinic acids in F. cucullata increased after exposure in a climatic chamber, which may be associated with a general activation of metabolic processes with an increase in temperature. At the same time, the content of fumarprotocetraric acid in C. laevigata lichens decreased at temperatures of +10 and +20 °C compared to the initial samples, which may be associated with its protective properties under the action of low-temperature stress.

The Fumarprotocetraric Acid Inhibits Tau Covalently, Avoiding Cytotoxicity of Aggregates in Cells.

Neurodegenerative disorders, including Tauopathies that involve tau protein, base their pathological mechanism on forming proteinaceous aggregates, which has a deleterious effect on cells triggering an inflammatory response. Moreover, tau inhibitors can exert their mechanism of action through noncovalent and covalent interactions. Thus, Michael’s addition appears as a feasible type of interaction involving an α, β unsaturated carbonyl moiety to avoid pathological confirmation and further cytotoxicity. Moreover, we isolated three compounds from Antarctic lichens Cladonia cariosa and Himantormia lugubris: protolichesterinic acid (1), fumarprotocetraric acid (2), and lichesterinic acid (3). The maleimide cysteine labeling assay showed that compounds 1, 2, and 3 inhibit at 50 µM, but compounds 2 and 3 are statistically significant. Based on its inhibition capacity, we decided to test compound 2 further. Thus, our results suggest that compound 2 remodel soluble oligomers and diminish β sheet content, as demonstrated through ThT experiments. Hence, we added externally treated oligomers with compound 2 to demonstrate that they are harmless in cell culture. First, the morphology of cells in the presence of aggregates does not suffer evident changes compared to the control. Additionally, the externally added aggregates do not provoke a substantial LDH release compared to the control, indicating that treated oligomers do not provoke membrane damage in cell culture compared with aggregates alone. Thus, in the present work, we demonstrated that Michael’s acceptors found in lichens could serve as a scaffold to explore different mechanisms of action to turn tau aggregates into harmless species.

Lichen Depsidones with Biological Interest.

Depsidones are some of the most abundant secondary metabolites produced by lichens. These compounds have aroused great pharmacological interest due to their activities as antioxidants, antimicrobial, and cytotoxic agents. Hence, this paper aims to provide up-to-date knowledge including an overview of the potential biological interest of lichen depsidones. So far, the most studied depsidones are fumarprotocetraric acid, lobaric acid, norstictic acid, physodic acid, salazinic acid, and stictic acid. Their pharmacological activities have been mainly investigated in in vitro studies and, to a lesser extent, in in vivo studies. No clinical trials have been performed yet. Depsidones are promising cytotoxic agents that act against different cell lines of animal and human origin. Moreover, these compounds have shown antimicrobial activity against both Gram-positive and Gram-negative bacteria and fungi, mainly Candida spp. Furthermore, depsidones have antioxidant properties as revealed in oxidative stress in vitro and in vivo models. Future research should be focused on further investigating the mechanism of action of depsidones and in evaluating new potential actions as well as other depsidones that have not been studied yet from a pharmacological perspective. Likewise, more in vivo studies are prerequisite, and clinical trials for the most promising depsidones are encouraged.

Intra-specific variation in lichen secondary compounds across environmental gradients on Signy Island, maritime Antarctic

Lichens produce various carbon-based secondary compounds (CBSCs) in response to abiotic conditions and herbivory. Although lichen CBSCs have received considerable attention with regard to responses to UV-B exposure, very little is known about intra-specific variation across environmental gradients and their role in protection against herbivory in the Antarctic. Here we report on the variation in CBSCs of two widely distributed and common Antarctic lichens, Usnea antarctica and Umbilicaria antarctica, between sites with different solar exposure (NW–SE) and along natural nitrogen (N) gradients which are associated with changing lichen-invertebrate associations on Signy Island (South Orkney Islands, maritime Antarctic). Fumarprotocetraric and usnic acid concentrations in Usnea showed no relationships with solar exposure, lichen-N or associated invertebrate abundance. However, fumarprotocetraric acid concentration was 13 times higher at inland sites compared to coastal sites along the N-gradients. Gyrophoric acid concentration in Umbilicaria was 33% lower in sun-facing (northerly exposed) habitats compared to more shaded (south-facing) rocks and declined with elevation. Gyrophoric acid concentration was positively correlated with the abundance and species richness of associated microarthropods, similar to the patterns found with lichen N. This initial investigation indicates that there can be large intraspecific variation in lichen CBSC concentrations across relative short distances (< 500 m) on Signy Island and raises further questions regarding current understanding of the role of CBSCs in Antarctic lichens in relation to biotic and abiotic pressures.

Three overlooked species of Bacidia from insular Laurimacaronesia

We discuss the taxonomy of three species of Bacidia occurring in insular Laurimacaronesia. Two of them, B. amylothelia (Vain.) Vain. and B. endoleucoides (Nyl.) Zahlbr., which were previously described from Angola and Madeira, respectively, are found here to belong in Bacidia s. str. (Ramalinaceae). Modern descriptions and illustrations are provided for the first time. Bacidia amylothelia is similar to B. areolata Gerasimova &amp; A. Beck, B. campalea (Tuck.) S. Ekman &amp; Kalb, B. fusconigrescens (Nyl.) Zahlbr., B. heteroloma (Vain.) Zahlbr., B. millegrana (Taylor) Zahlbr. and B. suffusa (Fr.) A. Schneid. and is reported here from the Canary Islands. Bacidia endoleucoides is most likely to be confused with B. absistens (Nyl.) Arnold, B. friesiana (Hepp) Körb., B. salazarensis B. de Lesd. and B. caesiovirens S. Ekman &amp; Holien and was found to be widely distributed in the Canary Islands and Azores in addition to Madeira. The third species, Bacidia deludens S. Ekman, Tønsberg &amp; van den Boom, is described here as new to science. Bacidia deludens is characterised by a greyish, crustose thallus with whitish soralia, pale apothecia with crystals in the hymenium and proper exciple, acicular ascospores with 3–19 septa, and the production of fumarprotocetraric acid as the consistently present major substance. It is described here from the Canary Islands and Madeira. Although conservatively treated here in Bacidia, we argue that it is likely to belong in the Malmideaceae. An identification key to all known species of Bacidia s. str. in insular Macaronesia is provided.

Efficiency and selectivity of ionic liquids in microwave-assisted extraction of major lichen phenolic compounds: a scalable process with recycling of ionic liquids.

Pseudevernia furfuracea, a lichen used classically for cosmetic applications, contains interesting metabolites possessing antimicrobial and anti-inflammatory or antioxidant properties. Ionic liquid combined to microwave-assisted extraction (IL-MAE) was successfully applied for metabolites extraction from Pseudevernia furfuracea. Three imidazolium and pyridinium-based ionic liquids (ILs): 1,3-dimethylimidazolium methylsulphate [C1 C1 Im][MeSO4 ], 1-ethyl-3-methylimidazolium ethylsulphate [C2 C1 Im][EtSO4 ], and N-ethylpyridinium ethylsulphate [C2 Py][EtSO4 ] were assessed for this process. The efficiency of the extraction method was evaluated using thin-layer chromatography (TLC) coupled to a Camag® spectrophotodensitometer and using high-performance liquid chromatography (HPLC) analysis. ILs under MAE showed extraction time efficiency (15 min vs. 24 h for conventional heating) and high selectivity in extracting the targeted metabolites: atranorin (AT), methyl-β-orcinol carboxylate (MOC), fumarprotocetraric acid (Fum. Ac.), and physodic acid (Phys. Ac.) despite the increased degradation of AT under MAE. We showed a tunable selectivity of ILs towards extracting metabolites by changing anion or cation due to the modification of the interaction between the IL and the metabolites. While [C2 Py][EtSO4 ] was the most efficient IL and could extract all the targeted metabolites, [C2 C1 Im][EtSO4 ] was the most selective. It fully extracted AT and partially Fum. Ac. Moreover, the lichen prepared by mixing procedure provided AT and Fum. Ac. more than the milled one. A 100 times scale-up extraction was successfully performed on mixed samples with full IL recycling after back extraction. IL-MAE is reliable for lichen metabolites extraction. The method is reproducible, scalable, with possible IL recycling, opening the door for potential industrial applications.

The amount of lichen secondary metabolites in Cladonia foliacea (Cladoniaceae, lichenised Ascomycota)

The terricolous species Cladonia foliacea (Cladoniaceae, lichenised Ascomycota) widely distributed in open, dry lowland steppe and rocky mountain grassland vegetation in Europe was chosen as a potential test organism for ecological experiments, since their thalli are producing cortical solar radiation-protective and UV screening pigment dibenzofuran usnic acid and medullary secondary substance depsidone fumarprotocetraric acid. Significant seasonal differences were found in the amounts of lichen secondary metabolites analysed by HPTLC and HPLC-PDA between summer and winter collected thalli in sandy grassland area in Hungary. The concentrations of usnic acid varied between 7.34 and 15.52 mg/g in summer collected samples and 13.90 and 21.61 mg/g in winter collected ones. A comparable amount (11.61±0.29 mg/g) was measured in pulverised samples. The concentrations of fumarprotocetraric acid varied between 0.60 and 3.01 mg/g in summer collected samples and 2.26 and 5.81 mg/g in winter collected thalli. A comparable amount (2.45±0.21 mg/g) was found in pulverised samples. The range of concentration values is comparable with data known from lichens. A higher amount of usnic acid is produced in winter probably to ensure sufficient protection also for summer. The fumarprotocetraric acid content of the medulla might contribute to the solar irradiation reflecting role of the pale lower surface lobes turning upwards in dry condition.