3-hydroxyphysodic Acid Research Articles

A Comparative Study of Isolated Secondary Metabolites from Lichens and Their Antioxidative Properties.

Free radicals play a critical role in the chemical processes that occur in all cells. Pharmaceutical companies manufacture a variety of synthetically prepared antioxidants, but it is known that many of these can be carcinogenic. As a result, efforts are being made to find natural antioxidants that do not have these side effects. Lichens may be suitable candidates because they contain secondary metabolites with proven antioxidant properties. This could be explained by the presence of compounds with phenolic groups in lichens. The radical scavenging reaction is a chemical reaction governed by stoichiometry, and our aim is to determine the efficacy of these reactions. The aim of this study is to compare metabolite activity based on the same amount of substance involved in radical scavenging, calculated in micromoles rather than weight concentration. This provides an accurate way of comparing radical scavenging activity. We tested superoxide anion scavenging activity and free radical scavenging activity of isolated lichen secondary metabolites and their mixtures in different ratios. The following compounds were isolated and tested for antioxidant activity: gyrophoric acid (Umbilicaria hirsuta), evernic acid (Evernia prunastri), physodic acid, 3-hydroxyphysodic acid, physodalic acid and atranorin (Hypogymnia physodes), and usnic acid (as a synthetic compound). Of all the tested compounds, 3-hydroxyphysodic acid, as well as mixtures containing this metabolite, showed the strongest scavenging activity. The results also demonstrated that calculation by amount of substance leads to a new consideration of antioxidant activity.

Hypogymnia papilliformis (Parmeliaceae), a new lichen from Far East Russia and China

AbstractHypogymnia papilliformis McCune, Tchabanenko & X. L.Wei is described as a new species of lichenized fungus from the Primorsky region of Russia and Shaanxi Province in China. It is a relatively rare species from mixed conifer-broadleaved forests in suboceanic climates, in mountainous areas at elevations of 600–1500 m. Most similar to H. delavayi, the species are distinguished by branching pattern and chemistry. Hypogymnia papilliformis is predominantly isotomically branched while H. delavayi usually develops subpinnate branching. The ceiling of the lobe cavity is mid brown to dark brown in H. papilliformis, while the ceiling is often pale brownish or white in H. delavayi. Also, H. papilliformis lacks 3-hydroxyphysodic acid, and thus has a K– medulla, while H. delavayi always contains 3-hydroxyphysodic acid as a major substance and is thus K+ slowly reddish brown. We provide a key to the six esorediate Hypogymnia species known from the Russian Far East.

Effects of Depsidones from Hypogymnia physodes on HeLa Cell Viability and Growth

The anti-proliferative activitiy of Hypogymnia physodes methanol extracts (ME) and its main constituents, physodalic acid (P1), physodic acid (P2), and 3-hydroxy physodic acid (P3), was tested on human cancer HeLa cell lines. Three lichen depsidones, P1, P2 and P3, were isolated from H. physodes ME using column chromatography and their structures were determined by UV, ESI TOF MS, 1H and 13C NMR. The content of P1, P2 and P3 in ME was determined using reversed-phase highperformance liquid chromatography with photodiode array detection. P1–3 represented even 70 % of the studied extract. The HeLa cells were incubated during 24 and 72 h in the presence of ME and depsidones P1, P2 and P3, at concentrations of 10–1000 μg/ml. Compounds P2 and P3 showed higher activity than compound P1. Half maximal inhibitory concentrations (IC50, μg/ml) of P1, P2, P3 and ME for 24-h incubation were 964, 171, 97 and 254 μg/ml, respectively, while for 72-h incubation they were 283, 66, 63 and 68 μg/ml. As far as we know, this is the first report on the effect of H. physodes ME and their depsidones on HeLa cells.

Effect of Hypogymnia physodes Extracts and their Depsidones on Micronucleus Distribution in Human Lymphocytes

Three lichen depsidones, physodalic acid (1), physodic acid (2), and 3-hydroxy physodic acid (3), were isolated from Hypogymnia physodes diethyl ether extract using column chromatography, and their structures determined by comparing their UV, 1H and 13C NMR spectroscopic and MS data with those given in the literature, as well as with data computed by CHEM draw ultra 11 software. The contents of 1, 2 and 3 were determined in the methanol (ME), acetone (AE), and diethyl ether (EE) extracts using reversed-phase high performance liquid chromatography with photodiode array detection. The extracts, isolated depsidones 1-3 and EE fraction F23 (consisting of 90% 2 and 3, in the ratio 5.5: 1) were evaluated for their in vitro effects on chromosome aberrations in peripheral human lymphocytes using the cytochalasin-B blocked micronucleus (CBMN) assay in doses of 1 microg/mL and 2 microg/mL of final culture solution. The frequency of MN was scored in binucleated cells, and nuclear proliferation index was calculated. It was found that 1, 2, 3, F23, and EE at 1.0 microg/mL exerted a beneficial effect on lymphocyte cells giving a significant decrease of the frequency of MN in comparison with the positive control Amifostin WR-2721.Among the tested depsidones at a concentration of 1 microg/mL, 3 exhibited the most prominent effect decreasing the frequency of MN by 30.3%, followed by 2 (28.2%) and 1 (22.0%). The extracts were less effective than the isolated depsidones.

Lichen substance concentrations in the lichen Hypogymnia physodes are correlated with heavy metal concentrations in the substratum

Lichen substances (i.e. lichen-specific carbon-based secondary compounds) are known to be involved in the uptake and immobilization of metal ions, though the biochemical mechanisms of this interaction are largely unexplained. Previous research on potential effects of lichen substances on heavy metal uptake and tolerance mostly focused on lichens in heavily polluted areas with exceptionally high metal concentrations. In the present study, we aimed at gathering information as to whether lichen substances might be involved in the fine-tuning of metal uptake even at not or low-polluted sites. Therefore, we studied lichen substance concentrations in the epiphytic lichen Hypogymnia physodes and metal concentrations in its substratum in a montane spruce forest of Germany. H. physodes produces two depsides and five depsidones, which had been shown to be involved in metal homeostasis, namely in Cu and Mn uptake, in previous laboratory experiments. The amount of lichen substances increased with increasing heavy metal concentration in the substratum, though the latter varied only in the range of a few μmolg−1 between the sample trees. Variability of lichen substance concentrations in H. physodes within the individual trees was low. Among the different lichen substances of H. physodes, the amount of the depsidone physodalic acid relative to the total of lichen substances was most closely correlated to the concentrations of Cu and Mn in the substratum, whereas the amount of the depsidone 3-hydroxyphysodic acid decreased both with increasing concentrations of these two metals and physodalic acid. Thus, our data suggest that lichen substances contribute to metal homeostasis not only in heavy metal-rich habitats, but also at not or low-polluted sites where the lichen substances apparently help to maintain constant intracellular metal concentrations despite of spatially varying availabilities of metal ions.

Effect of four lichen acids isolated from Hypogymnia physodes on viability of rat thymocytes

Four lichen acids, physodalic acid (F1), physodic acid (F2), 3-hydroxyphysodic acid (F3), and isophysodic acid (F4), were isolated from Hypogymnia physodes methanol extract using preparative reversed-phase high performance liquid chromatography and their structures were determined by UV, MS, 1H NMR and 13C NMR. This is the first report on the isolation of F4 from H. physodes. Isolated rat thymocytes were cultivated with increasing F1–F4 concentrations (0.1, 1, 10μg/well) and proliferative activity, viability, ROS (reactive oxygen species) production and MMP (mitochondrial membrane potential) disturbances were evaluated. Obtained results show significantly decreased thymocytes proliferation was observed when cells were treated with F1 (1μg, p<0.05; 10μg; p<0.001), F2 (10μg, p<0.05) and F3 compound (10μg, p<0.05). Significantly increased cytotoxicity was detected when cells were incubated with F1 (1μg, p<0.05; 10μg, p<0.01), F2 (10μg, p<0.05) and F3 compound (10μg, p<0.001). Increased H2DCF-DA fluorescence intensity, when cells were treated with F1 (1μg, p<0.001; 1μg, p<0.01; 10μg, p<0.001) and F2 (1μg, p<0.05; 10μg, p<0.01) compound, indicating the increase of intracellular ROS production. Simultaneously, increased ROS levels were followed with significantly decreased MMP when thymocytes were cultivated with F1 (0.1μg, p<0.001; 1μg, p<0.001; 10μg, p<0.001) and F2 compound (10μg, p<0.001). Thymocytes exposure to increased (0.1, 1, 10μg) concentrations of F3 and F4 compounds did not result with significant alterations in MMP and intracellular ROS production. We have shown that higher F1 and F2 concentrations induce thymocytes toxicity mainly through induction of oxidative stress, while cytotoxicity effect of F3 is followed with altered antioxidant/oxidant balance. The rigid 11H-dibenzo[b,e][1,4]dioxepin-11-one ring in the depsidone structure may play a important role for the examined biological activities.

Larvicidal activity of some secondary lichen metabolites against the mosquito Culiseta longiareolata Macquart (Diptera: Culicidae)

The larvicidal activity of some lichen metabolites, (+)-usnic acid, atranorin, 3-hydroxyphysodic acid and gyrophoric acid, against the second and third instar larvae of the mosquito Culiseta longiareolata were studied. All metabolites caused high larvicidal activities. When metabolites were compared on the basis of their LC50 values, the order of increasing toxicity was as follows: gyrophoric acid (0.41 ppm) > (+)-usnic acid (0.48 ppm) > atranorin (0.52 ppm) > 3-hydroxyphysodic acid (0.97 ppm). However, when LC90 values were compared, the order of toxicity was (+)-usnic acid (1.54 ppm) > gyrophoric acid (1.93 ppm) > 3-hydroxyphysodic acid (4.33 ppm) > atranorin (5.63 ppm). In conclusion, our results found that lichen secondary metabolites may have a promising role as potential larvicides.

Hypogymnia magnifica (Parmeliaceae), a new lichen from southwest China

Hypogymnia magnifica X.L.Wei & McCune is described as a new species of lichenized fungi from high elevations in Yunnan and Sichuan Provinces in southwestern China. Previously lumped under H. taiwanalpina, H. magnifica is readily distinguished from that species by its large size (commonly to 30 cm or more diam.), broad, rather appressed lobes that are contiguous to subcontiguous, sparse perforations in the upper and lateral surfaces, whitish color in the field (when dry), and lacking 3-hydroxyphysodic acid. Hypogymnia magnifica and H. taiwanalpina appear to be allopatric, with the former restricted to southwestern China and the latter restricted to far-east Asia (Taiwan and Japan).

Hypogymnia canadensis (Parmeliaceae), a new lichen from the Pacific Coast of North America

Hypogymnia canadensis Goward & McCune is described as a new species of lichenized fungi from the west coast of North America, ranging from southeastern Alaska to western Oregon, inland to southeastern British Columbia. In many respects similar to H. metaphysodes (Asahina) Rass., H. canadensis is distinguished from that species by an imbricate growth form, more open branching, lobe tips that readily become brown in exposed sites, a darker ceiling of the lobe cavity, and the presence of 3-hydroxyphysodic acid in the medulla (K slowly reddish brown). So far H. canadensis is known from conifers in cool suboceanic to oceanic climates.

Hypogymnia incurvoides(Parmeliaceae), a little-known Russian species discovered in eastern North America

Hypogymnia incurvoides Rass. was originally described from the Archangel'sk Region in northwestern Russia in 1967. Long neglected, it has since been found at several sites in Newfoundland and Nova Scotia in eastern Canada. Hypogymnia incurvoides is most similar to H. physodes, but is readily distinguished from that species by the presence of holes in the lobe tips and axils, black-margined lobes forming a regular lattice, and the absence of 3-hydroxyphysodic acid. This substance gives a K reddish brown reaction, which is consistently seen in H. physodes but not H. incurvoides. The species should be sought in other oceanic boreal to subarctic locations in the Northern Hemisphere.

The Antimicrobial Activity of Extracts of the Lichen Hypogymnia tubulosa and Its 3-Hydroxyphysodic Acid Constituent

The antimicrobial activity and the MIC values of the diethyl ether, acetone, chloroform, petroleum ether, and ethanol extracts of the lichen Hypogymnia tubulosa and its 3-hydroxyphysodic acid constituent have been investigated against some microorganisms. At least one of the extracts or 3-hydroxyphysodic acid showed antimicrobial activity against Aeromonas hydrophila, Bacillus cereus, Bacillus subtilis, Escherichia coli, Klebsiella pneumoniae, Listeria monocytogenes, Proteus vulgaris, Salmonella typhimurium, Staphylococcus aureus, Streptococcus faecalis, and Candida albicans. No antifungal activity of the extracts has been observed against ten filamentous fungi.

Intrathalline Variation in Concentration of Lichen Substances in Pseudevernia furfuracea

In Pseudevernia furfuracea olivetoric acid and atranorin decrease toward the base of the laciniae while physodic and chloratranorin increase. Oxyphysodic acid shows a contrasting pattern of intrathalline distribution: when olivetoric acid is absent (chemical race I) it decreases from the apex to the base of the laciniae, but when olivetoric acid is present (chemical races II and IV) the relative concentration of oxyphysodic acid increases in the same direction. Biogenetic considerations of these lichen products and some hypotheses of the origin of chemical variation in this species are discussed. For many years Pseudevernia furfuracea (L.) Zopf has been the subject of numerous chemical studies such as those developed by Culberson (1965) and Culberson et al. (1977). Current evidence suggests that in this species there are three races, having either physodic acid (race I), olivetoric acid (race III), or both (race II), as the major substances in the medulla. Moreover, some studies have attempted to correlate this chemical variation with ecological or geograph- ical factors (Hale 1968; Halvorsen & Bendiksen 1982; Hawksworth & Chapman 1971), but only a corre- lation with the distribution of the chemical races in an European northwestern-southeastern gradient has been observed. Oxyphysodic acid (3-hydroxyphysodic acid) is an- other substance found in P. furfuracea thalli and it is usually associated with physodic acid, although it is not always detected in chemical strain II, where olivetoric acid is present as well (C. Culberson, pers. comm. 1987). Culberson et al. (1977) did not detect this acid with physodic and olivetoric acids in spec- imens of the latter race. The low sensitivity of TLC may be responsible for this, but alternatively the con- centration of oxyphysodic acid could vary signifi- cantly along the laciniae, or even be absent in places, depending on the physiological activity of each part of the thallus. In a previous paper (L6pez & Manrique 1989) we described a new infrequent (1.3%) chemical race in Spain that contains olivetoric, physodic, and oxy- physodic acids together (race IV). For this reason we decided to investigate more thoroughly the chemistry of P. furfuracea. Intraspecific variation in chemistry as well as variations in substance concentration with-

ChemInform Abstract: 3‐HYDROXYPHYSODIC ACID. CHEMICAL CORROBORATION OF THE STRUCTURE OF THIS LICHEN DEPSIDONE

AbstractEs wird gezeigt, daß die früher veröffentlichte Struktur (Ia) für das Flechten‐Depsidon "Physodol" inkorrekt ist und daß diese Verbindung vielmehr mit der Titelverbindung (Ib) identisch ist, deren Struktur auf chemischem Weg bestätigt wird.

ChemInform Abstract: THE STRUCTURE OF LIVIDIC ACID. A DEPSIDONE FROM THE LICHEN PARMELIA FORMOSANA

AbstractDer Extrakt aus der Flechte Parmelia formosana liefert beim Behandeln mit Phenyldiazomethan ein Gemisch, aus dem chromatographisch der Benzylester (Ia) isoliert wird.

The structure of the lichen depsidone hydroxyphysodic acid. A carbon-13 magnetic resonance study

Unequivocal evidence for the structure of the lichen depsidone 3-hydroxyphysodic acid has been provided by analysis of long-range carbon-13-hydrogen coupling interactions and carbon-13 chemical shifts. This approach was also used to establish the isocoumarin structure (16) for that of the key decarboxycyclodehydration product obtained by degradation of 3-hydroxyphysodic acid. The breakdown of some carbon-13 additivity effects in the sterically congested ring is discussed. A possible mechanism for the formation of the isocoumarin (16) is outlined.

The structure of lividic acid. A depsidone from the lichen Parmelia formosana

The depsidone, lividic acid [1'-carboxy-2',3-dihydroxy-4.methoxy-6-(2"-oxoheptyl)-6-pentyldepsidone](1) has been isolated from the lichen Parmelia formosana Zahlbr. The structure of this compound followed from a combination of spectroscopic properties and chemical interconversions. The overall substitution pattern of this depsidone was confirmed by methylation with excess diazomethane, whereupon methyl 3-methoxy-2',4,4'-tri-O-methylisophysodate was obtained. Further, lividic acid was synthesized from 3-hydroxyphysodic acid by methylation of the corresponding benzyl ester and subsequent hydrogenolysis.

3-Hydroxyphysodic acid. Chemical corroboration of the structure of this lichen depsidone

The published structure of the lichen depsidone ?physodol? has been shown to be incorrect. This compound is in fact identical with 3- hydroxyphysodic acid (2) (1?-carboxy-2?,3,4-trihydroxy-6-(2?- oxoheptyl)-6?-pentyldepsidone). ��� Chemical evidence is presented to confirm the isocoumarin structure (10) for that of the key decarboxy anhydro derivative obtained by degradation of 3-hydroxyphysodic acid, and thus to disprove the alternative 5-hydroxy formulation suggested for this compound.