Next Article in Journal
Photophysical and Bactericidal Properties of Pyridinium and Imidazolium Porphyrins for Photodynamic Antimicrobial Chemotherapy
Next Article in Special Issue
Impact of In Vitro Gastrointestinal Digestion on the Bioaccessibility of Phytochemical Compounds from Eight Fruit Juices
Previous Article in Journal
Metal-based Heterogeneous Catalysts for One-Pot Synthesis of Secondary Anilines from Nitroarenes and Aldehydes
Previous Article in Special Issue
The Wonderful Activities of the Genus Mentha: Not Only Antioxidant Properties
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Molecules
Volume 26
Issue 4
10.3390/molecules26041121
molecules-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Phytochemical Investigation of New Algerian Lichen Species: Physcia Mediterranea Nimis

by
Marwa Kerboua
1,
Monia Ali Ahmed
1,
Nsevolo Samba
2,3,
Radhia Aitfella-Lahlou
2,4,5,
Lucia Silva
2,4,
Juan F. Boyero
6,
Cesar Raposo
6 and
Jesus Miguel Lopez Rodilla
2,4,*
1
Laboratory of Vegetal Biology and Environment, Biology Department, Badji Mokhtar University, Annaba 23000, Algeria
2
Chemistry Department, University of Beira Interior, 6201-001 Covilha, Portugal
3
Department of Clinical Analysis and Public Health, University Kimpa Vita, Uige 77, Angola
4
Fiber Materials and Environmental Technologies (FibEnTech), University of Beira Interior, 6201-001 Covilhã, Portugal
5
Laboratory of Valorisation and Conservation of Biological Resources, Biology Department, Faculty of Sciences, University M’Hamed Bougara, Boumerdes 35000, Algeria
6
Department of Analytical Chemistry, Nutrition and Food Science, Faculty of Chemistry, Chromatographic and mass analysis service (NUCLEUS), University of Salamanca, 37008 Salamanca, Spain
*
Author to whom correspondence should be addressed.
Molecules 2021, 26(4), 1121; https://doi.org/10.3390/molecules26041121
Submission received: 21 January 2021 / Revised: 17 February 2021 / Accepted: 18 February 2021 / Published: 20 February 2021
(This article belongs to the Special Issue Bioactive Compounds and Antioxidant Activity of Extracts from Different Natural Plants)
Browse Figures
Review Reports Versions Notes

Abstract

:
The present study provides new data concerning the chemical characterisation of Physcia mediterranea Nimis, a rare Mediterranean species belonging to the family Physciaceae. The phytochemical screening was carried out using GC-MS, HPLC-ESI-MS-MS, and NMR techniques. Hot extraction of n-hexane was carried out, followed by separation of the part insoluble in methanol: wax (WA-hex), from the part soluble in methanol (ME-hex). GC-MS analysis of the ME-hex part revealed the presence of methylbenzoic acids such as sparassol and atraric acid and a diterpene with a kaurene skeleton which has never been detected before in lichen species. Out of all the compounds identified by HPLC-ESI-MS-MS, sixteen compounds are common between WA-hex and ME-hex. Most are aliphatic fatty acids, phenolic compounds and depsides. The wax part is characterised by the presence of atranorin, a depside of high biological value. Proton 1H and carbon 13C NMR have confirmed its identification. Atranol, chloroatranol (depsides compound), Ffukinanolide (sesquiterpene lactones), leprolomin (diphenyl ether), muronic acid (triterpenes), and ursolic acid (triterpenes) have also been identified in ME-hex. The results suggested that Physcia mediterranea Nimis is a valuable source of bioactive compounds that could be useful for several applications as functional foods, cosmetics, and pharmaceuticals.

1. Introduction

From the Greek word, “leikhen” lichen was first used to designate a plant in the 4th century BC by Theophraste [1]. This small organism has been integrated into the fungal kingdom and results from the symbiotic association of a fungus called mycobiont and a green alga or a cyanobacterium called photobiont [2,3]. An estimated more than 17,000 species of lichen exist today, extending from the tropics to the polar regions [4]. The symbiosis gives lichens a specific structure and reproduction to each constituent alone. Unlike higher plants, they have no root, stem, or leaf, but a rudimentary vegetative apparatus: the thallus [5]. Moreover, they grow on a wide variety of substrates including soil, bark, bare rock surfaces, leaves of vascular plants, barnacle shells, and other lichens [4].
The mycobiont plays the most determining role in the morphology and structure of the lichen [6]. The thallus carries the elements necessary for reproduction. A great diversity of shapes and colours defines seven main types of lichens [7]. These organisms tolerate large temperature variations, extreme desiccation, and intense exposure to ultraviolet [8]. In addition, this tolerance to light and desiccation is better in lichen compared to isolated partners [9]. As a two or even multiple partnerships, lichens can respond more sensitive to different environmental signals in complex ecosystems than algae and fungi growing unprotected and living aposymbiotically according to Reyes et al., 1996 [10].
Lichens produce many unusual secondary metabolites that have not been discovered in other plants [11]. The uniqueness of many lichen substances attracted the first chemists’ attention in the mid-19th century. Before this, they were used as dyes for textiles and as additives for soap and perfume making, and a considerable number for healing diseases [12,13]. Lichens produce more than 800 potentially bioactive compounds [14,15,16]. Among these compounds are nitrogen, phosphorus and sulfur, polyols, carbohydrates, aliphatic and cycloaliphatic compounds, aromatic compounds, meta- and para-depsides, and depsidones, dibenzofurans, diphenylethers, naphtopyrans, biphenyls, diphenylmethanes, nostoclides, xanthones, quinones, naphthoquinones, and usnic acid. Esters, terpenes, steroids, terphenylquinones, and pulvinic acid derivatives also occur [16,17,18]. The majority of lichenic substances are small aromatic polyketides. They are synthesised during the mutualistic relationship (symbiosis) of lichens with green photobionts [13,19]. These substances are often absent in cyanobacterial lichens replaced by other interesting secondary metabolites, e.g., terpenes and terpenoids [20,21].
Recently, lichens have attracted several researchers worldwide to review their therapeutic and cosmetic value in traditional medicine. A wide range of species have revealed effective biological activities such as antioxidant, cardiovascular protectors, antimicrobial, anti-inflammatory, analgesic, antipyretic, antiviral, anti-insecticidal, antibiotic, antifungal, antidiabetic, and anticancer [22,23,24,25,26,27,28,29,30,31]. Various applications of lichen extracts as a treatment for skin conditions, wounds, respiratory and digestive problems, as well as for obstetric and gynaecological problems have also been recorded [32]. The pharmacological potential of lichens is due to the presence of unique secondary metabolites which subsequently became excessively attractive to the pharmaceutical and cosmetic industries. As a result, a multitude of optimised processes for the quantitative extraction of these lichen metabolites has been developed [33].
In Algeria, several investigations have been carried out to explore their diversity [34,35,36,37,38,39,40,41,42,43,44,45]. More than 1085 species of lichens have been identified, 64 of which are endemic [41]. Most research has been focused on their use as bioindicators of air pollution [42,43,46,47,48,49]. However, many regions remain unexplored, and research reports on lichen chemistry in this richly diverse country are very limited.
In the present investigation, we analysed the phytochemical profile of Physcia mediterranea Nimis (Figure 1), belonging to the family of Physicaceae. The lichen, a new species recently identified in Algeria, was collected in the El-Kala National Park, a remarkable and culturally rich region with more than 117 species of lichens identified. Several species of this area, both plant and animal, are protected in Algeria and belong to the IUCN (International Union for Conservation of Nature) red list. Indeed, the National park of El-Kala was classified as “Reserve of the biosphere”, by UNESCO, on 17 December 1990 [50]. It can be concluded that the Algerian species of lichens that have been little or not studied deserve special attention with research involving both the chemical part and the biological one. This would allow us to increase the contribution to discovering new compounds that may serve as models for new drugs with therapeutic properties. The literature search reveals that still no studies have been done on Physcia mediterranea Nimis species’ products. Therefore, our work aims to isolate and investigate the chemical constituents in different organic lichen extracts from Physcia mediterranea Nimis using GC-MS, followed by HPLC-ESI-MS-MS and NMR analysis.

2. Results and Discussion

2.1. GC-MS Analysis

The GC-MS analysis of ME-hex extract is shown in Figure 2. The analysis allowed us to detect ten components. Nine products were identified by comparing their mass spectra with reference spectra from the NIST and Wiley databases (Table 1). The major chemical compounds identified in ME-hex from Physcia mediterranea included derivatives of methylbenzoic acids (56%) and free fatty acids (36.1%). A diterpene with a Kauran skeleton ((−)-ent-Kauran-16a-ol) (Figure 3) was also identified (3.8%) [18]. Only 2.2% of the entire extract was not identified.
Derivatives of methylbenzoic acids are commonly present in different genera of lichen-like Stereocaulon halei [51], Parmotrema mesotropum [52], Cassipourea malosana [53], Cetraria islandica [54], Usnea longissima [55,56], Stereocaulon paschale [57], and Parmelia sulcata Taylor [58]. Atraric acid (Figure 2), widely present in some species such as Hypogymnia physodes, Evernia prunastri, and Parmelia sulcata, growing on the same host tree (Prunus domestica) [59], can be in the free or complexed form and serves as a basis for the composition of depsides and depsidones [60]. It is considered a specific antagonist of androgen receptors and therefore inhibits human prostate cancer growth [60,61]. Atraric acid shows nematocidal, antioxidant, antimicrobial, and anti-inflammatory properties in vitro, and inhibits carrageenan-induced oedema and wound healing activity in vivo [62,63].
Lichens contain many of the fatty acids commonly found in higher plants [64]. Indeed, many common lichen genera include species with multiple strains of fatty acids [65,66,67]. These lipid profiles, most often treated as chemotypes, have been used by many researchers to taxonomically classify certain lichens, such as Cladonia [68], Lepraria [69], Parmelia [70], Tephromela s.lat [71], and Mycoblastus sanguinarius [72]. Although they are widely present in some genera, fatty acids were of no taxonomic importance and were omitted during the first chemical studies on lichens [72,73,74]. By chemical and biochemical comparisons, a mechanistic relationship between polyketide and fatty acid biosynthesis has been recognised. The carbon backbones of the molecules are assembled by successive condensation of acyl units [75]. Another important fatty acid role was cell signal transduction [76] as well as chemical protection. Therefore, they allow the lichens to survive as environmental conditions change [8,9,10].
In our study, palmitic acid is present in high concentration (24%) compared to oleic (4.2%), stearic acid (3.8%), and linoleic (3.2%) acid (Table 1 and Figure 2). Various works using different growing conditions explain the variations in fatty acid in lichens [77,78]. Molina et al. (2003) studied the lichen Physconia distorta and suggested a close relationship between the synthesis of secondary metabolites and fatty acid metabolism. Mycobiota grown in a glucose-enriched medium favoured the production of fatty acids [79]. Another important factor that could influence the production of fatty acids in lichens is temperature. According to several studies, the degree of unsaturation varies with the season and decreases with increasing temperature [80,81]. In the thallus of Teloschistes flavicans, the saturated fatty acids, palmitic and stearic, were more abundant in February. In contrast, in August, when the average temperature was 23 °C, there was an increase in oleic and linoleic fatty acids [82].
In addition to the stress due to the decrease in temperature, nitrogen deprivation and light intensity are also known to promote fatty acid accumulation [80,82,83]. However, these factors are not necessarily the unique parameters that determine the fatty acid content, but rather genetics, combined with environmental conditions (e.g., altitude, air pollution, seasonal effects), must also be taken into account [67,84].

2.2. HPLC-ESI-MS-MS Analysis

2.2.1. WA-hex Extract Analysis of Physcia mediterranea

Forty-five peaks (Figure S1) were detected for the first time in a WA-hex (Table 2) using LC/ESI/MS/MS in negative mode. Identifying major compounds was simplified by interpreting their MS/MS spectra, provided in our system resource and comparison with the literature. The 37 compounds identified were mainly paraconic and aliphatic acids, depsides (aromatic polyketides), phenolic compounds, and diterpenes. Only eight compounds could not be identified. The representative chemical structures are presented in Figure S2.
Twenty-nine paraconic and aliphatic acids were identified: peaks 1–2, 5–7, 9, 13–19, 21–24, 26–29, 31, 35, 37, 39, 40, 42, and 44 using UHPLC-ESI-MS-MS analysis [13,18]. Among these compounds is fumaric acid (Figure S2), a valuable compound used in food, beverages, detergents, animal feed, pharmaceuticals, and various industrial products [85,86,87,88]. Similarly, for traumatic acid, a phytohormone belongs to the class of cytokinins, a study has demonstrated its positive influence on oxidative stress parameters in normal human fibroblasts [89]. It is also effective against breast cancer cells and has potential anticancer properties and tumour prevention activity. Traumatic acid leads to decreased cell proliferation and viability, GSH/GSSG ratio, and thiol group content. It increased caspase activity, membrane lipid peroxidation, and ROS content, simultaneously reducing breast cancer cell growth through the influence of oxidative stress on apoptosis [90].
Three phenolic compounds were identified and assigned to peaks 4, 8, and 10 using UHPLC-ESI-MS-MS. Peak 4 and peak 10 were identified as p-Coumaric acid 6,7-Dihydroxycoumarin (Esculetin) (Figure S2) whose molecular anions were at m/z 163.0395 and 177.0186, respectively. To our knowledge, this is the first time that compounds such as these have been found in lichen material. They possess diverse biological and pharmacological properties, including anti-asthma, anti-inflammatory, anti-nociceptive, antioxidative, antitumor, and antiviral activities [91,92,93,94,95,96,97]. Gingerol, a phenolic compound (Figure S2), has been identified in the extract WA-hex. Its peak 8 would correspond to [M − H]− ion at 293.1762 (Table 2).
Peak 11 and 12 were identified as depsides: atranorin ([M − H]− ion at m/z 373.0929) and chloroatranorin ([M − H]− ion at m/z 407.0539), respectively (Table 2, Figure S2). Atranorin, a derivative of β-orcinol, is one of the most common secondary metabolites of lichen [98]. It is particularly present in the lichen families Cladoniaceae, Lecanoraceae, Parmeliaceae, and Streocaulaceae [99,100,101]. In the Physcia genera, there are Physcia caesia [102], Physcia aipolia [103,104], Physcia alnophila [105], and Physcia sorediosa [106]. In recent years, atranorin has been the most extensively studied. Indeed, it has shown antioxidant [107], antimicrobial [108], anti-inflammatory [109], antinociceptive [110], wound healing [111], and photoprotective properties [112]. Additionally, it exerted strong inhibitory effects on cancer cell proliferation, migration, and actin cytoskeleton organisation [113].
Portentol, peak 32 ([M − H]− ion at m/z 309.1743) and Stephanol peak 36 ([M − H]− ion at m/z 397.2266), which are cycloaliphatic compounds were also identified in WA-hex (Table 2, Figure S2). Likewise for asebotoxin I, a toxic diterpene has been identified at the peak 43 ([M − H]− ion at m/z 425.2581). It was initially and only discovered in the plant Pieris japonica [114], but our results show the opposite, i.e., that the compound can be detected in lichen species.

2.2.2. ME-hex Extract Analysis of Physcia mediterranea

In the present study, the analysis of the phytochemical profile of ME-hex using UHPLC-ESI-MS/MS, in negative ion mode, resulted in the detection of 54 significant compounds indicated in Figure S3. Only four compounds could not be identified. The identified compounds are of paraconic and aliphatic acids, aromatic polyketides (depsides, depsones, and phenyl ethers), phenolic acids, sesquiterpenes lactones, triterpenes, carboxybenzaldehyde, and carboxyphthalide types (Table 3).
Thirty-seven paraconic and aliphatic acids corresponding to peaks 1–3, 5, 6, 8–10, 12, 14–19, 22, 24–30, 38, 39, and 41–52 were identified using UHPLC/ESI/MS/MS analysis. 3,5-Dimethoxyciclohexanecarboxylic acid (Figure 4) was identified as peak 1 (molecular anion at m/z 188.1043). The fragmentation of peak 1 produced ion at m/z 143.8651 [M − H − CO2]−, and 141.8670 [M − H − CO2]−. Peak 2, with an [M − H]− ion at m/z 294.0741, was identified as 6-(hydroxymethyl)-3,5-bis(methoxycarbonyl)-2,4-dimethylcyclohex-1-ene-1-carboxylic acid (Figure 4). The fragmentation of peak 1 produced ion at m/z 249.0771 [M − H − CO2]−, 234.0530 [M – H − CO2 − CH3]−, and 207.0695 [M − H − CO2 − CH3]−, confirming this compound. Peak 3 was identified as 3,5,6-hydroxymethyl-2,4-dimethylcyclohex-1-ene-1-carboxylic acid (Figure 4), which showed a [M − H]− at m/z 244.1311. Major diagnostic daughter MS ions of the compound were [M − H − CO−CH3OH]− and [M − H − H2O2]− (183.1025 and 176.6474 a.m.u, respectively). Peak 5 with a [M − H]− ion at m/z 226.1201 was identified as 5-formyl-3-hydroxymethyl-2,4,6-trimethylcyclohex-1-ene-1-carboxylic acid (Figure 4). The fragmentation of the peak produced ions at m/z 207.1023 and 97.0284 confirming this structure. Peak 6 was identified as 3,5-dihydroxy-2,4,6-trimethylciclohexenecarboxilic acid (molecular anion at m/z 200.1046) (Figure 4). Major daughter ions of peak 6 were at m/z 183.4492, 162.8385, and 114.9508. Peak 8 with a [M − H −]− ion at m/z 242.1153 was identified as 4-O-demethylbaeomycesic acid, and their major diagnostic daughter MS ions were [M − H − CO2]− and [M − H − 2CH3OH]−, (218.8167 and 172.6485 a.m.u, respectively). The 2,4-dihydroxy-3,5,6-trimethylcyclohexane-1-carboxylic acid (Figure 4) was identified as Peak 8 ([M − H]− at m/z 202.1202). The fragmentation of this compound produced three ions at m/z 197.6263, 164.8350 and 139.1124 confirming its structure. Peak 12 was identified as 4-hydroxy-2,5-dimethylcyclohex-1-ene-1-carboxylic acid (Figure 4), which showed a [M− H ]− peak at m/z 170.0936. Major diagnostic daughter MS ions were [M − H − CO]−, [M − H − CO2]−, and [M − H − CO2]− (193.0514, 140.9478, 124.9796, and 104.4180 a.m.u., respectively). Peak 14 was assigned to 6-(1-Oxopentyl)-1-cyclohexene-1-carboxylic acid (Figure 4) on its resolution molecular anion at m/z 210.1253 ([M − H]− peak). It produced major diagnostic MS ions at 164.8947, 146.9850, and 105.0336.
According to the results (Table 3), four depsides compounds were identified corresponding to peak 4, 11, 23, and 35 in the ME-hex extract. Peak 4 was identified as atranol (Figure 5) (molecular anion at m/z 152.0465). The fragmentation of peak 4 also produced ions at 123.0444 [M − H − CO2]−, 105.0335 [M − H − CO2− H2O2]−, and 81.0335 [M − H − CO2 − H2O2]− confirming this compound. Chloroatranol and 8-Hydroxydiffractaic acid identified at peak 11 and 23, showed [M − H]− ions at m/z 186.0079 and 390.1315, respectively. This is also chloratranorin (Figure 5) at peak 35, which shows [M − H]− ions at m/z 408.0611.
A despsone named allo-protolichesterinic acid was also detected in ME-hex extract. It would correspond to the peak 21 and shows [M − H]− ions at m/z 324.2303. Moreover, two phenolic acids were identified, corresponding to peak 13 and 31: p-Coumaric acid and dihydroxycoumarin (Figure 5). They showed [M − H]− ions at m/z 164.0470 and 178.0259 respectively. Peak 34 was identified as 7-chloro-3-oxo-1,3-dihydroisobenzofuran-5-carboxylic acid (carboxyphthalide) (Figure 5), which showed a [M − H]− peak at m/z 211.9873. It produced major diagnostic MS ions at m/z 138.9948 and 103.0180, confirming this compound.
It is similar for the compounds fukinanolide (sesquiterpene lactones) at peak 20 and leprolomin (diphenyl ether) (Figure 5) at peak 37. They showed a [M − H]− peak at m/z 312.2302 and 366.2403 respectively. In the study by Zhang et al. (2016), fukinanolide, also called bakkenolide A, extracted from the plant Petasites tricholobus, showed anti-inflammatory properties in the treatment of leukaemia [115]. In the lichens group, the sesquiterpene has only been identified in Cetraria islandica [116].
Finally, two triterpenes, muronic acid at peak 32 and ursolic acid at peak 39, were also identified (Figure 5) and produced major diagnostic MS ions at m/z 390.1314, 456.3605. Muronic acid was previously identified in Usnea rubicunda, Usnea subfloridana [117], and Punctelia microsticta [118]. On the other hand, ursolic acid (3β-hydroxy-urs-12-ene-28-oic acid) is widespread in the vegetable kingdom [119]. The lichenic species Ramalina hierrensis [16], Ramalina hierrensis [120], Stereocaulon evolutum [121], and Pannaria tavaresii [122] also contain this compound.

2.3. NMR Analysis

1H and C13 NMR analysed the two samples from WA-hex and ME-hex. The results we obtained reveal the predominance of the secondary metabolite atranorin only in the WA-hex sample. The structure has been characterised, and the NMR spectra are compared with those of a previously isolated sample of Physcia sorediosa [106,123], demonstrated in Figures S4 and S5.
Atranorin (Figure 6) is the most common secondary metabolite in lichens and is mainly found on lichens’ surface (cortex) [124]. It acts as a photo-buffer because it reflects harmful UV rays to the thallus’s surface and allows the lichens to live in areas receiving intense solar radiation [125].
Several factors can influence atranorin concentrations in lichens. They fluctuate with the seasons [126,127] and vary according to the habitat [128]. The method of preparation and extraction of lichens can also influence the concentration of this metabolite. Conventional organic solvents (such as hexane and acetone) are commonly used for its extraction as hydrophobic [129]. Our study used n-hexane with Soxhlet extraction method, which is of choice in studying organic analytes extracted from lichens [130]. It is still used to date to extract organic air pollutants, organochlorinated pesticides and insecticides from the lichen matrix [131,132].
According to Komaty et al. (2015), the lichen grinding method can affect the extraction efficiency and even be used to selectively increase the extraction efficiency of certain secondary metabolites such as atranorin [133]. To achieve a higher yield, use a blender instead of a ball mill, as it selectively grinds the cortex into a fine powder, which can be recovered from the larger medulla pieces. A ball mill or a mortar and pestle technique will reduce the whole lichen to powder, which will reduce the extraction efficiency of the atranorin [129]. Pseudevernia furfuracea is a lichen widely used as a raw material in the perfume and cosmetics industries due to its richness in aromatic compounds [33,134,135,136,137]. Microwave-assisted extraction of this lichen has increased atranorin extraction efficiency by a factor of five [133].

3. Materials and Methods

3.1. Lichen Material

The saxicolous lichen specimen Physcia mediterranea Nimis was collected at Ain Tebib (Sector Oum tboul) on the rock, at an altitude of 120 m above sea level, coordinate 36°49′ 09″ N; 08°31′ 33″ E in June 2017 (Figure 7). Ain Tebib station is located in the national park of El Kala (80,000 ha). The collected lichen samples were packed in polyethene bags and stored at 4 °C until further processed. Professor Monia Ali Ahmed has identified Physcia mediterranea Nimis (Figure 1), lichenologist and research director of the Pathology of Ecosystems team at the University of Badji-Mokhtar, Annaba, Algeria. Botanical description of Physcia mediterranea Nimis is in Figure S6. The identification was confirmed by Pr Jean Michel Sussey, lichenologist at the French Association of Lichenology (AFL). This sample has been deposited in Badji-Mokhtar University, Annaba, code AAM-1.

3.2. Sample Preparation

The lichens were washed with tap water to remove the dust and other foreign materials. The washed samples were dried under shade for a week. The lichen was ground using a grinder. The preparations were then pulverised into powdered form by using heavy-duty blender.

3.3. Preparation of Physcia mediterranea Extracts

The powder samples (24 g) of Physcia mediterranea Nimis were extracted with the solvent n-hexane (500 mL) using a Soxhlet extractor 24 h. After complete extraction, the solvent was evaporated using a rotary evaporator under reduced pressure to obtain n-hexane extract (1.026 g). It was then extracted with hot methanol (60 °C) to obtain two parts: insoluble precipitate representing lichen wax (WA-hex) and the methanol soluble part (ME-hex). Both extracts (WA-hex/ME-hex) were completely evaporated using a rotary evaporator under reduced pressure to obtain dry extracts (0.552 g/0.300 g), respectively.

3.4. Instrumentation and Analysis Parameters

In this present study, the chemical composition of the two fractions WA-hex and ME-hex of Algerian Physcia mediterranea Nimis was analysed using the HPLC-ESI-MS-MS method. Moreover, we combined GC-MS’s ability with the targeted metabolomics of HPLC-ESI-MS-MS methods to characterise the composition of n-hexane extract (ME-hex) for the first time. In addition, most compounds potentially present in the wax (WA-hex) are detected and characterised using the NMR method.

3.4.1. GC-MS Analysis

For the GC-MS analysis, an Agilent MS220 (Varian, Inc. Walnut Creek, CA, USA) mass spectrometer coupled to a 7890A GC. The oven temperature was initially set to 50 °C, held for 5 min, then a ramp of 30 °C/min was applied up to 270 °C that was held for an additional5 min. MS spectra were acquired in EI mode with a mass range from 50 to 600 a.m.u. Before being injected into the GC-MS system, the ME-hex fraction was pre-esterified with diazomethane, in order to identify eventual less polar compounds in this fraction. It was then solubilised in dichloromethane and injected into the apparatus.

3.4.2. NMR Analysis

Proton 1H and carbon 13C NMR spectroscopy were recorded on a Brüker Advance III 400 MHz spectrometer (Brüker Scientific Inc, Billerica, MA, USA) at 400 MHz for proton and 100 MHz for carbon. The recovered WA was dissolved in deuterated solvent’s (CDCl3) (5 mg/mL), at room temperature. The solution was transferred to 5 mm outside diameter tubes, and the spectra were acquired at room temperature. The deuterated solvent’s residual peak signal was for 1H spectra at 7.26 ppm and 13C spectra at 77.2 ppm. The chemical deviations (δ) are expressed in parts per million (ppm) and the coupling constants (J) in Hertz. The data was processed using TOPSPIN 3.5 software (Brüker Scientific Inc.).

3.4.3. HPLC-ESI-MS-MS

The method was carried out on an orbitrap Thermo q-Exactive mass spectrometer coupled to a Vanquish HPLC. A Kinetex XB-C18 (Phenomenex) with a particle size of 2.6 microns, 100 mm of length, and a diameter of 2.1 mm was used as a column. The mobile phases were 0.1% formic aqueous solution (A) and, acetonitrile (B). The gradient program (time (min), % B) was: (0.00, 50); (20.00, 100); (25.00, 100); (26.00, 50). The flow rate was 0.200 mL min−1 and the injection volume was 10 µL.
The ionisation electrospray in negative mode was used. The following analysis parameters were: electrospray voltage −3.8 kV, sheath gas flow rate, 30; auxiliary gas unit flow rate, 10; drying gas temperature, 310 °C; capillary temperature, 320 °C; S-lens and RF level, 55. The acquisition was performed in a mass range from 100 to 1000 a.m.u. An auto MS2 program was used with a fragmentation voltage of 30.

4. Conclusions

Knowledge of the chemical constituents of lichens is invaluable as this information will be useful for the synthesis of potential new chemical substances. Many researchers report such phytochemical screening of various lichens [103,106,116,117,123,124]. A growing body of evidence indicates that lichens’ secondary metabolites play an essential role in human health and may be nutritionally important [24,26,27,29,30,31,58,112,118,119,120]. In the present study, we have identified and chemically characterised Algerian Physcia mediterranea Nimis for the first time. The extraction of these metabolites was carried out with hot hexane. A methanol-crystallisation process allowed us to characterise and identify the atranorine depside, as a major component of lichen wax. In this work, several aromatic acids, a kaurane, and fatty acids have been identified by GC-MS of ME-hex. The UHPLC-ESI-MS-MS technique has been used to analyse the crystallised fraction, WA-hex, whose major product is atranorin and chloroatranorin with the minority products identified. In ME-hex fraction, this technique identifies paraconic and aliphatic acids, depsides, depsones, phenolic compounds, sesquiterpenes, triterpenes, and phenyl ethers in addition to other minority derivatives. This study reveals for the first time the different compounds of Physcia medeterranea considered as a rare international species; furthermore, it highlights the importance of the lichens of Algeria as a promising source of bioactive molecules.

Supplementary Materials

The following are available online, Figure S1 (UHPLC/ESI/MS/MS chromatogram of Physcia mediterranea (WA-hex). Figure S2: Chemical structure of Compounds identified in WA-hex of Physcia mediterranea by UHPLC/ESI/MS/MS. Figure S3: UHPLC/ESI/MS/MS chromatogram of Physcia mediterranea (ME-hex). Figure S4: 1H NMR spectrum of WA-hex, 400 MHz. Figure S5: C13 NMR spectrum of WA-hex, 100 MHz. Figure S6: Botanical description of Physcia mediterranea Nimis collected in El Kala region (Algeria).

Author Contributions

Conceptualisation, M.K. and M.A.A.; methodology of chemical analysis, J.M.L.R., N.S. and M.K.; validation, J.M.L.R., N.S. and M.A.A.; investigation, M.K. and N.S.; writing—original draft preparation, R.A.-L., M.K. and M.A.A.; references, R.A.-L., M.K. and M.A.A.; writing—review and editing, R.A.-L.; supervision, J.M.L.R. and M.A.A.; project administration, L.S., J.F.B., C.R., J.M.L.R., and M.A.A.; funding acquisition, J.M.L.R. All authors have read and agreed to the published version of the manuscript.

Funding

The authors thank FibEnTech-UBI, which is financed by National Funds from Fundação para a Ciência e a Tecnologia (FCT) and Community Funds (UID/Multi/00195).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Lichen of Physcia mediterranea Nimis was used in this study. Professor Monia Ali Ahmed identified Physcia mediterranea Nimis, lichenologist and research director of the Pathology of Ecosystems team at the University of Badji-Mokhtar, Annaba, Algeria. The identification was confirmed by Pr Jean Michel Sussey, lichenologist at the French Association of Lichenology (AFL). This sample has been deposited in Badji-Mokhtar University, Annaba, code AAM-1.

Acknowledgments

We thank the Ministry of Higher Education and Scientific Research (MESRS) of Algeria for the financial support to this study, and we would like to warmly thank Grira Abdeslem, Principal Forestry Inspector, El Kala National Park, for his presence and his invaluable assistance in the field.

Conflicts of Interest

The authors declare no conflict of interest.

Sample Availability

Samples of all compounds used in the study are available from the authors.

References

  1. Boullard, B. Guerre et Paix Dans le Règne Végétal; Ellipses: Paris, France, 1990. [Google Scholar]
  2. Armaleo, D.; Müller, O.; Lutzoni, F.; Andrésson, Ó.S.; Blanc, G.; Bode, H.B.; Collart, F.R.; Dal Grande, F.; Dietrich, F.; Grigoriev, I.V.; et al. The lichen symbiosis re-viewed through the genomes of Cladonia grayi and its algal partner Asterochloris glomerata. BMC Genom. 2019, 20, 1–33. [Google Scholar] [CrossRef] [Green Version]
  3. Weber, B.; Büdel, B. Fungi and lichens. In Encyclopedia of Earth Sciences Series; Springer: Amsterdam, The Netherlands, 2011; pp. 401–409. [Google Scholar]
  4. Taylor, T.N.; Krings, M. Fossil Plants: Fungi and Lichens. In Encyclopedia of Geology; Elsevier: Amsterdam, The Netherlands, 2004; pp. 436–443. ISBN 9780123693969. [Google Scholar]
  5. Adams, D.G.; Bergman, B.; Nierzwicki-Bauer, S.A.; Duggan, P.S.; Rai, A.N.; Schüßler, A. Cyanobacterial-plant symbioses. In The Prokaryotes: Prokaryotic Biology and Symbiotic Associations; Springer: Berlin/Heidelberg, Germany, 2013; pp. 359–400. ISBN 9783642301940. [Google Scholar]
  6. Werth, S.; Sork, V.L. Identity and genetic structure of the photobiont of the epiphytic lichen Ramalina menziesii on three oak species in southern California. Am. J. Bot. 2010, 97, 821–830. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  7. Van-Haluwyn, C.; Asta, J.; Gaveriaux, J.P. Guide des Lichens de France: Lichens Des Arbres; Belin, Ed.; Belin: Paris, France, 2009; ISBN 978-2701147000. [Google Scholar]
  8. Meeßen, J.; Sánchez, F.J.; Brandt, A.; Balzer, E.M.; de la Torre, R.; Sancho, L.G.; de Vera, J.P.; Ott, S. Extremotolerance and Resistance of Lichens: Comparative Studies on Five Species Used in Astrobiological Research I. Morphological and Anatomical Characteristics. Orig. Life Evol. Biosph. 2013, 43, 283–303. [Google Scholar] [CrossRef] [PubMed]
  9. Kranner, I.; Cram, W.J.; Zorn, M.; Wornik, S.; Yoshimura, I.; Stabentheiner, E.; Pfeifhofer, H.W. Antioxidants and photoprotection in a lichen as compared with its isolated symbiotic partners. Proc. Natl. Acad. Sci. USA 2005, 102, 3141–3146. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  10. Reyes, A.; Lopez-Bilbao, M.G.; Del, M.; Molina, C. Relationship between Isozymic Variability and Envir-onmental Conditions in the Lichen Xanthoria parietina. Phyton 1996, 36, 265–275. [Google Scholar]
  11. Goga, M.; Elečko, J.; Marcinčinová, M.; Ručová, D.; Bačkorová, M.; Bačkor, M. Lichen Metabolites: An Overview of Some Secondary Metabolites and Their Biological Potential. In Co-Evolution of Secondary Metabolites; Springer: Cham, Germany, 2018; pp. 1–36. [Google Scholar]
  12. Kaushik, A.; Giri, P.P. Lichens. In Lichen-Derived Products; Wiley: Hoboken, NJ, USA, 2020; pp. 179–187. [Google Scholar]
  13. Upreti, D.K.; Divakar, P.K.; Shukla, V.; Bajpai, R. Recent Advances in Lichenology: Modern Methods and Approaches in Lichen Systematics and Culture Techniques; Springer: New Delhi, India, 2015; Volume 2, ISBN 9788132222354. [Google Scholar]
  14. Yamamoto, Y.; Kinoshita, Y.; Kinoshita, K.; Koyama, K.; Takahashi, K. A zearalenone derivative from the liquid culture of the lichen, Baeomyces placophyllus. J. Hattori Bot. Lab. 2002, 92, 285–289. [Google Scholar]
  15. Chicita, F.C. Chemical and Botanical Guide to Lichen Products; UNC Press: Chapel Hill, NC, USA, 1969; ISBN 978-0-8078-6859-1. [Google Scholar]
  16. Moreira, A.S.N.; Braz-Filho, R.; Mussi-Dias, V.; Vieira, I.J.C. Chemistry and biological activity of Ramalina lichenised fungi. Molecules 2015, 20, 8952–8987. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  17. Oksanen, I. Ecological and biotechnological aspects of lichens. Appl. Microbiol. Biotechnol. 2006, 73, 723–734. [Google Scholar] [CrossRef]
  18. Huneck, S.; Yoshimura, I.; Huneck, S.; Yoshimura, I. Identification of Lichen Substances. In Identification of Lichen Substances; Springer: Berlin/Heidelberg, Germany, 1996; pp. 11–123. [Google Scholar]
  19. Calcott, M.J.; Ackerley, D.F.; Knight, A.; Keyzers, R.A.; Owen, J.G. Secondary metabolism in the lichen symbiosis. Chem. Soc. Rev. 2018, 47, 1730. [Google Scholar] [CrossRef]
  20. Elshobary, M.E.; Osman, M.E.; Abo-Shady, A.M.; Komatsu, E.; Perreault, H.; Sorensen, J.; Piercey-Normore, M.D. Algal carbohydrates affect polyketide synthesis of the lichen-forming fungus Cladonia rangiferina. Mycologia 2016, 108, 646–656. [Google Scholar] [CrossRef] [Green Version]
  21. Elkhateeb, W.; Daba, G. Occurrence of terpenes, polyketides, and tannins in some Japanese lichens and green mosses. Egypt. Pharm. J. 2020, 19, 216. [Google Scholar] [CrossRef]
  22. Solárová, Z.; Liskova, A.; Samec, M.; Kubatka, P.; Büsselberg, D.; Solár, P. Anticancer potential of lichens’ secondary metabolites. Biomolecules 2020, 10, 87. [Google Scholar] [CrossRef] [Green Version]
  23. Jha, B.N.; Shrestha, M.; Pandey, D.P.; Bhattarai, T.; Bhattarai, H.D.; Paudel, B. Investigation of antioxidant, antimicrobial and toxicity activities of lichens from high altitude regions of Nepal. BMC Complement. Altern. Med. 2017, 17, 1–8. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  24. Thadhani, V.M.; Karunaratne, V. Potential of Lichen Compounds as Antidiabetic Agents with Antioxidative Properties: A Review. Oxid. Med. Cell. Longev. 2017, 2017. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  25. Rodriguez, C.M.; Bennett, J.P.; Johnson, C.J. Lichens: Unexpected anti-prion agents? Prion 2012, 6, 11–16. [Google Scholar] [CrossRef] [Green Version]
  26. Millot, M.; Girardot, M.; Dutreix, L.; Mambu, L.; Imbert, C. Antifungal and anti-biofilm activities of acetone lichen extracts against Candida albicans. Molecules 2017, 22, 651. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  27. Zambare, V.P.; Christopher, L.P. Biopharmaceutical potential of lichens. Pharm. Biol. 2012, 50, 778–798. [Google Scholar] [CrossRef] [PubMed]
  28. White, P.A.S.; Oliveira, R.C.M.; Oliveira, A.P.; Serafini, M.R.; Araújo, A.A.S.; Gelain, D.P.; Moreira, J.C.F.; Almeida, J.R.G.S.; Quintans, J.S.S.; Quintans-Junior, L.J.; et al. Antioxidant activity and mechanisms of action of natural compounds isolated from lichens: A systematic review. Molecules 2014, 19, 14496–14527. [Google Scholar] [CrossRef]
  29. Shukla, V.; Joshi, G.P.; Rawat, M.S.M. Lichens as a potential natural source of bioactive compounds: A review. Phytochem. Rev. 2010, 9, 303–314. [Google Scholar] [CrossRef]
  30. Behera, B.C.; Mahadik, N.; Morey, M. Antioxidative and cardiovascular-protective activities of metabolite usnic acid and psoromic acid produced by lichen species Usnea complanata under submerged fermentation. Pharm. Biol. 2012, 50, 968–979. [Google Scholar] [CrossRef] [Green Version]
  31. Crawford, S.D. Lichens used in traditional medicine. In Lichen Secondary Metabolites: Bioactive Properties and Pharmaceutical Potential; Springer International Publishing: Berlin/Heidelberg, Germany, 2015; pp. 27–80. ISBN 9783319133744. [Google Scholar]
  32. Crawford, S.D. Lichens Used in Traditional Medicine. In Lichen Secondary Metabolites; Springer International Publishing: Berlin/Heidelberg, Germany, 2019; pp. 31–97. [Google Scholar]
  33. Komaty, S.; Letertre, M.; Dang, D.; Jungnickel, H.; Laux, P. Sample preparation for an optimised extraction of localised metabolites in lichens: Application to Pseudevernia furfuracea. Talanta 2015. [Google Scholar] [CrossRef]
  34. Rafika, B.; Monia, A. Antibacterial Activity of the Chloroform, Acetone, Methanol and queous Extracts of Algerian Lichens. Jordan J. Pharm. Sci. 2018, 11, 55–67. [Google Scholar]
  35. Monia, A.A.; Rafika, B.; Tarek, H. Lichen diversity in the edough peninsula, North East of Algeria. Bot. Complut. 2018, 42, 9–18. [Google Scholar] [CrossRef] [Green Version]
  36. Brakni, R.; Serradj, M.; Fernandez, X.; Michel, T. Chemical and biological investigation of Algerian lichens. Planta Med. 2016, 82, P657. [Google Scholar] [CrossRef]
  37. Aljabeili, H.S.; Barakat, H.; Abdel-Rahman, H.A. Chemical Composition, Antibacterial and Antioxidant Activities of Thyme Essential Oil (Thymus vulgaris). Food Nutr. Sci. 2018, 9, 433–446. [Google Scholar] [CrossRef] [Green Version]
  38. Serradj, M.; El Oualidi, J.; Slimani, A.; Boumedris, Z. Contribution to the lichens;inventory from the Oubeira Lake (NE Algeria). Bull. Inst. Sci. Rabat Sect. Sci. Vie 2013, 35, 15–17. [Google Scholar]
  39. Amrani, S.; Nacer, A.; Noureddine, N.E.; Seaward, M.R. Lichenological exploration of Algeria: Historical overview and annotated bibliography, 1799–2013. Willdenowia 2015, 45, 15–34. [Google Scholar] [CrossRef] [Green Version]
  40. Rebbas, K.; Boutabia, L.; Touazi, Y.; Gharzouli, R.; Djellouli, Y.; Alatou, D. Inventaire des lichens du Parc national de Gouraya (Béjaïa, Algérie). Phytotherapie 2011, 9, 225–233. [Google Scholar] [CrossRef]
  41. Mohamed, A.H.; Miara, M.D.; Rebbas, K.; Slimani, A.; Ravera, S.; Hamerelain, A.S. Mise à jour de l’inventaire des lichens d’Algérie. Rev. Ecol. 2014, 10, 75–103. [Google Scholar]
  42. Rahali, M. Cartographie de la pollution plombique dans la région d’Alger en utilisant un lichen (Xanthoria parientina) comme bioaccumulateur. Pollut. Atmos. 2002, 421–432. [Google Scholar] [CrossRef] [Green Version]
  43. Khedim, R.; Maatoug, M.; Benhassaini, H.; Hammou, M.A. Macrolichens New to Algeria and Other Interesting Species from Theniet-el-Had National Park. Herzogia 2018, 31, 252–267. [Google Scholar] [CrossRef]
  44. Brakni, R.; Ali Ahmed, M.; Burger, P.; Schwing, A.; Michel, G.; Pomares, C.; Hasseine, L.; Boyer, L.; Fernandez, X.; Landreau, A.; et al. UHPLC-HRMS/MS Based Profiling of Algerian Lichens and Their Antimicrobial Activities. Chem. Biodivers. 2018, 15, e1800031. [Google Scholar] [CrossRef] [PubMed]
  45. Slimani, A.; Ahmed, A.; Serradj, M.; Hamel, T.; Coste, C. Contribution a l’etude de la flore lichenique dans la zeenaie de Bougous (foret de Ramel Tonal) au niveau du Pare National d’El Kala Nord Est Algerien. Synthèse Rev. Des Sci. Et De La Technol. 2013, 27, 22–29. [Google Scholar]
  46. Bouiadjra, S.B.; Ramdani, M.; Djellouli, R.; ELkhiati, N.; Zerey, W.E.; Bouterfas, K.; Roger, F. Air quality in the metropolitan area of Sidi Bel Abbes (Algeria) through the lichens species as bio-indicators. Moroc. J. Chem. 2017, 5. [Google Scholar] [CrossRef]
  47. Ghennam, K.; Fatiha, A. Biosurveillance par SIG de la qualité de l’air d’une cité méditerraneene (Alger), diversité lichénique, indice de pureté atmosphérique (IPA) et indice d’impact. Revue d’Écologie 2017, 72, 134–146. [Google Scholar]
  48. Maizi, N.; Alioua, A.; Tahar, A. Jumelage des bio-indicateurs et d’un réseau de surveillance de la qualité de l’air pour la détection de la pollution par le SO 2 dans la région de Annaba (Algérie). Biotechnol. Agron. Soc. Env. 2012, 16, 149–158. [Google Scholar]
  49. Serradj-Ali Ahmed, M.; Boumedris, Z.E.; Djebar, M.R.; Tahar, A. Responses of antioxidants in Flavoparmelia caperata (L.) Hale to the atmospheric pollution air at two urban and semi-urban areas in the region of Annaba (East of Algeria). Pollut. Atmos. 2014. [Google Scholar] [CrossRef] [Green Version]
  50. Djamel, S.; Yamna, D.; Djamel, A. Biological diversity of the National Park of El-Kala (Algeria), valorisation and protection. Biodivers. J. 2014, 5, 525–532. [Google Scholar]
  51. Putra, P.P.; Abdullah, S.S.; Rahmatunisa, R.; Junaidin, J.; Ismed, F. Structure, activity, and drug-likeness of pure compounds of Sumatran lichen (Stereocaulon halei) for the targeted ACE2 protein in COVID-19 disease. Pharmaciana 2020, 10, 135. [Google Scholar] [CrossRef]
  52. Mallavadhani, U.V.; Tirupathamma, R.S.; Sagarika, G.; Ramakrishna, S. Isolation, Chemical Modification, and Anticancer Activity of Major Metabolites of the Lichen Parmotrema mesotropum. Chem. Nat. Compd. 2019, 55, 825–831. [Google Scholar] [CrossRef]
  53. Nishiyama, Y.; Noda, Y.; Nakatani, N.; Shitan, N.; Sudo, T.; Kato, A.; Mutiso, P.B.C. Structure of constituents isolated from the bark of Cassipourea malosana and their cytotoxicity against a human ovarian cell line. J. Nat. Med. 2019, 73, 289–296. [Google Scholar] [CrossRef] [PubMed]
  54. Vladimirova, I.N.; Georgiyants, V.A. Extracted compounds from Cetraria islandica. Chem. Nat. Compd. 2013, 49, 347–348. [Google Scholar] [CrossRef]
  55. Akbar, S.; Akbar, S. Dolichousnea longissima (Ach) Articus/Parmelia perlata Esch. (Parmeliaceae). In Handbook of 200 Medicinal Plants; Springer International Publishing: Berlin/Heidelberg, Germany, 2020; pp. 873–878. [Google Scholar]
  56. Yu, X.L.; Yang, X.Y.; Gao, X.L.; Bai, R.F.; Yin, X.; Su, G.Z.; Qu, C.H.; Chai, X.Y.; Tu, P.F. Phenolic constituents from lichen Usnea longissima. Zhongguo Zhongyao Zazhi 2016, 41, 1864–1869. [Google Scholar] [CrossRef] [PubMed]
  57. Carpentier, C.; Queiroz, E.F.; Marcourt, L.; Wolfender, J.L.; Azelmat, J.; Grenier, D.; Boudreau, S.; Voyer, N. Dibenzofurans and pseudodepsidones from the lichen Stereocaulon paschale collected in northern Quebec. J. Nat. Prod. 2017, 80, 210–214. [Google Scholar] [CrossRef]
  58. Ari, F.; Ulukaya, E.; Oran, S.; Celikler, S.; Ozturk, S.; Ozel, M.Z. Promising anticancer activity of a lichen, Parmelia sulcata Taylor, against breast cancer cell lines and genotoxic effect on human lymphocytes. Cytotechnology 2015, 67, 531–543. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  59. Stojanović, I.Ž.; Radulović, N.S.; Mitrović, T.L.; Stamenković, S.M.; Stojanović, G.S. Volatile constituents of selected Parmeliaceae lichens. J. Serb. Chem. Soc. 2011, 76, 987–994. [Google Scholar] [CrossRef]
  60. Hoffmann, H.R.; Matusch, R.; Baniahmad, A. Isolation of Atraric Acid, Synthesis of Atraric Acid Derivatives, and Use of Atraric Acid and the Derivatives thereofe for the Treatment of Benigne Prostatic Hyperplasia, Prostate Carcinoma and Spinobulbar Muscular Atrophy. U.S. Patent 8,481,519, 9 July 2013. [Google Scholar]
  61. Hessenkemper, W.; Roediger, J.; Bartsch, S.; Houtsmuller, A.B.; van Royen, M.E.; Petersen, I.; Grimm, M.-O.; Baniahmad, A. A natural androgen receptor antagonist induces cellular senescence in prostate cancer cells. Mol. Endocrinol. 2014, 28, 1831–1840. [Google Scholar] [CrossRef] [Green Version]
  62. Güvenç, A.; Akkol, E.K.; Süntar, I.; Keleş, H.; Yildiz, S.; Çaliş, I. Biological activities of Pseudevernia furfuracea (L.) Zopf extracts and isolation of the active compounds. J. Ethnopharmacol. 2012, 144, 726–734. [Google Scholar] [CrossRef]
  63. Ahad, A.M.; Goto, Y.; Kiuchi, F.; Tsuda, Y.; Kondo, K.; Sato, T. Nematocidal principles in “Oakmoss Absolute” and nematocidal activity of 2, 4-Dihydroxybenzoates. Chem. Pharm. Bull. 1991, 39, 1043–1046. [Google Scholar] [CrossRef] [Green Version]
  64. Yamamoto, Y.; Watanabe, A. Fatty acid their composition of lichens and their phyco-and mycobionts. J. Gen. Appl. Microbiol. 1974, 20, 83–86. [Google Scholar] [CrossRef] [Green Version]
  65. Sonesson, M.; Grimberg, Å.; Sveinbjörnsson, B.; Carlsson, B.Å. Seasonal variation in concentrations of carbohydrates and lipids in two epiphytic lichens with contrasting, snow-depth related distribution on subarctic birch trees. Bryologist 2011, 114, 443–452. [Google Scholar] [CrossRef]
  66. Řezanka, T.; Dembitsky, V.M. Fatty acids of lichen species from Tian Shan Mountains. Folia Microbiol. 1999, 44, 643–646. [Google Scholar] [CrossRef]
  67. Vu, T.H.; Catheline, D.; Delmail, D.; Boustie, J.; Legrand, P.; Lohezic-Le Devehat, F. Gas chromatographic analysis to compare the fatty acid composition of fifteen lichen species, with a focus on Stereocaulon. Lichenologist 2016, 48, 323–337. [Google Scholar] [CrossRef]
  68. Culberson, C. Biogenetic relationships of the lichen substances in the framework of systematics. Bryologist 1986, 89, 91–98. [Google Scholar] [CrossRef]
  69. Fehrer, J.; Slavíková-Bayerová, Š.; Orange, A. Large genetic divergence of new, morphologically similar species of sterile lichens from Europe (Lepraria, Stereocaulaceae, Ascomycota): Concordance of DNA sequence data with secondary metabolites. Cladistics 2008, 24, 443–458. [Google Scholar] [CrossRef]
  70. Hale, M.E. A Monograph of the Lichen Genus Parmelia Acharius sensu Strict (Ascomycotina: Pameliaceae); Smithsonian Institution Press: Washington, DC, USA, 1987. [Google Scholar]
  71. Haugan, R.; Timdal, E. Tephromela perlata and T. talayana, with notes on the T. aglaea-complex. Graph. Scr. 1994, 6, 17–26. [Google Scholar]
  72. Spribille, T.; Klug, B.; Mayrhofer, H. A phylogenetic analysis of the boreal lichen Mycoblastus sanguinarius (Mycoblastaceae, lichenised Ascomycota) reveals cryptic clades correlated with fatty acid profiles. Mol. Phylogenet. Evol. 2011, 59, 603–614. [Google Scholar] [CrossRef] [PubMed]
  73. Lumbsch, H.T. The Use of Metabolic Data in Lichenology at the Species and Subspecific Levels. Lichenologist 1998, 30, 357–367. [Google Scholar] [CrossRef]
  74. Hawksworth, D.L. Lichen chemotaxonomy. In Lichenology: Progress and Problems; Proceedings of an international Symposium; Academic Press: Cambridge, MA, USA, 1976; pp. 139–184. [Google Scholar]
  75. Stocker-Wörgötter, E. Metabolic diversity of lichen-forming ascomycetous fungi: Culturing, polyketide and shikimate metabolite production, and PKS genes. Nat. Prod. Rep. 2008, 25, 188–200. [Google Scholar] [CrossRef]
  76. Sumida, C.; Graber, R.; Nunez, E. Role of fatty acids in signal transduction: Modulators and messengers. Prostaglandins Leukot. Essent. Fat. Acids 1993, 48, 117–122. [Google Scholar] [CrossRef]
  77. Bychek-Gushchina, I. Study of biochemical aspects of lichen symbiosis. I. Lipids and fatty acids in cultured lichen symbionts. Biochemistry 1997, 62, 490–494. [Google Scholar]
  78. Torres, A.; Dor, I.; Rotem, J.; Srebnik, M.; Dembitsky, V.M. Characterization of surface n-alkanes and fatty acids of the epiphytic lichen Xanthoria parietina, its photobiont a green alga Trebouxia sp., and its mycobiont, from the Jerusalem hills. Eur. J. Biochem. 2003, 270, 2120–2125. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  79. Molina, M.C.; Crespo, A.; Vicente, C.; Elix, J.A. Differences in the composition of phenolics and fatty acids of cultured mycobiont and thallus of Physconia distorta. Plant Physiol. Biochem. 2003, 41, 175–180. [Google Scholar] [CrossRef]
  80. Dembitsky, V.M.; Rezanka, T.; Bychek, I.A. Seasonal variation of lipids and fatty acids from tree-growing lichens of the genus Physcia. Phytochemistry 1994, 36, 601–608. [Google Scholar] [CrossRef]
  81. Dembitsky, V.M.; Rezanka, T.; Bychek, I. A Seasonal variability of lipids and fatty acids in the tree-growing lichen Xanthoria parientina L. J. Exp. Bot. 1994, 45, 403–408. [Google Scholar] [CrossRef]
  82. Reis, R.A.; Iacomini, M.; Gorin, P.A.J.; Souza, L.M.; Grube, M.; Cortes Cordeiro, L.M.; Sassaki, G.L. Fatty acid composition of the tropical lichen Teloschistes flavicans and its cultivated symbionts. FEMS Microbiol. Lett. 2005, 247, 1–6. [Google Scholar] [CrossRef] [Green Version]
  83. Goss, R.; Wilhelm, C. Lipids in Algae, Lichens and Mosses. In Lipids in Photosynthesis; Springer: Dordrecht, The Netherlands, 2009; pp. 117–137. [Google Scholar]
  84. Temina, M.; Levitsky, D.O.; Dembitsky, V.M. Chemical Constituents of the Epiphytic and Lithophilic Lichens of the Genus Collema. Rec. Nat. Prod. 2010, 4, 79–86. [Google Scholar]
  85. Ilica, R.A.; Kloetzer, L.; Galaction, A.I.; Caşcaval, D. Fumaric acid: Production and separation. Biotechnol. Lett. 2019, 41, 47–57. [Google Scholar] [CrossRef] [PubMed]
  86. Sebastian, J.; Hegde, K.; Kumar, P.; Rouissi, T.; Brar, S.K. Bioproduction of fumaric acid: An insight into microbial strain improvement strategies. Crit. Rev. Biotechnol. 2019, 39, 817–834. [Google Scholar] [CrossRef]
  87. Haghikia, A.; Linker, R.; Gold, R. Fumarsäure in der Therapie der Multiplen Sklerose. Nervenarzt 2014, 85, 720–726. [Google Scholar] [CrossRef] [PubMed]
  88. Papadaki, A.; Papapostolou, H.; Alexandri, M.; Kopsahelis, N.; Papanikolaou, S.; de Castro, A.M.; Freire, D.M.G.; Koutinas, A.A. Fumaric acid production using renewable resources from biodiesel and cane sugar production processes. Environ. Sci. Pollut. Res. 2018, 25, 35960–35970. [Google Scholar] [CrossRef] [PubMed]
  89. Jabłońska-Trypuć, A.; Pankiewicz, W.; Czerpak, R. Traumatic Acid Reduces Oxidative Stress and Enhances Collagen Biosynthesis in Cultured Human Skin Fibroblasts. Lipids 2016, 51, 1021–1035. [Google Scholar] [CrossRef] [Green Version]
  90. Jabłońska-Trypuć, A.; Krętowski, R.; Wołejko, E.; Wydro, U.; Butarewicz, A. Traumatic acid toxicity mechanisms in human breast cancer MCF-7cells. Regul. Toxicol. Pharmacol. 2019, 106, 137–146. [Google Scholar] [CrossRef] [PubMed]
  91. Lee, H.C.; Liu, F.C.; Tsai, C.N.; Chou, A.H.; Liao, C.C.; Yu, H.P. Esculetin Ameliorates Lipopolysaccharide-Induced Acute Lung Injury in Mice Via Modulation of the AKT/ERK/NF-κB and RORγt/IL-17 Pathways. Inflammation 2020, 43, 962–974. [Google Scholar] [CrossRef] [PubMed]
  92. Wang, C.; Pei, A.; Chen, J.; Yu, H.; Sun, M.L.; Liu, C.F.; Xu, X. A natural coumarin derivative esculetin offers neuroprotection on cerebral ischemia/reperfusion injury in mice. J. Neurochem. 2012, 121, 1007–1013. [Google Scholar] [CrossRef]
  93. Hongyan, L. Esculetin Attenuates Th2 and Th17 Responses in an Ovalbumin-Induced Asthmatic Mouse Model. Inflammation 2016, 39, 735–743. [Google Scholar] [CrossRef] [PubMed]
  94. Han, M.H.; Park, C.; Lee, D.S.; Hong, S.H.; Choi, I.W.; Kim, G.Y.; Choi, S.H.; Shim, J.H.; Chae, J.I.; Yoo, Y.H.; et al. Cytoprotective effects of esculetin against oxidative stress are associated with the upregulation of Nrf2-mediated NQO1 expression via the activation of the ERK pathway. Int. J. Mol. Med. 2017, 39, 380–386. [Google Scholar] [CrossRef] [Green Version]
  95. Srikrishna, D.; Godugu, C.; Dubey, P.K. A Review on Pharmacological Properties of Coumarins. Mini-Rev. Med. Chem. 2016, 18, 113–141. [Google Scholar] [CrossRef] [PubMed]
  96. Stefanachi, A.; Leonetti, F.; Pisani, L.; Catto, M.; Carotti, A. Coumarin: A Natural, Privileged and Versatile Scaffold for Bioactive Compounds. Molecules 2018, 23, 250. [Google Scholar] [CrossRef] [Green Version]
  97. Musa, M.; Cooperwood, J.; Khan, M.O. A Review of Coumarin Derivatives in Pharmacotherapy of Breast Cancer. Curr. Med. Chem. 2008, 15, 2664–2679. [Google Scholar] [CrossRef] [Green Version]
  98. Studzińska-Sroka, E.; Galanty, A.; Bylka, W. Atranorin-an interesting lichen secondary metabolite. Mini-Rev. Med. Chem. 2017, 17, 1633–1645. [Google Scholar] [CrossRef] [PubMed]
  99. Huneck, S.; Feige, G.B.; Schmidt, J. Chemie von Cladonia furcata und Cladonia rangiformis. Herzogia 2004, 17, 51–58. [Google Scholar]
  100. Nybakken, L.; Helmersen, A.M.; Gauslaa, Y.; Selås, V. Lichen compounds Restrain lichen feeding by bank voles (Myodes glareolus). J. Chem. Ecol. 2010, 36, 298–304. [Google Scholar] [CrossRef]
  101. Culberson, C.F. Chemical and Botanical Guide to Lichen Products, 2nd ed.; Otto Koeltz Science Publishers: Koenigstein, Germany, 1979. [Google Scholar]
  102. Elix, J.A. A Catalogue of Standardized Chromatographic Data and Biosynthetic Relationships for Lichen Substances; Australian National University: Canberra, Australia, 2014. [Google Scholar]
  103. Ranković, B.; Mišić, M.; Sukdolak, S. The antimicrobial activity of substances derived from the lichens Physcia aipolia, Umbilicaria polyphylla, Parmelia caperata and Hypogymnia physodes. World J. Microbiol. Biotechnol. 2008, 24, 1239–1242. [Google Scholar] [CrossRef]
  104. Solhaug, K.A.; Larsson, P.; Gauslaa, Y. Light screening in lichen cortices can be quantified by chlorophyll fluorescence techniques for both reflecting and absorbing pigments. Planta 2010, 231, 1003–1011. [Google Scholar] [CrossRef]
  105. Brodo, I.M.; Freebury, C.; Alfonso, N. Notes on the Lichens Physcia aipolia and P. alnophila in North America. Evansia 2013, 30, 110–119. [Google Scholar] [CrossRef]
  106. Maldonado Montaño, A.; Menesses, R.; Bravo, J.A.; Vila, J.L.; Bello, A. Presence of atranorin in Physcia sorediosa. Rev. Boliv. Quím. 2016, 33, 175–178. [Google Scholar]
  107. Kosanić, M.; Ranković, B.; Stanojković, T.; Rančić, A.; Manojlović, N. Cladonia lichens and their major metabolites as possible natural antioxidant, antimicrobial and anticancer agents. LWT Food Sci. Technol. 2014, 59, 518–525. [Google Scholar] [CrossRef]
  108. Pompilio, A.; Pomponio, S.; Di Vincenzo, V.; Crocetta, V.; Nicoletti, M.; Piovano, M.; Garbarino, J.A.; Di Bonaventura, G. Antimicrobial and antibiofilm activity of secondary metabolites of lichens against methicillin-resistant Staphylococcus aureus strains from cystic fibrosis patients. Future Microbiol. 2013, 8, 281–292. [Google Scholar] [CrossRef] [PubMed]
  109. Melo, M.G.D.; Araújo, A.A.S.; Serafini, M.R.; Carvalho, L.F.; Bezerra, M.S.; Ramos, C.S.; Bonjardim, L.R.; Albuquerque-Júnior, R.L.C.; Lima, J.T.; Siqueira, R.S.; et al. Anti-inflammatory and toxicity studies of atranorin extracted from Cladina kalbii Ahti in rodents. Braz. J. Pharm. Sci. 2011, 47, 861–872. [Google Scholar] [CrossRef] [Green Version]
  110. Melo, M.G.D.; Araújo, A.A.S.; Rocha, C.P.L.; Almeida, E.M.S.A.; De Souza Siqueira, R.; Bonjardim, L.R.; Quintans-Júnior, L.J. Purification, physicochemical properties, thermal analysis and antinociceptive effect of atranorin extracted from Cladina kalbii. Biol. Pharm. Bull. 2008, 31, 1977–1980. [Google Scholar] [CrossRef] [Green Version]
  111. Barreto, R.S.S.; Albuquerque-Júnior, R.L.C.; Pereira-Filho, R.N.; Quintans, J.S.S.; Barreto, A.S.; DeSantana, J.M.; Santana-Filho, V.J.; Santos, M.R.V.; Bonjardim, L.R.; Araújo, A.A.S.; et al. Evaluation of wound healing activity of atranorin, a lichen secondary metabolite, on rodents. Braz. J. Pharmacogn. 2013, 23, 310–319. [Google Scholar] [CrossRef] [Green Version]
  112. Harikrishnan, A.; Veena, V.; Lakshmi, B.; Shanmugavalli, R.; Theres, S.; Prashantha, C.N.; Shah, T.; Oshin, K.; Togam, R.; Nandi, S. Atranorin, an antimicrobial metabolite from lichen Parmotrema rampoddense exhibited in vitro anti-breast cancer activity through interaction with Akt activity. J. Biomol. Struct. Dyn. 2020. [Google Scholar] [CrossRef] [PubMed]
  113. Galanty, A.; Koczurkiewicz, P.; Wnuk, D.; Paw, M.; Karnas, E.; Podolak, I.; Węgrzyn, M.; Borusiewicz, M.; Madeja, Z.; Czyż, J.; et al. Usnic acid and atranorin exert selective cytostatic and anti-invasive effects on human prostate and melanoma cancer cells. Toxicol. Vitr. 2017, 40, 161–169. [Google Scholar] [CrossRef] [PubMed]
  114. Hikino, H.; Ito, K.; Takemoto, T. Stereostructure of Asebotoxin I and II, Toxins of Pieris japonica. Chem. Pharm. Bull. 1969, 17, 854–855. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  115. Zhang, L.; Hong, Z.; Zhang, R.R.; Sun, X.Z.; Yuan, Y.F.; Hu, J.; Wang, X. Bakkenolide A inhibits leukemia by regulation of HDAC3 and PI3K/Akt-related signaling pathways. Biomed. Pharmacother. 2016, 83, 958–966. [Google Scholar] [CrossRef]
  116. Solberg, Y. Chemical constituents of the lichen species Cetraria islandica. J. Hattori Bot. Lab. 1986, 60, 391–406. [Google Scholar]
  117. Simirgiotis, M.J.; Quispe, C.; Areche, C.; Sepúlveda, B. Phenolic compounds in Chilean Mistletoe (quintral, Tristerix tetrandus) analysed by UHPLC-Q/Orbitrap/MS/MS and its antioxidant properties. Molecules 2016, 21, 245. [Google Scholar] [CrossRef] [Green Version]
  118. Huneck, S. New Results on the Chemistry of Lichen Substances. In Progress in the Chemistry of Organic Natural Products; Springer: Berlin/Heidelberg, Germany, 2001; Volume 81. [Google Scholar]
  119. Seo, D.Y.; Lee, S.R.; Heo, J.W.; No, M.H.; Rhee, B.D.; Ko, K.S.; Kwak, H.B.; Han, J. Ursolic acid in health and disease. Korean J. Physiol. Pharmacol. 2018, 22, 235–248. [Google Scholar] [CrossRef] [Green Version]
  120. Gonzalez, A.G.; Bermejo Barrera, J.; Rodriguez Perez, E.M.; Hernandez Padron, C.E. chemical constituents of the lichen Ramalina hierrensis. Planta Med. 1992, 58, 214–218. [Google Scholar] [CrossRef] [PubMed]
  121. Vu, T.H.; Lamer, A.C.L.; Lalli, C.; Samson, J.B.M.; Dévéhat, F.L.L.; Seyec, J. Le Depsides: Lichen metabolites active against hepatitis C virus. PLoS ONE 2015, 10, e0120405. [Google Scholar] [CrossRef] [PubMed]
  122. Rubio, C.; Galloway, D.J.; Quilhot, W. Secondary products from Pannaria tavaresii (Lichens). J. Chil. Chem. Soc. 2005, 50, 667–669. [Google Scholar] [CrossRef]
  123. De Carvalho, M.G.; De Carvalho, G.J.A.; Braz-Filho, R. Chemical Constituents from Ouratea floribunda: Complete 1H and 13C NMR Assignments of Atranorin and its New Acetyl Derivative. J. Braz. Chem. Soc. 2000, 11, 143–147. [Google Scholar] [CrossRef]
  124. Norouzi, H.; Azizi, A.; Gholami, M.; Sohrabi, M.; Boustie, J. Chemotype variations among lichen ecotypes of Umbilicaria aprina as revealed by LC-ESI-MS/MS: A survey of antioxidant phenolics. Environ. Sci. Pollut. Res. 2020, 27, 40296–40308. [Google Scholar] [CrossRef] [PubMed]
  125. Vos, C.; McKinney, P.; Pearson, C.; Heiny, E.; Gunawardena, G.; Holt, E.A. The optimal extraction and stability of atranorin from lichens, in relation to solvent and pH. Lichenologist 2018, 50, 499–512. [Google Scholar] [CrossRef]
  126. Lohezic-Le Devehat, F.; Legouin, B.; Couteau, C.; Boustie, J.; Coiffard, L. Lichenic extracts and metabolites as UV filters. J. Photochem. Photobiol. B Biol. 2013, 120, 17–28. [Google Scholar] [CrossRef]
  127. Bjerke, J.W.; Elvebakk, A.; Domínguez, E.; Dahlback, A. Seasonal trends in usnic acid concentrations of Arctic, alpine and Patagonian populations of the lichen Flavocetraria nivalis. Phytochemistry 2005, 66, 337–344. [Google Scholar] [CrossRef]
  128. Marante, F.J.T.; Castellano, A.G.; Rosas, F.E.; Aguiar, J.Q.; Barrera, J.B. Identification and quantitation of allelochemicals from the lichen Lethariella canariensis: Phytotoxicity and antioxidative activity. J. Chem. Ecol. 2003, 29, 2049–2071. [Google Scholar] [CrossRef]
  129. Solhaug, K.A.; Lind, M.; Nybakken, L.; Gauslaa, Y. Possible functional roles of cortical depsides and medullary depsidones in the foliose lichen Hypogymnia physodes. Flora Morphol. Distrib. Funct. Ecol. Plants 2009, 204, 40–48. [Google Scholar] [CrossRef]
  130. Forbes, P.; Wat, L.; Strumpher, J. Comparative Perspectives on Extraction Methods for Organic Metabolites and Pollutants from Lichens. In Lichen-Derived Products; Wiley: Hoboken, NJ, USA, 2020; pp. 27–73. [Google Scholar]
  131. Augusto, S.; Pinho, P.; Branquinho, C.; Pereira, M.J.; Soares, A.; Catarino, F. Atmospheric Dioxin and Furan Deposition in Relation to Land-Use and Other Pollutants: A Survey with Lichens; Kluwer Academic Publishers: Dordrecht, The Netherlands, 2004; Volume 49. [Google Scholar]
  132. Van der Wat, L.; Forbes, P.B.C. Comparison of extraction techniques for polycyclic aromatic hydrocarbons from lichen biomonitors. Environ. Sci. Pollut. Res. 2019, 26, 11179–11190. [Google Scholar] [CrossRef]
  133. Van Der Wat, L.; Forbes, P.B.C. Lichens as biomonitors for organic air pollutants. TrAC Trends Anal. Chem. 2015, 64, 165–172. [Google Scholar] [CrossRef] [Green Version]
  134. Caccamese, S.; Toscano, R.M.; Bellesia, F.; Pinetti, A. Methyl β-Orcinolcarboxylate and Depsides from Parmelia Furfuracea. J. Nat. Prod. 1985, 48, 157–158. [Google Scholar] [CrossRef]
  135. Gunzinger, J.; Tabacchi, R. Isolement et identification de l’acide furfurique, nouvelle depsidone du lichen Pseudevernia furfuracea (L.) Ach. Helv. Chim. Acta 1985, 68, 1936–1939. [Google Scholar] [CrossRef]
  136. Culberson, W.L.; Culberson, C.F.; Johnson, A. Pseudevernia furfuracea-Olivetorina Relationships: Chemistry and Ecology. Mycologia 1977, 69, 604. [Google Scholar] [CrossRef]
  137. Joulain, D.; Tabacchi, R. Lichen extracts as raw materials in perfumery. Part 2: Treemoss. Flavour Fragr. J. 2009, 24, 105–116. [Google Scholar] [CrossRef] [Green Version]
Molecules 26 01121 g001 550
Figure 1. Algerian Physcia mediterranea Nimis photographed in the national park of El Kala, Algeria.
Figure 1. Algerian Physcia mediterranea Nimis photographed in the national park of El Kala, Algeria.
Molecules 26 01121 g001
Molecules 26 01121 g002 550
Figure 2. GC-MS chromatogram of Physcia mediterranea (ME-hex part). RT = retention time. (1), (2), (3), (4) = derivatives of methylbenzoic acids; (5), (6), (7), (8) = fatty acids; (9) = diterpene; (10) = unknown.
Figure 2. GC-MS chromatogram of Physcia mediterranea (ME-hex part). RT = retention time. (1), (2), (3), (4) = derivatives of methylbenzoic acids; (5), (6), (7), (8) = fatty acids; (9) = diterpene; (10) = unknown.
Molecules 26 01121 g002
Molecules 26 01121 g003 550
Figure 3. Chemical structure of (a) (−)-ent-Kauran-16a-ol and (b) Atraric acid.
Figure 3. Chemical structure of (a) (−)-ent-Kauran-16a-ol and (b) Atraric acid.
Molecules 26 01121 g003
Molecules 26 01121 g004 550
Figure 4. Chemical structure of Compound 3, 4, 5, 6, 8, 9, 12, 14, and 38 identified in the ME-hex of Physcia mediterranea by UHPLC/ESI/MS/MS.
Figure 4. Chemical structure of Compound 3, 4, 5, 6, 8, 9, 12, 14, and 38 identified in the ME-hex of Physcia mediterranea by UHPLC/ESI/MS/MS.
Molecules 26 01121 g004
Molecules 26 01121 g005 550
Figure 5. Chemical structure of compound 7, 11, 13, 20, 23, 31, 32, 37, and 39 identified in ME-hex of Physcia mediterranea by UHPLC/ESI/MS/MS.
Figure 5. Chemical structure of compound 7, 11, 13, 20, 23, 31, 32, 37, and 39 identified in ME-hex of Physcia mediterranea by UHPLC/ESI/MS/MS.
Molecules 26 01121 g005
Molecules 26 01121 g006 550
Figure 6. Chemical structure of atranorin isolated from WA-hex of Physcia mediterranea.
Figure 6. Chemical structure of atranorin isolated from WA-hex of Physcia mediterranea.
Molecules 26 01121 g006
Molecules 26 01121 g007 550
Figure 7. The topography of Algeria, indicating El Kala region boundaries (36°49′ 09″ N; 08°31′ 33″ E).
Figure 7. The topography of Algeria, indicating El Kala region boundaries (36°49′ 09″ N; 08°31′ 33″ E).
Molecules 26 01121 g007
Table
Table 1. Identification of metabolites in ME-hex of Physcia mediterranea by GC-MS analysis.
Table 1. Identification of metabolites in ME-hex of Physcia mediterranea by GC-MS analysis.
N°RTCompoundMass%Synonyms
119:56Methyl 2-hydroxy-4-methoxy-6-methylbenzoate1960.3Orsellinic Acid Methyl Ester 4-Methyl Ether (Sparassol)C10H12O4
220:01Methyl 2,4-dihydroxy-3,5,6-trimethylbenzoate21052.3Derivatives of methylbenzoic acidsC11H14O4
320:43Methyl 4-hydroxy-2-methoxy-3,6-dimethylbenzoate2101.12-metoxy-Atraric acidC10H12O4
420:64Methyl 2,4-dihydroxy-3,6-dimethylbenzoate1962.3Atraric acidC10H12O4
522:55Methyl hexadecanoate27024.2Palmitic acidC16H32O2
623:5313-Methyl-17-norkaur-15-ene2722.4Linoleic acidC18H32O2
724:19Methyl cis, cis-9,12-octadecadienoate2943.2Oleic acidC18H34O2
824:25Methyl (Z)-9-octadecenoate2964.9Stearic acidC18H34O2
924:48Methyloctadecanoate2983.8(−)-ent-Kauran-16α-olC2OH34O
1026:21Unknown3762.2Unknown
T% 95.5
RT = retention time; T% = total of compounds identified (%) in the extracts.
Table
Table 2. Identification of metabolites in WA-hex of Physcia mediterranea by UHPLC/ESI/MS/MS.
Table 2. Identification of metabolites in WA-hex of Physcia mediterranea by UHPLC/ESI/MS/MS.
N°RT[M − H]−TMMMMTCompounds Identification
10.03146.9397147.9475C4H4O6ADihydroxyfumaric acid
21.80116.9276117.9354C4H6O4AButendioic acid
32.72187.0971188.1049C9H16O2A3,5-dimethoxyciclohexanecarboxylic acid
45.51163.0395164.0473C9 H8 O3Bp-Coumaric acid
56.57227.1286228.1364C12H20O4ATrans-dodec-2-enedioic acid (traumatic acid)
67.46282.2077283.2155C16 H29 NO3AN-dodecanoyl-l-Homoserine lactone
78.09174.9556175.9634C5 H4 O7A2-Hydroxy-3,4-dioxopentanedioc acid
88.15293.1762294.1840C17H26O4BGingerol
914.37295.2280296.2358C18 H32 O3A18-Hydroxylinoleic acid
1015.36177.0186178.0264C9 H6 O4B6,7-Dihydroxycoumarin (esculetin)
1115.44373.0929374.1007C19 H18 O8CAtranorin
1216.47407.0539408.0617C19 H17 O8ClCChloroatranorin
1317.57277.2175278.2203C18H30O2AOctadeca-9,12,15-trienoic acid (linolenelaidic acid)
1417.77265.1480266.1558C15 H22 O4A(4E,6E,9E)-Pentadeca-4,6,9-trienedioic acid
1518.03253.2331254.2249C16 H30 O2APalmitoleic acid
1618.34241.2173242.2251C15H30O2APentadecanoic acid
1718.50279.0936280.2409C18 H32 O2ALinoleic acid
1818.91267.2331268.2409C17 H32 O2Acis-9-Heptadecenoic acid (margaroleic acid)
1919.15255.2329256.2407C16 H32 O2APalmitic acid
2019.35459.3271460.3349C25H48O7UnknownUnknown
2119.51281.2487282.2565C18 H34 O2AOleic acid
2219.75459.3272460.3350C25 H48 O7AMethyl glucose isostearate
2319.89269.2488270.2566C17 H34 O2AHeptadecanoic acid (margaric acid)
2419.99307.2645308.2723C20 H36 O2A11,14-Eicosadienoic acid
2520.07457.3722458.3800C27 H54 O5UnknownUnknown
2620.18295.2645296.2723C19H36O2A10E-nonadecenoic acid
2720.54283.2643284.2721C18 H36 O2AStearic acid (octadecanoic acid)
2820.82309.2800310.2878C20 H38 O2AEicosenoic acid (gondoic acid)
2921.71311.2957312.3035C20H40O2AArachidic acid (eicosanoic acid)
3022.62297.1532298.1610C12H26O8UnknownUnknown
3122.77339.3268340.3346C22H44O2ADocosanoic acid (behenic acid)
3223.08309.1743310.1821C17 H26 O5DPortentol
3323.58353.2003354.2081C19H30O6UnknownUnknown
3423.81311.1689312.1767C13 H30 O8UnknownUnknown
3523.89367.3579368.3657C24H48O2ALignoceric acid (tetracosanoic acid)
3624.01397.2266398.2344C21H34O7DStephanol
3724.60293.1793294.1871C17H26O4AHeptadecatrienedioic acid
3824.94325.1844326.1922C14 H30 O8UnknownUnknown
3925.33395.3895396.3973C26H52O2AHexacosanoic acid or cerotic acid
4025.89337.2055338.2133C19 H30 O5A6-Oxononadeca-8,11-dienedioic acid
4126.10339.2000340.2078C15 H32 O8UnknownUnknown
4226.24381.2317382.2395C21 H34 O6A19-Acetoxylichesterinic acid
4326.75425.2581426.2659C23H38O7EAsebotoxin I
4427.53321.2106322.2184C19H30O4ANonadecatrienedioic acid
4527.85304.9143305.9221UnknownUnknownUnknown
RT = retention time; TM = theoretical mass (m/z); [M − H]− = measured mass as negative ion (m/z); MM = molecular mass; MT = metabolite type; A = paraconic and aliphatic acids; B = phenolic compounds; C = depsides; D = cycloaliphatic compounds; E = diterpenes.
Table
Table 3. Identification of metabolites in ME-hex of Physcia mediterranea by UHPLC/ESI/MS/MS.
Table 3. Identification of metabolites in ME-hex of Physcia mediterranea by UHPLC/ESI/MS/MS.
N°RT[M – H]−TMMMMTCompoundsMS2 Ions
12.72187.0970188.1043C9H16O4A3,5-Dimethoxyciclohexanecarboxylic acid141.8670; 123.0807
22.75293.0669294.0741C14H14O7A6-(Hydroxymethyl)-3,5-bis(methoxycarbonyl)-2,4-dimethylcyclohex-1-ene-1-carboxylic acid234.0530; 207.0695
33.08243.1239244.1311C12H20O5A3,5,6-Hydroxymethyl-2,4-dimethylcyclohex-1-ene-1-carboxylic acid183.1025; 176.6474
43.15151.0393152.0465C8H8O3BAtranol123.0444; 105.0335; 81.0335
53.20225.1129226.1201C12H18O4A5-Formyl-3-hydroxymethyl-2,4,6-trimethylcyclohex-1-ene-1-carboxylic acid207.1023; 97.0284
63.47199.0973200.1046C10H16O4A3,5-Dihydroxy-2,4,6-trimethylciclohexenecarboxilic acid183.4492; 162.8385; 114.9508
73.58149.0237150.0310C8H6O3C4-Formylbenzoic acid−
83.82241.1081242.1153C12H18O5A5-Formyl-3,6-dihydroxymethyl-2,4-dimethylcyclohex-1-enecarboxylic acid218.816; 172.6485
94.10201.1129202.1202C10H18O4A2,4-Dihydroxy-3,5,6-trimethylcyclohexane-1-carboxylic acid197.6263; 164.8350; 139.1124
104.95199.1337200.1409C11H20O3A2-Hydroxy-10-undecenoic acid−
115.00185.0006186.0079C8H7ClO3BChloroatranol−
125.21169.0863170.0936C9H14O3A4-Hydroxy-2,5-dimethylcyclohex-1-ene-1-carboxylic acid124.9796; 104.4180
135.49163.0392164.0470C9H8O3Dp-Coumaric acid−
145.60209.1181210.1253C12H18O3A6-(1-Oxopentyl)-1-cyclohexene-1-carboxylic acid146.9850; 105.0336
155.87215.1286216.1359C11H20O4AUndecanedioic acid−
166.57227.1288228.1360C12H20O4Atrans-Dodec-2-enedioic acid−
177.44282.2078283.2150C16H29NO3AN-Dodecanoyl-l-Homoserine lactone−
189.53243.1601244.1675C13H24O4ATridecanedioic acid−
1910.59311.2230312.2302C18H32O4A9Z-Octadecenedioic acid−
2010.87233.1547234.1619C15H22O2EFukinanolide−
2110.89323.2230324.2303C19H32O4Fallo-Protolichesterinic acid−
2212.28313,2388314.2266C18H34O4AOctadecanedioic acid−
2313.04389.1245390.1315C20H22O8B8-Hydroxydiffractaic acid−
2413.34293.2124294.2202C18H30O3A2-Hydroxylinolenic acid−
2513.58291.1968292.2041C18H28O3Aα-Licanic acid−
2614.38295.2279296.2351C18H32O3A2-Hydroxylinoleic acid−
2714.40295,2278296,2352C18H32O3A18-hydroxylinoleic acid−
2815.19297.2436298.2508C18H34O3A9-Oxooctadecanoic acid−
2915.25295.2280296.2351C18H32O3ACoriolic acid−
3015.36297.2435298.2508C18H34O3ARicinoleic acid−
3115.43177.0187178.0259C9H6O4D6,7-Dihydroxycoumarin−
3215.46365.2330366.2403C21H34O5GMuronic acid−
3316.37471.3481472.3553C30H48O4UnknownUnknown−
3416.47210.9834211.9873C9H5ClO4H7-chloro-3-oxo-1,3-dihydroisobenzofuran-5-carboxylic acid138.9948, 103.0180
3516.51407.0540408.0611C19H17ClO8BChloroatranorin−
3617.59387.2544388.2616C24H36O4UnknownUnknown−
3717.68389.1242390.1314C20H22O8ILeprolomin−
3818.04253.2173254.2244C16H30O2APalmitoleic acid−
3918.33241.2172242.2245C15H30O2APentadecanoic acid−
4018.35455.3531456.3605C30H48O3GUrsolic acid−
4118.50279.2330280.2403C18H32O2ALinoleic acid−
4218.66299.2595300.2667C18H36O3A2-Hydroxyoctadecanoic acid−
4318.78279.2332280.2403C18H32O2ALinoleic acid−
4419.16255.2329256.2401C16H32O2APalmitic acid−
4519.49281.2485282.2559C18H34O2AOleic acid−
4619.62269.2488270.2561C17H34O2A15-Methylhexadecanoic acid−
4719.89269.2488270.2561C17H34O2AHeptadecanoic acid−
4820.07327.2543328.2616C20H40O3A2-Hydroxyeicosanoic acid−
4920.54283.2643284.2716C18H36O2AStearic acid (octadecanoic acid)−
5020.74309.2801310.2875C20H38O2AEicosenoic acid (Gondoic acid)−
5121.14297.2801298.2873C19H38O2ANonadecanoic acid−
5221.70311.2957312.3029C20H40O2AArachidic acid (Eicosanoic acid)−
5322.68637.4841638.4908C41 H66 O5UnknownUnknown−
5424.51605.3483606. 3561C37H50O7UnknownUnknown−
RT = retention time (min); TM = theoretical mass (m/z); [M – H]− = measured mass as negative ion (m/z); MM = molecular mass; MT = metabolite type; A = paraconic and aliphatic acids; B = depsides; C = carboxybenzaldehyde; D = phenolic acids; E = sesquiterpene lactones; F = depsones; G = triterpene; H = carboxyphthalide; I = diphenyl ether.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Kerboua, M.; Ahmed, M.A.; Samba, N.; Aitfella-Lahlou, R.; Silva, L.; Boyero, J.F.; Raposo, C.; Lopez Rodilla, J.M. Phytochemical Investigation of New Algerian Lichen Species: Physcia Mediterranea Nimis. Molecules 2021, 26, 1121. https://doi.org/10.3390/molecules26041121

AMA Style

Kerboua M, Ahmed MA, Samba N, Aitfella-Lahlou R, Silva L, Boyero JF, Raposo C, Lopez Rodilla JM. Phytochemical Investigation of New Algerian Lichen Species: Physcia Mediterranea Nimis. Molecules. 2021; 26(4):1121. https://doi.org/10.3390/molecules26041121

Chicago/Turabian Style

Kerboua, Marwa, Monia Ali Ahmed, Nsevolo Samba, Radhia Aitfella-Lahlou, Lucia Silva, Juan F. Boyero, Cesar Raposo, and Jesus Miguel Lopez Rodilla. 2021. "Phytochemical Investigation of New Algerian Lichen Species: Physcia Mediterranea Nimis" Molecules 26, no. 4: 1121. https://doi.org/10.3390/molecules26041121

Article Metrics

Back to TopTop