Environmental drivers of ground-floor bryophytes diversity in temperate forests.

Bryophytes play important role in forest ecosystem functioning and their distribution and diversity are driven by numerous environmental factors. The aim of the present study was to bring new insights in deeper understanding of terrestrial bryophytes diversity in temperate forests, as well as to determine the environmental factors which have predominant influence on ground-floor bryophytes. The survey was conducted in Fruska Gora Mountain (Serbia) across seven forest sites where ground-bryophytes were sampled. Soil moisture, temperature, and pH were measured as soil characteristics, while herbaceous cover, litter cover, stream distance, number of trees and shrubs were used as characteristics of stand structure. Species richness, Shannon diversity index, and evenness index were used as diversity measures. Generalised linear model and canonical correspondence analysis (CCA) were used to test the influence of environmental variables on bryophyte diversity. Litter cover was the most important explanatory variable, followed by soil moisture, stream distance and tree number, respectively. Overall, the stand structure was found to have a greater impact on ground-floor bryophyte diversity compared to soil characteristics. Identification of the most significant ecological factors affecting the diversity and distribution of bryophytes in forest ecosystems is of great importance in forest ecology with the aim of defining adequate management methods to preserve the biodiversity of forests, with particular emphasis on endangered and rare bryophyte species.

Bis-Bibenzyls from the Liverwort Pellia endiviifolia and Their Biological Activity.

Based on previous investigations where bis-bibenzyls isolated from liverworts showed various biological activities (cytotoxic, antimicrobial, and antiviral), we investigated their cytotoxic activity in several human cancer cell lines. From the methylene-chloride/methanol extract of the liverwort Pellia endiviifolia, three bis-bibenzyls of the perrottetin type were isolated, namely perrottetin E, 10'-hydroxyperrottetin E, and 10,10'-dihydroxyperrottetin E. The last two were found for the first time in this species. Their structures were resolved using 1D and 2D NMR, as well as by comparison with data in the literature. Cytotoxic activity of the isolated compounds was tested on three human leukemia cell lines, HL-60 (acute promyelocytic leukemia cells), U-937 (acute monocytic leukemia cells), and K-562 (human chronic myelogenous leukemia cells), as well as on human embryonal teratocarcinoma cell line (NT2/D1) and human glioblastoma cell lines A-172 and U-251, and compared to the previously isolated bis-bibenzyls (perrottetins) of similar structure. The isolated compounds exhibited modest activity against leukemia cells and significant activity against NT2/D1 and A-172. Overall, the most active cytotoxic compounds in this investigation were perrottetin E (1), isolated in this work from Pellia endiviifolia, and perrottetin F phenanthrene derivative (7), previously isolated from Lunularia cruciata and added for a comparison of their cytotoxic activity.

Cytotoxic Activity of Riccardin and Perrottetin Derivatives from the Liverwort Lunularia cruciata.

Seven new bisbibenzyls (1-7) were isolated from the methanol extract of the liverwort Lunularia cruciata along with one previously known bibenzyl and five known bisbibenzyls. The structures of compounds 1-7 were elucidated on the basis of the spectroscopic data. These newly isolated bisbibenzyls may be divided into two groups, the acyclic bisbibenzyls, perrottetins (1-3), and the cyclic analogues, riccardins (4-7). Besides standard perrottetin and riccardin structures (1 and 4, respectively), they contain phenanthrene (3 and 5), dihydrophenanthrene (2), and quinone moieties (6 and 7), rarely found in natural products. The new compounds 3 and 5, as well as the known riccardin G, exhibited cytotoxic activity against the A549 lung cancer cell line with IC50 values of 5.0, 5.0, and 2.5 µM, respectively.

Diarylheptanoids from Alnus viridis ssp. viridis and Alnus glutinosa: Modulation of Quorum Sensing Activity in Pseudomonas aeruginosa.

Diarylheptanoids from the barks of Alnus viridis ssp. viridis (green alder) and Alnus glutinosa (black alder) were explored for anti-quorum sensing activity. Chemicals with anti-quorum sensing activity have recently been examined for antimicrobial applications. The anti-quorum sensing activity of the selected diarylheptanoids was determined using two biosensors, namely Pseudomonas aeruginosa PAO1 and Chromobacterium violaceum CV026. Although all of the investigated compounds negatively influenced the motility of P. aeruginosa PAO1, four were able to inhibit biofilm formation of this human opportunistic pathogen for 40-70 %. Three of the diarylheptanoids (3, 4, and 5) negatively influenced the biosynthesis of pyocyanin, which is under the control of quorum sensing. Platyphyllenone (7) and hirsutenone (5) were able to inhibit the biosynthesis of violacein in C. violaceum CV026, with 5 being able to inhibit the synthesis of both biopigments. Only one of the tested diarylheptanoids (1) was shown to significantly decrease the production of acyl homoserine lactones (AHL) in P. aeruginosa PAO1, more specifically, production of the long chain N-(3-oxododecanoyl)-l-HSL. On the other side, four diarylheptanoids (2-5) significantly reduced the synthesis of 2-alkyl-4-quinolones, part of the P. aeruginosa quinolone-mediated signaling system. To properly assess therapeutic potential of these compounds, their in vitro antiproliferative effect on normal human lung fibroblasts was determined, with doses affecting cell proliferation between 10 and 100 µg/mL. This study confirms that the barks of green and black alders are rich source of phytochemicals with a wide range of biological activities that could further be exploited as natural agents against bacterial contaminations and infections.

Diarylheptanoids from green alder bark and their potential for DNA protection.

Nine diarylheptanoids, 1-9, catechin (11), and a phenolic glucoside, 10, were isolated from the bark of green alder (Alnus viridis). Four of the isolated compounds, i.e., 2, 5, 8, 10, are new. The structures of 1-11 were determined on the basis of spectroscopic data. All isolated compounds were evaluated for their in vitro protective effects on chromosome aberrations in peripheral human lymphocytes using cytokinesis-block micronucleus (CBMN) assay. Almost all of them exerted a pronounced effect of decreasing DNA damage of human lymphocytes, acting stronger than the known synthetic protector amifostine.

Satureja horvatii essential oil: in vitro antimicrobial and antiradical properties and in situ control of Listeria monocytogenes in pork meat.

The dominant compounds in Satureja horvatii oil were p-cymene (33.14%), thymol (26.11%) and thymol methyl ether (15.08%). The minimum inhibitory concentration (MIC) varied from 0.03 to 0.57 mg/mL for bacteria, and from 0.56 to 2.23 mg/mL for yeast strains, while minimum bactericidal/yeast-cidal concentration (MBC/MYC) varied from 0.07 to 1.15 mg/mL and 1.11 to 5.57 mg/mL for bacteria and yeasts, respectively. The antiradical potential of the essential oil was evaluated using hydroxyl radical (•OH) generated in Fenton reaction. The meat preserving potential of essential oil from Satureja horvatii was investigated against L. monocytogenes. Essential oil successfully inhibited development of L. monocytogenes in pork meat. Sensorial evaluation on flavor and color of meat was performed. The color and flavor of meat treated with essential oil improved after 4 days of storage. S. horvatii essential oil can act as a potent inhibitor of food spoiling microorganisms, in meat products and also can be a useful source of natural antioxidants.

Assessment of the chemical composition and in vitro antimicrobial potential of extracts of the liverwort Scapania aspera.

The chemical composition of Scapania aspera extracts was determined by solid phase micro extraction gas chromatography-mass spectrometry (SPME GC-MS) and 96 constituents were identified. The dominant compounds in the methanol extract were beta-barbatene (25.1%), o-cymene (14.0%), alpha-barbatene (5.7%), allo-aromadendrene (4.9%) and beta-bourbonene, while in the ethanol extract, o-cymene (17.8%), beta-barbatene (17.6%), alpha-thujene (6.7%), octen-1-ol acetate (4.9%) and beta-bazzanene (2.4%) were the major components. In the ethyl acetate extract, beta-barbatene (14.3%), undecane (11.8%), 2-methyldecane (11.2%), decane (10.9%) and o-cymene (3.6%) were major components. The antimicrobial activity of the different extracts was evaluated against pathogenic and food spoilage microorganisms using disc diffusion and micro-broth dilution methods. The minimal inhibitory concentration (MIC) of extracts of S. aspera varied from 0.4 to 1.5 mg/mL and 1 to 3 mg/mL for yeast and bacterial strains, respectively. The zone of inhibition of the methanol extract for yeast strains was higher than that for bacterial strains. The results suggest that S. aspera extracts have potential as natural antimicrobial agents.

Antimicrobial Potential and Chemical Characterization of Serbian Liverwort (Porella arboris-vitae): SEM and TEM Observations.

The chemical composition of Porella arboris-vitae extracts was determined by solid phase microextraction, gas chromatography-mass spectrometry (SPME GC-MS), and 66 constituents were identified. The dominant compounds in methanol extract of P. arboris-vitae were ß-caryophyllene (14.7%), α-gurjunene (10.9%), α-selinene (10.8%), ß-elemene (5.6%), γ-muurolene (4.6%), and allo-aromadendrene (4.3%) and in ethanol extract, ß-caryophyllene (11.8%), α-selinene (9.6%), α-gurjunene (9.4%), isopentyl alcohol (8.8%), 2-hexanol (3.7%), ß-elemene (3.7%), allo-aromadendrene (3.7%), and γ-muurolene (3.3%) were the major components. In ethyl acetate extract of P. arboris-vitae, undecane (11.3%), ß-caryophyllene (8.4%), dodecane (6.4%), α-gurjunene (6%), 2-methyldecane (5.1%), hemimellitene (4.9%), and D-limonene (3.9%) were major components. The antimicrobial activity of different P. arboris-vitae extracts was evaluated against selected food spoilage microorganisms using microbroth dilution method. The Minimal Inhibitory Concentration (MIC) varied from 0.5 to 1.5 mg/mL and 1.25 to 2 mg/mL for yeast and bacterial strains, respectively. Significant morphological and ultrastructural alterations due to the effect of methanolic and ethanolic P. arboris-vitae extracts on S. Enteritidis have also been observed by scanning electron microscope and transmission electron microscope, respectively. The results provide the evidence of antimicrobial potential of P. arboris-vitae extracts and suggest its potential as natural antimicrobial agents for food preservation.

Composition of n-alkanes in natural populations of Pinus nigra from Serbia - chemotaxonomic implications.

This is the first report on the composition and variability of the needle-wax n-alkanes in natural populations of Pinus nigra in Serbia. Samples of 195 trees from seven populations belonging to several infraspecific taxa (ssp. nigra, var. gocensis, ssp. pallasiana, and var. banatica) were analyzed. In general, the size of the n-alkanes ranged from C(16) to C(33) , with the exception of ssp. nigra, for which it ranged from C(18) to C(33) . The most abundant were C(23) -, C(25) -, C(27) -, and C(29) -alkanes. The needle waxes of Populations I-III and V were characterized by a higher content of C(23) -, C(25) -, and C(27) -alkanes and a lower content of C(24) -, C(26) -, C(28) -, and C(30) -alkanes, compared to the other populations, and the trees of these populations could be assigned to ssp. nigra. The samples of Population VI were characterized by higher amounts of C(22) -, C(24) -, C(30) -, and C(32) -alkanes and lower amounts of C(25) - and C(27) -alkanes, and the trees could be considered as ssp. pallasiana. The samples of Population VII, consisting of trees belonging to var. banatica, were richer in C(29) -, C(31) -, and C(33) -alkanes. The wax compositions of Populations IV and V, both composed of trees previously determined as P. nigra var. gocensis, showed a tendency of splitting. Indeed, the alkane composition of Population IV was closer to that of ssp. pallasiana pines, while that of Population V was more similar to that of ssp. nigra pines. From the results presented here, it is obvious that in the central part of the Balkan Peninsula, significant diversification and differentiation of the populations of black pine exists, and these populations could be defined as different intraspecific taxa. Our results also indicate the validity of n-alkanes as chemotaxonomic characters within this aggregate.

Identification of volatile components of liverwort (Porella cordaeana) extracts using GC/MS-SPME and their antimicrobial activity.

Chemical constituents of liverwort (Porella cordaeana) extracts have been identified using solid-phase microextraction-gas chromatography mass spectrometry (SPME-GC/MS). The methanol, ethanol and ethyl acetate extracts were rich in terpenoids such as sesquiterpene hydrocarbons (53.12%, 51.68%, 23.16%), and monoterpene hydrocarbons (22.83%, 18.90%, 23.36%), respectively. The dominant compounds in the extracts were ß-phellandrene (15.54%, 13.66%, 12.10%) and ß-caryophyllene (10.72%, 8.29%, 7.79%, respectively). The antimicrobial activity of the extracts was evaluated against eleven food microorganisms using the microdilution and disc diffusion methods. The minimum inhibitory concentration (MIC) varied from 0.50 to 2.00 mg/mL for yeast strains (Saccharomyces cerevisiae 635, Zygosacharomyces bailii 45, Aerobasidium pullulans L6F, Pichia membranaefaciens OC 71, Pichia membranaefaciens OC 70, Pichia anomala CBS 5759, Pichia anomala DBVPG 3003 and Yarrowia lipolytica RO13), and from 1.00 to 3.00 mg/mL for bacterial strains (Salmonella enteritidis 155, Escherichia coli 555 and Listeria monocytogenes 56Ly). Methanol extract showed better activity in comparison with ethanol and ethyl acetate extracts. High percentages of monoterpene and sesquiterpene hydrocarbons could be responsible for the better antimicrobial activity.

Chemical and principal-component analyses of the essential oils of Apioideae taxa (Apiaceae) from central Balkan.

The volatile constituents of the essential oils of 23 taxa belonging to the Apioideae subfamily were studied in detail. The investigated taxa were Pimpinella serbica (Vis.) Bentham & Hooker, Libanotis montana Cr., Cnidium silaifolium (Jacq.) Simk. ssp. orientale (Boiss.) Tutin, Bupleurum praealtum L., B. sibthorpianum S. S. var. diversifolium (Roch.) Hay, Aegopodium podagraria L., Torilis anthriscus (L.) Gmel., Orlaya grandiflora (L.) Hoffm., Laserpitium siler L., Laser trilobum (L.) Brokh., Chaerophyllum aureum L., C. hirsutum L., C. temulum L., Pastinaca sativa L., P. hirsuta Pancic., Tordylium maximum L., Physospermum cornubiense (L.) DC., Peucedanum alsaticum L., P. oreoselinum (L.) Moench, P. cervaria (L.) Cuss., P. austriacum (Jacq.) Koch, P. longifolium W. et K., and P. officinale L. All of these species grow wild in the central part of the Balkan Peninsula. The essential oils were found to be complex mixtures of various compounds, more than 100 constituents being in each taxon, with contributions of main products never exceeding 25% of the total content. Sesquiterpene hydrocarbons were found to be the main group of constituents of all taxa, except for Peucedanum species, where monoterpene hydrocarbons were identified as the main components. The chemotaxonomic value of the essential-oil composition is discussed according to results of principal-component analysis (PCA). The essential-oil composition mainly reflects current taxonomic relationships between the investigated taxa.