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Plants of the genus Prangos are intensively investigated as potential new sources of bioactive isolated products. In this work, the chemical composition of volatile constituents (essential oils and headspace volatiles) and dichloromethane extracts, as well as antimicrobial and antibiofilm activities of essential oils and MFDEs (methanol fractions of dichloromethane extracts) of Prangos trifida from Serbia, were investigated. Volatiles of roots, leaves, stems and fruits, and fatty acids and phytosterols in dichloromethane extracts of roots and fruits were analyzed by GC-FID-MS, whereas coumarins in MFDEs by LC-MS and some isolated coumarins by 1 H-NMR. Minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations/minimum fungicidal concentrations (MBCs/MFCs) of essential oils and MFDEs were determined against 13 microorganisms. Antibiofilm activity was assessed against four microorganisms. Additionally, congo red and ergosterol binding assays were conducted to elucidate selected mechanisms of antibiofilm action in the case of Candida albicans . Total of 52 volatile constituents, 16 fatty acids, eight phytosterols and 10 coumarins were identified. Essential oils demonstrated significant activity, surpassing that of commercial food preservatives, against six tested molds from the Aspergillus , Penicillium and Trichoderma genera, as well as against bacteria Staphylococcus aureus and Bacillus cereus . Most of the oils strongly inhibited the formation of biofilms by S. aureus, Listeria monocytogenes and Escherichia coli . MFDEs exhibited noteworthy effects against B. cereus and the tested Aspergillus species, particularly A. niger , and significantly inhibited C. albicans biofilm formation. This inhibition was linked to a marked reduction in exopolysaccharide production, while antifungal mechanisms associated with ergosterol remained unaffected.

The composition and anticholinesterase activity of the dried MeOH extracts of Hieracium scheppigianum and H. naegelianum underground parts (rhizomes and roots), as well as the anticholinesterase activity of the dried, previously chemically characterised MeOH extracts of the flowering aerial parts of these two and 26 other Hieracium species in the strict sense (s. str.), were investigated. Furthermore, the anticholinesterase activity of 12 selected secondary metabolites of these extracts was evaluated. Using semi-preparative LC-MS, five caffeoylquinic acids and the sesquiterpene lactone crepiside E were isolated from H. scheppigianum underground parts extract. All these compounds were also identified in the underground parts extract of H. naegelianum. Quantitative LC-MS analysis showed that the analysed underground parts extracts were rich in both caffeoylquinic acids (139.77 and 156.62 mg/g of extract, respectively) and crepiside E (126.88 and 116.58 mg/g). In the Ellman method, the tested extracts showed an interesting anti-AChE and/or anti-BChE activity (IC 50 =0.56-1.58 mg/mL), which can be explained, at least partially, by the presence of some of their constituents. Among the metabolites tested, the best activity was revealed for the flavonoids apigenin, luteolin and diosmetin, and the sesquiterpene lactone 8-epiixerisamine A (IC 50 =68.09-299.37 μM)., (© 2023 Wiley-VHCA AG, Zurich, Switzerland.)

Alchemilla viridiflora Rothm., Rosaceae is a herbaceous plant widespread in central Greece, Bulgaria, North Macedonia and Serbia with Kosovo. Liquid chromatography-mass spectrometry analysis leads to the identification of 20 compounds in methanol extract, mainly ellagitannins and flavonoid glycosides. Given that various plant extracts have traditionally been used to treat hypertension and that some of the analyzed methanol extract constituents have beneficial cardiovascular effects, we hypothesized that some of these effects are achieved by inhibiting angiotensin I-converting enzyme (ACE). The dose-dependent ACE inhibitory activities of A. viridiflora and miquelianin were observed with an IC 50 of 2.51 ± 0.00 µg/mL of A. viridiflora extract compared to the IC 50 of 5.4139 ± 0.00 µM for miquelianin. The contribution of the single compounds to the tested activity was further analyzed through the in silico experimental approach. Computational docking results showed that tiliroside, ellagic acid pentose and galloyl-hexahydroxydiphenoyl-glucose exhibited even better binding affinity for the ACE active site than miquelianin, for which ACE activity was confirmed by an in vitro assay., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Dry MeOH extract of Ferula heuffelii (Apiaceae) underground parts was tested for spasmolytic, gastroprotective and antioxidant activities. HPLC analysis revealed that chlorogenic acid (CGA; 34.6 mg/g) was its main constituent. Extract in vitro exhibited notable total antioxidant activity (FRAP value=1.0 μmol Fe 2+ /mg), and scavenging of DPPH (SC 50 =62.5 μg/ml) and • OH radicals (49.5 % at 20 μg/ml in 2-deoxyribose assay). In vitro on isolated rat ileum, extract exhibited significant spasmolytic activity, i. e., it showed 124.6 % of maximal atropine effect on spontaneous contractions (at 100 μg/ml), and reduced spasmogenic effect of KCl (80 mm) to 44.4 % (at 60 μg/ml) and of highest applied concentration of ACh to 26.3 % (at 120 μg/ml). In parallel experiments, spasmolytic effect of CGA was also demonstrated. In acute EtOH-induced gastric ulceration model in rats, extract (100 mg/kg p.o.) showed significant gastroprotective effect (gastric damage score 0.50), similar to ranitidine (20 mg/kg p.o.). Obtained results showed that tested F. heuffelii polar extract represents new herbal preparation with potential use against some gastrointestinal complaints., (© 2022 Wiley-VHCA AG, Zurich, Switzerland.)

The genus Ramonda includes three Paleoendemic and Tertiary relict species that survived in refugial habitats of the Balkan Peninsula ( R. nathaliae and R. serbica ) and the Iberian Peninsula ( R. myconi ). They are all "resurrection plants," a rare phenomenon among flowering plants in Europe. Ramonda myconi and R. nathaliae are diploids (2 n  = 2 x  = 48), while R. serbica is a hexaploid (2 n  = 6 x  = 144). The two Balkan species occur in sympatry in only two localities in eastern Serbia, where tetraploid potential hybrids (2 n  = 4 x  = 96) were found. This observation raised questions about the existence of gene flow between the two species and, more generally, about the evolutionary processes shaping their genetic diversity. To address this question, genetic markers (AFLP) and an estimate of genome size variation were used in a much larger sample and at a larger geographic scale than previously. The combination of AFLP markers and genome size results suggested ongoing processes of interspecific and interploidy hybridization in the two sites of sympatry. The data also showed that interspecific gene flow was strictly confined to sympatry. Elsewhere, both Ramonda species were characterized by low genetic diversity within populations and high population differentiation. This is consistent with the fact that the two species are highly fragmented into small and isolated populations, likely a consequence of their postglacial history. Within sympatry, enormous variability in cytotypes was observed, exceeding most reported cases of mixed ploidy in complex plant species (from 2 x to >8 x ). The AFLP profiles of non-canonical ploidy levels indicated a diversity of origin pathways and that backcrosses probably occur between tetraploid interspecific hybrids and parental species. The question arises whether this diversity of cytotypes corresponds to a transient situation. If not, the question arises as to the genetic and ecological mechanisms that allow this diversity to be maintained over time., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Lazarević, Siljak-Yakovlev, Sanino, Niketić, Lamy, Hinsinger, Tomović, Stevanović, Stevanović and Robert.)

Present study investigated triterpene profile, antihyperalgesic and antiedematous activities of Hieracium scheppigianum flowering aerial parts dichloromethane extract (SCH), and antihyperalgesic and antiedematous activities of previously chemically characterised polyphenol-rich H. glabratum and H . calophyllum flowering aerial parts methanol extracts (GLA and CAL, respectively). α - and β -Amyrin and their acetates, and lupeol acetate were identified and quantified in SCH by GC-FID and GC-MS. In carrageenan-induced localised inflammation model in rats, SCH and GLA (50-200 mg/kg, p.o.) produced significant and dose-dependent antihyperalgesic effect of 26.9%-56.2% (ED 50 =163.0 ± 26.5 mg/kg) and 25.3%-51.6% (ED 50 =211.6 ± 70.6 mg/kg), respectively, and CAL (200 mg/kg, p.o.) exhibited effect of 38.1%. Extracts did not significantly reduce paw edema. SCH and GLA, which demonstrated higher (over 50%) antihyperalgesic efficacy, were tested in a rotarod test (200 mg/kg, p.o.) and no alteration of motor coordination was observed. Also, acute administration of SCH and GLA in mice (2000 mg/kg, p.o.) caused neither mortality nor toxicity.

Four sesquiterpene lactones (SLs), including three undescribed proline-SL conjugates, the guaianolides calophyllamine A and 8-epiixerisamine A, and the eudesmanolide calophyllamine B, were isolated from the methanol extract of Hieracium calophyllum R. Uechtr. (Compositae) flowering heads. Another known guaianolide, crepiside E, was detected in Hieracium L. species for the first time. Their structures were elucidated using extensive 1D and 2D NMR spectroscopy in combination with HRMS. The isolated SLs were used as external standards for qualitative and quantitative LC-MS analysis of the dry methanol extracts of the flowering aerial parts of 28 Hieracium species from the Balkan Peninsula. Guaianolides were the dominant SLs in 27 species studied. The chemosystematic significance of detected SLs was evaluated using multivariate statistics (PCA, nMDS and UPGMA). Differentiation between the main groups was well supported. All four compounds significantly and equally contributed to the differences between the species. In addition, the eudesmanolide calophyllamine B could be a significant chemosystematic marker for H. sect. Villosa (Griseb.) Gremli s.l. and Glauciformia (Freyn) Zahn-Italica (Fr.) Av. Touv., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Introduction: Hieracium s. str. represents one of the largest and most complex genera of flowering plants. As molecular genetics seems unlikely to disentangle intricate relationships within this reticulate species complex, analysis of flavonoids and phenolic acids, known as good chemosystematic markers, promise to be more reliable. Data about pharmacological activity of Hieracium species are scarce., Objective: Evaluation of the chemosystematic significance of flavonoids and phenolic acids of methanol extracts of aerial flowering parts of 28 Hieracium species from the Balkans. Additionally, investigation of antioxidant potentials of the extracts., Methods: Comparative qualitative and quantitative analysis of flavonoids and phenolic acids was performed by LC-MS. Multivariate statistical data analysis included non-metric multidimensional scaling (nMDS), unweighted pair-group arithmetic averages (UPGMA) and principal component analysis (PCA). Antioxidant activity was evaluated using three colorimetric tests., Results: Dominant phenolics in almost all species were luteolin type flavonoids, followed by phenolic acids. Although the investigated Hieracium species share many compounds, the current classification of the genus was supported by nMDS and UPGMA analyses with a good resolution to the group level. Hieracium naegelianum was clearly separated from the other investigated species. Spatial and ecological distances of the samples were likely to influence unexpected differentiation of some groups within H. sect. Pannosa. The vast majority of dominant compounds significantly contributed to differences between taxa. The antioxidant potential of the extracts was satisfactory and in accordance with their phenolics composition., Conclusions: Comparative LC-MS analysis demonstrated that flavonoids and phenolic acids are good indicators of chemosystematic relationships within Hieracium, particularly between non-hybrid species and groups from the same location. Copyright © 2017 John Wiley & Sons, Ltd., (Copyright © 2017 John Wiley & Sons, Ltd.)

Many Heracleum L. taxa (Apiaceae) are used as food and spices, and in traditional medicine. In this work, the chemical composition of Heracleum pyrenaicum subsp. orsinii (Guss.) F. Pedrotti and Pignatti root, leaf and fruit essential oils, their antimicrobial activity and cytotoxic effect on malignant and normal cells were investigated for the first time. The composition of the oils was analyzed by GC and GC-MS. Monoterpenes prevailed in the root oil, with β -pinene (38.6%) being dominant, while in the leaf oil, sesquiterpenes, mostly ( E )-nerolidol (20.5%) and ( E )-caryophyllene (17.0%), were the most abundant constituents. The fruit oil contained the majority of aliphatic esters, mainly octyl acetate (36.8%) and octyl hexanoate (22.1%). The antimicrobial activity was determined by microdilution method against eight bacteria and eight fungi (standard strains, clinical or food isolates). The best antibacterial activity, better than the activity of ampicillin, was shown by the root oil against Salmonella typhimurium , Escherichia coli and Pseudomonas aeruginosa . The strongest antifungal activity, stronger than the activity of ketoconazole, was exhibited by the leaf and root oils against Trichoderma viride , and by the root oil against Aspergillus ochraceus . The cytotoxic effect of the oils, determined by MTT test, was prominent against malignant HeLa, LS174 and A549 cells (IC 50  = 6.49-14.56 μg/mL). On the other hand, the oils did not show toxicity against normal MRC-5 cells at tested concentrations (IC 50  > 200.00 μg/mL). It can be concluded that investigated H . pyrenaicum subsp. orsinii oils represent potential new raw materials for food and pharmaceutical industry.

The compositions of hydrodistillated essential oils of Laser trilobum (L.) Borkh. rhizomes and fruits from Serbia, were investigated using GC and GC/MS. In the dark-blue rhizome oil forty-six compounds (93.1% of the total oil) were identified, with a-pinene (31.5%), y-terpinene (9.0%), p-cymene (7.9%), β-pinene (6.1%) and 1,4-dimethylazalene (6.0%) as the major components. In the colorless fruits oil, twenty components (96.8% of the total oil) were identified, and the main constituents were limonene (51.6%) and perillaldehyde (26.8%). The antimidrobial activity of the oils was tested using the broth microdilution method against nine bacterial and two fungal strains. The oils revealed significant antimicrobial effect, mainly better than that of thymol, used as a reference compound. The strongest activity was recorded for the rhizome oil against Escherichia coli, Klebsiella pneumoniae and Candida albicans (MICs=25 μg/mL), and the fruit oil against C. albicans ATCC 10259 (MIC=12.5 μg/mL).

Although some widespread, native cow parsnips (Heracleum L. spp., Apiaceae) had broad medicinal and culinary applications throughout history, the knowledge about their volatile constituents is insufficient. This work investigates the composition and bioactivities of H. sphondylium L. (HSPH), H. sibiricum L. (HSIB) and H. montanum Schleich. ex Gaudin (HMON) essential oils. The composition was tested by GC and GC-MS. (Z)-β-Ocimene was the most abundant in HSPH (28.9%) and HMON (20.4%) root oils, while in HSIB root oil, β-pinene (26.2%), methyl eugenol (22.3%) and elemicin (25.6%) prevailed. Leaf and flower oils were dominated by various sesquiterpenes (germacrene D, β-sesquiphellandrene, (E)-β-farnesene and/or (E)-caryophyllene) and/or phenylpropanoids (apiole, methyl eugenol, elemicin and/or (Z)-isoelemicin). Octyl acetate (57.5-67.1%) was the main constituent of all fruit oils. The antimicrobial activity was screened by a microdilution method against eight bacteria and eight fungi. The strongest antimicrobial effect, in several cases better than the activity of antibiotics, was shown by HSPH (MICs = 0.12-3.30 mg mL -1 ) and HMON (MICs = 0.10-1.30 mg mL -1 ) flower oils against bacteria, and HSIB fruit oil against fungi (MICs = 0.15-0.40 mg mL -1 ). The MTT test revealed that the oils were not or weakly cytotoxic against human malignant HeLa, LS174 and/or A549 cells (except HSPH root oil; IC 50 = 5.72-24.31 μg mL -1 ) and that tested oils were not toxic against human normal MRC-5 cells (at 200.00 μg mL -1 ). Significant activity observed against microorganisms that are the common cause of foodborne diseases, food contamination and/or hospital-acquired infections justifies certain traditional uses of the investigated plants and represents a good basis for further research of these Heracleum oils.

Background: Polyploidy is one of the most important evolutionary pathways in flowering plants and has significantly contributed to their diversification and radiation. Due to the prevalence of reticulate evolution spanning three ploidy levels, Knautia is considered one of the taxonomically most intricate groups in the European flora. On the basis of ITS and plastid DNA sequences as well as AFLP fingerprints obtained from 381 populations of almost all species of the genus we asked the following questions. (1) Where and when did the initial diversification in Knautia take place, and how did it proceed further? (2) Did Knautia undergo a similarly recent (Pliocene/Pleistocene) rapid radiation as other genera with similar ecology and overlapping distribution? (3) Did polyploids evolve within the previously recognised diploid groups or rather from hybridisation between groups?, Results: The diversification of Knautia was centred in the Eastern Mediterranean. According to our genetic data, the genus originated in the Early Miocene and started to diversify in the Middle Miocene, whereas the onset of radiation of sect. Trichera was in central parts of the Balkan Peninsula, roughly 4 Ma. Extensive spread out of the Balkans started in the Pleistocene about 1.5 Ma. Diversification of sect. Trichera was strongly fostered by polyploidisation, which occurred independently many times. Tetraploids are observed in almost all evolutionary lineages whereas hexaploids are rarer and restricted to a few phylogenetic groups. Whether polyploids originated via autopolyploidy or allopolyploidy is unclear due to the weak genetic separation among species. In spite of the complexity of sect. Trichera, we present nine AFLP-characterised informal species groups, which coincide only partly with former traditional groups., Conclusions: Knautia sect. Trichera is a prime example for rapid diversification, mostly taking place during Pliocene and Pleistocene. Numerous cycles of habitat fragmentation and subsequent reconnections likely promoted hybridisation and polyploidisation. Extensive haplotype sharing and unresolved phylogenetic relationships suggest that these processes occurred rapidly and extensively. Thus, the dynamic polyploid evolution, the lack of crossing barriers within ploidy levels supported by conserved floral morphology, the highly variable leaf morphology and unstable indumentum composition prevent establishing a well-founded taxonomic framework.

The objective of this research was to analyze the chemical composition, antimicrobial and cytotoxic activity of Heracleun pyrenaicum subsp. pollinianum (Bertol.) F. Pedrotti & Pignatti (HPP) and H. orphanidis Boiss. (HO) essential oils. The composition of the oils was analyzed by GC and GC-MS. β-Pinene (35.1%) was the most abundant compound in HPP root oil, while (Z)-falcarinol (80.0%) dominated in HO root oil. (E)-Nerolidol (28.5%) was the main constituent in HPP leaf oil. HPP fruit oil, as well as HO leaf and fruit oils mainly contained aliphatic esters, mostly octyl acetate (50.5-84.5%). Antimicrobial screening was performed by microdilution method against eight bacterial and eight fungal strains. The strongest antibacterial activity was shown by both root oils (MICs 0.02-0.60 mg/mL and MBCs 0.04-2.50 mg/mL for HPP, and MICs 0.02-1.25 mg/mL and MBCs 0.04-2.50 mg/mL for HO), while the best antifungal potential was exhibited by HPP fruit oil (MICs 0.30-0.60 mg/mL and MFCs 0.60-1.25 mg/mL) and HO leaf oil (MICs 0.15-0.63 mg/mL and MFCs 0.30-1.25 mg/mL). The tested root and fruit oils exhibited strong cytotoxic effect, which was determined by MTT test against HeLa (IC50 7.53-21.07 µg/mL) and LS174 (IC50 24.16-58.86 µg/mL) cell lines.

The antimicrobial and cytotoxic activities of isolates (CHCl3 and MeOH extracts and selected metabolites) obtained from the underground parts of the Balkan endemic plant Ferula heuffelii Griseb. ex Heuff. were assessed. The CHCl3 and MeOH extracts exhibited moderate antimicrobial activity, being more pronounced against Gram-positive than Gram-negative bacteria, especially against Staphylococcus aureus (MIC=12.5 μg/ml for both extracts) and Micrococcus luteus (MIC=50 and 12.5 μg/ml, resp.). Among the tested metabolites, (6E)-1-(2,4-dihydroxyphenyl)-3,7,11-trimethyl-3-vinyldodeca-6,10-dien-1-one (2) and (2S*,3R*)-2-[(3E)-4,8-dimethylnona-3,7-dien-1-yl]-2,3-dihydro-7-hydroxy-2,3-dimethylfuro[3,2-c]coumarin (4) demonstrated the best antimicrobial activity. Compounds 2 and 4 both strongly inhibited the growth of M. luteus (MIC=11.2 and 5.2 μM, resp.) and Staphylococcus epidermidis (MIC=22.5 and 10.5 μM, resp.) and compound 2 additionally also the growth of Bacillus subtilis (MIC=11.2 μM). The cytotoxic activity of the isolates was tested against three human cancer cell lines, viz., cervical adenocarcinoma (HeLa), chronic myelogenous leukemia (K562), and breast cancer (MCF-7) cells. The CHCl3 extract exhibited strong cytotoxic activity against all cell lines (IC50 <11.0 μg/ml). All compounds strongly inhibited the growth of the K562 and HeLa cell lines. Compound 4 exhibited also a strong activity against the MCF-7 cell line, comparable to that of cisplatin (IC50 =22.32±1.32 vs. 18.67±0.75μM)., (Copyright © 2015 Verlag Helvetica Chimica Acta AG, Zürich.)

The present study evaluated the anti-Helicobacterpylori activity of Alchemilla glabra Neygenf. (A. sect. Alchemilla), A. monticola Opiz (A. sect. Plicatae S.E. Fröhner), A. fissa Günther & Schummel (A. sect. Calycinae (Buser) Buser) and A. viridiflora Rothm. (A. sect. Calycinae), and identified ellagic acid and quercetin-3-O-β-glucoside. Anti-H. pylori activity was tested against ten clinical isolates and one reference strain (ATCC 43504). The methanol extracts were more active than the dichloromethane and cyclohexane extracts. The ranges of concentrations were between 4 μg/mL for methanol extracts of A. viridiflora, A. glabra and A. monticola, and 256 μg/mL for cyclohexane extracts of A. viridiflora, A. glabra and A. fissa. The best overall activity was obtained with A. monticola extracts. No significant difference was found in the ellagic acid contents of the methanol extracts of the tested Alchemilla species (0.2-0.3 mg/mL), and anti-H. pylori activity was similar (4-32 μg/mL). Ellagic acid exhibited strong activity at very low concentrations (0.125-0.5 μg/mL), while the second identified compound, quercetin-3-O-β-D-glucoside, was also very active in concentration of 2-16 μg/mL.

Antimicrobial properties of extracts of underground parts of three Laserpitium L. (Apiaceae) species, namely Laserpitium latifolium L., Laserpitium zernyi Hayek and Laserpitium ochridanum Micevski, were investigated. The investigated species are widely used as functional foods, as spices and for preparations in traditional medicine for treating complaints connected with infection and inflammation. Furthermore, antimicrobial and antibiofilm effects of laserpitine, the most abundant compound in the chloroform extract of L. latifolium, and guaianolide sesquiterpene lactones, such as, isomontanolide, montanolide and tarolide, principal components of the extracts of L. zernyi and L. ochridanum were assessed. The antimicrobial activity was tested using the microdilution method against five pathogenic bacteria and five fungi, as well as in the microplate biofilm assay on two Candida clinical isolates (C. albicans and C. krusei). Among the extracts, L. latifolium showed the most prominent activity. Isolated metabolites exerted higher effects against fungal than against bacterial strains, isomontanolide being the most active. Interestingly, all constituents showed higher potential on inhibition of biofilm formation than fluconazole, a reference compound. Tested metabolites may be good novel agents with high antifungal and antibacterial potential that might find practical applications in food industry as food preservatives in order to retard the growth of food spoiling microbes, but only after detailed safety assessments.

The Balkans are a major European biodiversity hotspot, however, almost nothing is known about processes of intraspecific diversification of the region's high-altitude biota and their reaction to the predicted global warming. To fill this gap, genome size measurements, AFLP fingerprints, plastid and nuclear sequences were employed to explore the phylogeography of Cerastium dinaricum. Range size changes under future climatic conditions were predicted by niche-based modeling. Likely the most cold-adapted plant endemic to the Dinaric Mountains in the western Balkan Peninsula, the species has conservation priority in the European Union as its highly fragmented distribution range includes only few small populations. A deep phylogeographic split paralleled by divergent genome size separates the populations into two vicariant groups. Substructure is pronounced within the southeastern group, corresponding to the area's higher geographic complexity. Cerastium dinaricum likely responded to past climatic oscillations with altitudinal range shifts, which, coupled with high topographic complexity of the region and warmer climate in the Holocene, sculptured its present fragmented distribution. Field observations revealed that the species is rarer than previously assumed and, as shown by modeling, severely endangered by global warming as viable habitat was predicted to be reduced by more than 70% by the year 2080., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Plants from the genus Ferula L. (Apiaceae) were used for various purposes in traditional medicine of different nations throughout the history. Ferula heuffelii Griseb. ex Heuffel is a perennial species endemic for Balkan peninsula. Ten compounds which belong to classes of prenyl-furocoumarin-, prenyl-dihydrofurochromone-, prenyl-benzoyl- and prenyl-benzoylfuranone-type sesquiterpenoids, as well as sesquiterpene coumarins and phenylpropanoids, were, for the first time, isolated from the CHCl3 extract of the underground parts of this plant and identified. Furthermore, extract and three isolated compounds, i.e., latifolone (1), dshamirone (4), and (2S*,3R*)-2-[(3E)-4,8-dimethylnona-3,7-dien-1-yl]-2,3-dihydro-7-hydroxy-2,3-dimethylfuro[3,2-c]coumarin (6) were, for the first time, evaluated for their in vitro antispasmodic activities in three experimental models: spontaneous contraction, and ACh- and KCl-induced contraction of an isolated rat ileum. The extract (0.1-1.3 mg/ml) and compound 6 (1-10 μg/ml) exhibited dose-dependent effect in all three models. Compound 1 (1-6 μg/ml) affected spontaneous contractions and those induced by KCl, while compound 4 (8 μg/ml) displayed only moderate activity with ACh-induced contractions. It can be concluded that tested compounds contribute to exhibited antispasmodic activity of crude extract. Additionally, extract (0.1-1.3 mg/ml) was tested for in vitro relaxant activity on an isolated rat trachea, and relaxed the KCl-induced contractions in a dose-dependent manner., (Copyright © 2014 Verlag Helvetica Chimica Acta AG, Zürich.)

In this paper antinociceptive and anti-edematous effects are examined of the essential oils of the underground parts of two Balkan endemic Laserpitium species (Apiaceae), L. zernyi and L. ochridanum. Furthermore, the essential oil of the underground parts of L. ochridanum is chemically characterised by GC and GC-MS. Antinociceptive and anti-edematous effects were measured in a rat model of localized inflammation, induced by carrageenan, using apparatus for the modified paw-pressure test, and plethysmometer, respectively. The effects of both Laserpitium essential oils were measured after oral gavage administration to male Wistar rats in doses of 25, 50 and 100 mg/kg. The main constituents of L. ochridanum essential oil were: alpha-pinene (33.2%), alpha-bisabolol (10.3%) and chamazulene (14.9%). The essential oil of L. zernyi was previously shown to be rich in alpha-pinene (31.6%) and alpha-bisabolol (30.9%). Both examined essential oils produced a significant dose-dependent antinociceptive effect. The corresponding ED50 +/- SEM in producing antinociception were 45.9 +/- 4.9 mg/kg and 42.4 +/- 2.1 mg/kg for L. zernyi and L. ochridanum oil, respectively. Both essential oils also significantly reduced paw edema in a dose-dependent manner. The estimated ED50 +/- SEM values for the anti-edematous effect were 36.3 +/- 4.5 mg/kg for L. zernyi oil and 45.1 +/- 11.3 mg/kg for L. ochridanum oil. These results suggest that the essential oils of both investigated Laserpitium species may be effective against pain and edema present in various inflammatory conditions.

Chloroform extracts of the underground parts of two Balkan endemic Laserpitium species, Laserpitium zernyi Hayek and Laserpitium ochridanum Micevski, were chemically investigated. Five unknown guaianolides from the class of slovanolides, of which four were additionally 2β-esterified, as well as two lactones, previously identified in other Laserpitium species, were isolated from the L. ochridanum extract. From the L. zernyi extract one slovanolide derivative was isolated for the first time in the genus Laserpitium. In addition, the phenylpropanoid latifolone and six known sesquiterpene lactones, characterised as derivatives of slovanolide and silerolide, were isolated from the extracts of both species. The cytotoxic activities of the total extracts and the isolated compounds were tested using MTT and SRB assays on the two human breast cancer cell lines, MCF 7/6 and MCF 7/AZ. The extracts exerted cytotoxic activities with the IC(50) values ranging 65.21-348.25 μg/mL. The L. ochridanum extract was most potent in the MTT test with IC(50) values of 65.21 and 66.09 μg/mL in the MCF 7/AZ and MCF 7/6 cell lines, respectively. The highest cytotoxic activity exerted 2β,8α-di-angeloyloxy-10β-hydroxy-6αH-guaian-3,(7-11)-dien-12,6-olide, a slovanolide derivative with an additional double bond in lactone ring, on highly invasive MCF 7/6 cell line, with IC(50) value 0.7 μM in both assays tested. Generally, guaianolides with a higher number of ester moieties at the positions 2β, 8α, 10β or 11α exhibited IC(50) values in the micromolar range, while eudesmanolides and guaianolides with a lower number of esters did not induce significant cytotoxicity., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The essential oil obtained by hydrodistillation from the roots of Anthriscus nemorosa (Bieb.) Sprengel (Umbelliferae) was analyzed by GC and GC-MS. Among sixty-two compounds identified (representing 89.0% of the total oil), the main components were: n-nonane (12.1%), n-hexadecanol (6.9%), delta-cadinene (6.4%), beta-pinene (6.0%) and germacrene D (5.4%). Furthermore, the antimicrobial activity of the oil was evaluated against the Gram-positive bacteria Staphylococcus epidermidis (ATCC 12228) and Bacillus subtilis (ATCC 6633), the Gram-negative bacterium Escherichia coli (ATCC 25922), and a yeast Candida albicans (ATCC 10259 and ATCC 24433) using the broth microdilution method.

The composition and antimicrobial activity of the essential oil from the underground parts (roots and rhizomes) of Laserpitium zernyi Hayek (Apiaceae) were investigated. The pale blue essential oil, obtained by hydrodistillation, was analyzed by GC and GC-MS. Forty-three compounds were identified (94.3% of total oil). The main constituent, beside alpha-pinene (31.6%) was alpha-bisabolol (30.9%), so this oil may be a novel potential natural source of this sesquiterpene alcohol. The antimicrobial activity was tested using the microdilution method against Gram (+) bacteria (Staphylococcus aureus, S. epidermidis, Micrococcus luteus, Enterococcus faecalis), Gram (-) bacteria (Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli), and two strains of yeast (Candida albicans). L. zernyi oil showed significant antibacterial activity against S. epidermidis, S. aureus and M. luteus, but lower activities against the other tested strains.

The phylogeography of typical alpine plant species is well understood in Europe. However, the genetic patterns of boreo-montane species are mostly unstudied. Therefore, we analysed the AFLPs of 198 individuals of Polygonatum verticillatum over a major part of its European distribution. We obtained a total of 402 reproducible fragments, of which 96.8% were polymorphic. The average Phi(ST) over all samples was high (73.0%). The highest number of private fragments was observed in the Cantabrian Mountains; the highest genetic diversities of the populations were detected in populations from the Alps. BAPS, Principal Coordinates and Cluster analyses revealed a deep split between the Cantabrian population and all other samples. The latter further distinguished two major groups in western and eastern Europe. These results suggest a complex biogeographical history of P. verticillatum. The Cantabrian population was most probably isolated for the longest time. Furthermore, putative glacial survival centres might have existed in the western group around the glaciated Alps and in the eastern group in the foothills of the Carpathian and Balkan mountain systems. The origin of the Scandinavian populations is still unresolved, but an origin from the southeastern Alps or the western Balkans appears the most likely scenario.

The essential oil from the aerial parts of Marrubium incanum Desr. (Lamiaceae), obtained by hydrodistillation, was analyzed by GC and GC-MS. Forty-six compounds were identified, representing 96.3% of the total oil. The main components of the oil were (E)-caryophyllene (27.0%), germacrene D (26.2%) and bicyclogermacrene (11.5%). The microbial growth inhibitory properties of the isolated essential oil were determined using the agar diffusion and broth microdilution method against seven bacterial species (Staphylococcus aureus ATCC 25923, S. epidermidis ATCC 12228, Micrococcus flavus ATCC 10240, Enterococcus faecalis ATCC 29212, Escherichia coli ATCC 25922, Klebsiella pneumoniae NCIMB 9111, and Pseudomonas aeruginosa ATCC 27853), and two strains of the yeast Candida albicans (ATCC 10259 and ATCC24433). The essential oil showed activity against all the microorganisms tested, but differences in microbial susceptibility were registered.

The present study investigated the antiinflammatory, gastroprotective and antioxidant activities of a CH(2)Cl(2) extract of western Balkan endemic Hieracium gymnocephalum Griseb. ex Pant. (Compositae). The carrageenan-induced rat paw oedema test was used as an experimental model for screening the antiinflammatory activity. The extract was administrated p.o. in doses of 25, 50, 100 and 200 mg/kg to rats and its effects compared with indomethacin, used p.o. as a reference drug. The results showed that the investigated extract reduced the oedema in a concentration-dependent manner. The obtained antiinflammatory effect was 5.9%, 11.7%, 31.2% and 44.1% at doses of 25, 50, 100 and 200 mg/kg, respectively, being statistically significant at a dose of 100 mg/kg. Indomethacin had a strong antiinflammatory effect of 73.4% at a dose of 8 mg/kg, but caused large gastric lesions. When the plant extract in the highest tested dose (200 mg/kg) was concomitantly given with indomethacin, the antiinflammatory effect was slightly enhanced, but the gastric lesions were significantly reduced. The antioxidant activity of the H. gymnocephalum extract, investigated using DPPH radical assay, OH-radical assay and TBA-test, was not substantial.

Methanol extracts of aerial flowering parts of four endemic Stachys taxa: S. anisochila VIS. et PANCIC, S. beckeana DORFLER & HAYEK, S. plumosa GRISEB. and S. alpina L. ssp. dinarica MURB. were investigated on their antioxidant activity. The extracts were studied for total antioxidant activity (TAA), along with 1,1-diphenyl 2-picryl hydrazyl (DPPH) and OH radical scavenging activity, and lipid peroxidation (LP). High correlations between total phenolics content, TAA and scavenging DPPH radical indicate that polyphenols are the main antioxidants. All Stachys extracts, with the exception of S. plumosa, exhibited high anti-DPPH activity (IC50<50 microg/ml). In concentration range from 6.25 to 50 microg/ml, all extracts scavenged OH radical above 40%, with maximal inhibitions for S. anisochila, S. alpina ssp. dinarica and S. beckeana extracts of 50.22%, 50.94% and 64.97%, respectively. Only S. plumosa extract achieved maximal activity of 60.67% at 100 microg/ml. As for LP, IC50 values for S. beckeana and S. alpina ssp. dinarica extracts were 25.07 and 49.00 microg/ml, respectively, while S. anisochila and S. plumosa extracts did not reach 50% of LP inhibition.