Large Scale Geographical Mapping of Essential Oil Volatiles in Heracleum (Apiaceae): Identification of Novel Compounds and Unraveling Cryptic Variation

Conspecific populations of plants in their native range are expected to show considerable variation due to long‐term ecological and evolutionary factors. We investigated the levels of secondary metabolites in Heracleum including H. persicum a valuable medicinal plant to depict the magnitude of cryptic variation and the potential significance of novel chemical traits. The essential oil volatiles from fruits of 34 populations from different species of Heracleum in Iranian distribution range and a native of H. sphondylium and an invasive population of H. persicum from Norway were analyzed with GC/MS. Out of 48 compounds identified, a contrasting pattern in the level of two major compounds, octyl acetate and hexyl butyrate was found among all studied species. Interestingly, a significant geographic pattern was observed; the hexyl butyrate/octyl acetate ratio was high (range 1.8 – 3.2) in the northwestern Iranian populations of H. persicum compared to that in northern and central populations (range 0.3 – 0.9). Four populations from Zagros mountains also exhibited a unique composition. Anethole was found in two populations of H. persicum from central Zagros, which has not been previously reported for essential oil of fruits of Heracleum so far. The results suggest high efficiency of large scale sampling from distribution range of species in identifying novel compounds. The unique pattern of geographic structuring also provides novel information to unravel cryptic variation in Heracleum.     

Chemical Diversity and Antimicrobial Activity of Volatile Compounds from Zanthoxylum zanthoxyloides Lam. according to Compound Classes,Plant Organs and Senegalese Sample Locations

The chemical diversity of Zanthoxylum zanthoxyloides growing wild in Senegal was studied according to volatile compound classes, plant organs and sample locations. The composition of fruit essential oil was investigated using an original targeted approach based on the combination of gas chromatography (GC) and liquid chromatography (LC) both coupled with mass spectrometry (MS). The volatile composition of Z. zanthoxyloides fruits exhibited relative high amounts of hydrocarbon monoterpenes (24.3 – 55.8%) and non‐terpenic oxygenated compounds (34.5 – 63.1%). The main components were (E)‐β‐ocimene (12.1 – 39%), octyl acetate (11.6 – 21.8%) and decanol (9.7 – 15.4%). The GC and GC/MS profiling of fruit essential oils showed a chemical variability according to geographical locations of plant material. The LC/MS/MS analysis of fruit oils allowed the detection of seven coumarins in trace content. The chemical composition of fruit essential oils was compared with volatile fractions of leaves and barks (root and trunk) from the same plant station. Hexadecanoic acid, germacrene D and decanal were identified as the major constituents of leaves whereas the barks (root and trunk) were dominated by pellitorine (85.8% and 57%, respectively), an atypic linear compound with amide group. The fruit essential oil exhibited interesting antimicrobial activities against Staphylococcus aureus and Candida albicans, particularly the alcohol fraction of the oil.     

Chemical Composition and Antimicrobial Activity of the Essential Oil from Aerial Parts and Roots of Eryngium barrelieri Boiss. and Eryngium glomeratum Lam. from Tunisia

The study of chemical composition and biological activity of unexplored essential oils may open new perspectives on their potential use in facing major health concerns such as drug‐resistant infections. The present study investigates the chemical composition and antimicrobial effects of previously unstudied essential oils obtained from genus Eryngium: Eryngium glomeratum Lam . and Eryngium barrelieri Boiss . The chemical compositions of the essential oils from aerial parts and roots of both species were studied using GC and GC/MS analytical technics. The analysis led to the identification of 102 compounds totalizing 85 – 94% of all detected compounds. Essential oils were characterized by the predominance of oxygenated sesquiterpenes. The oils obtained from aerial parts were tested against 36 microbial strains by agar dilution method and showed minimum inhibitory concentrations (MIC) in the range of 2 – 625 μg/ml. A strong antibacterial activity against multiresistant Pseudomonas aeruginosa was observed especially from E. glomeratum essential oil with MIC value up to 2 μg/ml. These findings give significant information about the pharmacological activity of these essential oils, which suggest their potential use to develop new remedies, or as sources of active compounds.     

Diversity of Secondary Metabolites in the Liverwort Syzygiella rubricaulis (Nees) Stephani (Jamesoniellaceae,Marchantiophyta) from Neotropical High Mountains

In order to evaluate the chemical diversity of Syzygiella rubricaulis (Nees ) Stephani , a species with a disjunct distribution in the neotropical high mountains, a phytochemical study was carried out with samples from 12 different populations of different altitudes from four South American countries. The chemical profiles of lipophilic extracts were analyzed by GC/MS for each population and 50 different compounds were found with the predominance and richness of sesquiterpenes. The majority of the compounds were found only in one population and the total number of substances ranged from 1 to 15 among the populations, but these numbers were not correlated with altitude, and characterize each population as distinct, based on similarity analysis. The qualitative and quantitative variations of metabolites found are a response to different conditions, under which they live, mostly likely altitudinal conditions. Further studies on the quantification of these chemicals may provide information on their ecological roles and importance for the distribution of S. rubricaulis at different altitudes. Despite the known richness of secondary metabolites produced by bryophytes, they are still poorly explored in the context of the ecological expressions.     

Inhibitory Activity and Chemical Characterization of Daucus carota subsp. maximus Essential Oils

The essential oils (EOs) of green seeds from Daucus carota subsp. maximus growing wild in Pantelleria Island (Sicily, Italy) were characterized. EOs were extracted by steam distillation, examined for their inhibitory properties against food‐borne Gram‐positive and Gram‐negative bacteria and analyzed for the chemical composition by gas chromatography (GC) and mass spectrometry (MS). Undiluted EOs showed a large inhibition spectrum against Gram‐positive strains and also vs. Acinetobacter spp. and Stenotrophomonas maltophilia. The minimum inhibition concentration (MIC) was in the range 1.25 – 2.50 μl/ml for the most sensitive strains. The chemical analysis indicated that D. carota subsp. maximus EOs included 34 compounds (five monoterpene hydrocarbons, six oxygenated monoterpenes, 14 sesquiterpene hydrocarbons, four oxygenated sesquiterpenes, camphorene and four other compounds), accounting for 95.48% of the total oil, and that the major chemicals were carotol, β‐bisabolene, and isoelemicin.     

Variation in Chemical Composition and Antibacterial Activity of the Essential Oil of Wild Populations of Phlomis olivieri

Various species of the genus Phlomis have been reported to produce metabolites demonstrating significant pharmacological efficiency. In this study, the essential oils from twelve populations of Phlomis olivieri collected from natural habitats were investigated for their chemical components. The hydrodistillated essential oil analyzed by GC‐FID and GC/MS. Analyses revealed 27 compounds, constituting 90.52 – 98.51% of the essential oils. Results indicated that the major components of the essential oils from various populations of P. olivieri were germacrene D (26.54 – 56.41%), bicyclogermacrene (6.38 – 30.55%), β‐caryophyllene (5.32 – 24.52%) and α‐pinene (1.29 – 15.53%). Principal component analyses (PCA) was used to identify any geographical variations in essential oil composition. Notably, three groups of Iranian P. olivieri populations were determined according to the major compounds. Results of the in vitro antibacterial activity indicated that P. olivieri essential oils showed good inhibitory activities against bacteria, especially Bacillus subtilis. The results of this study gave new insights for cultivation and industrial uses of P. olivieri in Iran.     

Essential Oils of Myrtaceae Species Growing Wild in Tunisia: Chemical Variability and Antifungal Activity Against Biscogniauxia mediterranea,the Causative Agent of Charcoal Canker

The chemical composition of five Eucalyptus species and five Myrtus communis L. populations was investigated using GC/MS and GC‐FID. For Eucalyptus essential oils, 32 compounds, representing 88.56 – 96.83% of the total oil according to species, were identified. The main compounds were 1,8‐cineole, α‐pinene, p‐cymene, γ‐gurjunene, α‐aromadendrene, and β‐phellandrene. For Myrtle essential oils, 26 compounds, representing 93.13 – 98.91% of the total oil were identified. α‐Pinene, 1,8‐cineole, linalool, and myrtenyl acetate were found to be the major compounds. Principal component analysis (PCA) showed chemical differentiation between Eucalyptus species and between Myrtle populations. Biscogniauxia mediterranea, the causative agent of charcoal canker, was identified according to its morphological and molecular characteristics. Essential oils of the investigated Eucalyptus species and Myrtle populations were tested for their antifungal capacity against this fungus. The antifungal activity varied according to the essential oil composition. Biscogniauxia mediterranea exhibited powerful resistance to some essential oils including them of Eucalyptus lehmannii and Eucalyptus sideroxylon but it was very sensitive to Eucalyptus camaldulensis oil (IC₅₀ = 3.83 mg/ml) and M. communis oil from Zaghouan (IC₅₀ = 1 mg/ml). This sensitivity was found to be correlated to some essential oil compounds such as p‐cymene, carvacrol, cuminaldehyde, and linalool.     

Chemical Composition of Essential Oils and Aromatic Waters from Different Italian Anthemis maritima Populations

The chemical composition of the essential oils and aromatic waters isolated from six Italian Anthemis maritima populations was determined by GC‐FID and GC/MS analyses. In total, 122 and 100 chemical compounds were identified in the essential oils and the aromatic waters, respectively. The main compound classes represented in the oils were monoterpene hydrocarbons, oxygenated monoterpenes, sesquiterpene hydrocarbons, oxygenated sesquiterpenes, and terpene esters. Multivariate chemometric techniques such as cluster analysis (CA) and principal coordinate analysis (PCO) were used to classify the samples according to the geographical origin. Statistical analysis allowed the attribution of the analyzed populations to different chemotype groups.     

HS‐SPME/GC Analysis Reveals the Population Variability of Terpene Contents in Juniperus communis Needles

Juniperus communis var. communis L. is an aromatic plant – typical boreal element of flora. In the extensive literature concerning J. communis, there is much data on the composition and the content of essential oil of needles and coneberries, but a detailed analysis of terpene distribution within and between populations is missing. A representative pool of 74 J. communis individuals originating from ten populations of Northern Poland was investigated in order to evaluate the intra‐ and interpopulational variability of the terpene pattern. Headspace solid‐phase microextraction (HS‐SPME) coupled with GC/MS and GC/FID was applied in achiral and enantioselective analysis. The majority of the samples (85%), despite different origin, were similar in the terpene pattern. High diversity of terpenes was observed within the populations and low diversity between them. High variation of enantiomeric composition was in accordance with large variation of individual compounds in general (achiral analysis). J. communis samples from Northern Poland could be distinguished by the α‐pinene/sabinene ratio, and they were divided into three chemical races.     

Genetic diversity and maternal origin of Northeast African and South American donkey populations

To investigate the mtDNA variation and origin of maternal lineages in South American donkeys and to reassess the domestication of donkeys in northeast Africa, we analyzed sequences (489 bp of the D‐loop) from 323 domestic donkeys sampled from Peru, Brazil, Ethiopia and Egypt. Altogether, the 323 sequences displayed 53 different haplotypes (45 in Ethiopia, 14 in Egypt, eight in Peru and six in Brazil). Among the four populations, Egyptian donkeys possessed the highest haplotype diversity (0.910 ± 0.032), followed by Brazilian donkeys (0.879 ± 0.060). The Clade I haplotypes dominated in Peruvian donkeys (65%), whereas Clade II haplotypes dominated in Brazilian donkeys (67%). Estimates of F_ST values showed a high genetic differentiation between Peruvian and Brazilian donkey populations (F_ST = 0.4066), which could be explained by the complex introduction history of South American donkeys. Phylogeographic analysis indicates that northeast Africa could be the most probable domestication center for Clade I donkeys. Analysis of molecular variance confirmed a weak genetic structure in domestic donkey populations among four continents (Europe, Asia, Africa and South America).     

Discovering variation of secondary metabolite diversity and its relationship with disease resistance in Cornus florida L.

Understanding intraspecific relationships between genetic and functional diversity is a major goal in the field of evolutionary biology and is important for conserving biodiversity. Linking intraspecific molecular patterns of plants to ecological pressures and trait variation remains difficult due to environment‐driven plasticity. Next‐generation sequencing, untargeted liquid chromatography–mass spectrometry (LC‐MS) profiling, and interdisciplinary approaches integrating population genomics, metabolomics, and community ecology permit novel strategies to tackle this problem. We analyzed six natural populations of the disease‐threatened Cornus florida L. from distinct ecological regions using genotype‐by‐sequencing markers and LC‐MS‐based untargeted metabolite profiling. We tested the hypothesis that higher genetic diversity in C. florida yielded higher chemical diversity and less disease susceptibility (screening hypothesis), and we also determined whether genetically similar subpopulations were similar in chemical composition. Most importantly, we identified metabolites that were associated with candidate loci or were predictive biomarkers of healthy or diseased plants after controlling for environment. Subpopulation clustering patterns based on genetic or chemical distances were largely congruent. While differences in genetic diversity were small among subpopulations, we did observe notable similarities in patterns between subpopulation averages of rarefied‐allelic and chemical richness. More specifically, we found that the most abundant compound of a correlated group of putative terpenoid glycosides and derivatives was correlated with tree health when considering chemodiversity. Random forest biomarker and genomewide association tests suggested that this putative iridoid glucoside and other closely associated chemical features were correlated to SNPs under selection.     

Geographically Related Variation in Epicuticular Wax Traits of Pinus nigra Populations from Southern Carpathians and Central Balkans – Taxonomic Considerations

The chemical composition of epicuticular waxes of nine populations from three Pinus nigra J. F. Arnold subspecies (namely subsp. nigra, subsp. banatica (Borbás ) Novák , and subsp. pallasiana (Lamb .) Holmboe ) from Southern Carpathians and central Balkan Peninsula were analyzed using GC/MS and GC/FID chromatography, and multivariate statistical techniques with respect to biogeography and taxonomy. In the needle waxes, four primary alcohols and 14 n‐alkanes ranging from C₂₁ to C₃₃ were identified, and the most abundant compounds were the four odd‐numbered n‐alkanes C₂₇, C₂₅, C₂₃, and C_29. Multivariate statistical analyses (CDA and CA) have shown existence of three P. nigra groups and suggested clinal differentiation as a mechanism of genetic variation across a geographic area: the first group consisted of the southernmost populations of subsp. pallasiana from Macedonia, the second consisted of the northernmost subsp. banatica populations from Romania, while all populations in Serbia described as three different subspecies (nigra, banatica, and pallasiana) formed the third group together with subsp. nigra population from Bosnia and Herzegovina. According to simple linear regression, geographic latitude and four bioclimatic parameters were moderately correlated with the contents of epicuticular wax compounds that are important in population discrimination, while stepwise multiple regression showed that latitude participated in most of the regression models for predicting the composition of the epicuticular waxes. These results agree with CDA and CA analysis, and confirmed the possibility of recognition of fine geographic differentiation of the analyzed P. nigra populations.     

Chemical Composition and Interpopulation Variability of Essential Oils of Taxus baccata L. from Serbia

The composition of the essential oil of the twigs and needles of Taxus baccata L. from three natural populations in Serbia has been determined by GC/MS analysis. Of the 91 detected compounds, 87 were identified. The most abundant compound classes were aliphatic alcohols, terpenes, aliphatic hydrocarbons, and aliphatic aldehydes, which together comprised ca. 86.92% of the total oil composition. The dominant constituents were oct‐1‐en‐3‐ol (23.48%), (3Z)‐hex‐3‐en‐1‐ol (11.46%; aliphatic alcohols), and myrtenol (11.38%; oxygenated monoterpene). The PCA of 22 selected compounds revealed differentiations of populations based on geographic distribution. The CA showed that Populations I and II from the Dinaric Alps were similar, and that Population III from the Balkan mountain system was distinct. This was the first investigation of interpopulation variability of T. baccata populations based on essential oil composition. The results of this study were compared with those of previous studies concerning volatile compounds produced by Taxus species. The results indicate that the essential oil content of T. baccata populations from this study is unique, mostly resembling the population from southeast Serbia.     

Aroma Profile of Rubus ulmifolius Flowers and Fruits During Different Ontogenetic Phases

The chemical composition of spontaneous volatile emission from Rubus ulmifolius flowers and fruits during different stages of development was evaluated by HS‐SPME‐GC/MS. In total, 155 chemical compounds were identified accounting 84.6 – 99.4% of whole aroma profile of flowers samples and 92.4 – 96.6% for fruit samples. The main constituents were α‐copaene, β‐caryophyllene, germacrene D, (E,E)‐α‐farnesene, 1,7‐octadien‐3‐one,2‐methyl‐6‐methylene, tridecane, (E)‐2‐hexenol acetate, (E)‐3‐hexenol acetate and cyperene. The results give a chemotaxonomic contribution to the characterization of the VOCs emitted from flowers and fruits during their ontogenic development.     

Genetic and Chemical Diversity in Perovskia abrotanoides Kar. (Lamiaceae) Populations Based on ISSRs Markers and Essential Oils Profile

Genetic and the essential oil composition variability among twelve Perovskia abrotanoides populations (PAbPs) growing wild in Iran were assessed by ISSR markers, GC‐FID and GC/MS, respectively. Nine selected ISSR primers produced 119 discernible bands, of them 96 (80.7%) being polymorphic. Genetic similarity values among populations ranged between 0.07 and 0.79 which indicated a high level of genetic variation. Polymorphic information content, resolving power and marker index generated by ISSR primers were, 0.31, 6.14, and 3.32, respectively. UPGMA grouped PAbPs into four main clusters. Altogether, 38 chemical compounds were identified in the oils, and a relatively high variation in their contents was found. Camphor (11.9 – 27.5%), 1,8‐cineole (11.3 – 21.3%), α‐bisabolol (0.0 – 13.1%), α‐pinene (5.9 – 10.8%), and δ‐3‐carene (0.1 – 10.5%) were the major compounds. Oxygenated monoterpenes (32.1 – 35.8%) and monoterpene hydrocarbons (25.7 – 30.4%) were the main groups of compounds in the oils studied. Cluster analysis and principal‐component analysis were used to characterize the samples according to oil components. Four main chemotypes were found to be Chemotype I (camphor/1,8‐cineol), Chemotype II (1,8‐cineole/camphor), Chemotype III (camphor/1,8‐cineol/α‐bisabolol), and Chemotype IV (camphor/δ‐3‐carene/α‐bisabolol). The information, provided here on P. abrotanoides populations, will be useful to introduce this plant into agricultural systems.     

Chemical Variability,Antioxidant and Antifungal Activities of Essential Oils and Hydrosol Extract of Calendula arvensis L. from Western Algeria

The chemical composition of the essential oils and hydrosol extract from aerial parts of Calendula arvensis L. was investigated using GC‐FID and GC/MS. Intra‐species variations of the chemical compositions of essential oils from 18 Algerian sample locations were investigated using statistical analysis. Chemical analysis allowed the identification of 53 compounds amounting to 92.3 – 98.5% with yields varied of 0.09 – 0.36% and the main compounds were zingiberenol 1 (8.7 – 29.8%), eremoligenol (4.2 – 12.5%), β‐curcumene (2.1 – 12.5%), zingiberenol 2 (4.6 – 19.8%) and (E,Z)‐farnesol (3.5 – 23.4%). The study of the chemical variability of essential oils allowed the discrimination of two main clusters confirming that there is a relation between the essential oil compositions and the harvest locations. Different concentrations of essential oil and hydrosol extract were prepared and their antioxidant activity were assessed using three methods (2,2‐diphenyl‐1‐picrylhydrazyl, Ferric‐Reducing Antioxidant Power Assay and β‐carotene). The results showed that hydrosol extract presented an interesting antioxidant activity. The in vitro antifungal activity of hydrosol extract produced the best antifungal inhibition against Penicillium expansum and Aspergillus niger, while, essential oil was inhibitory at relatively higher concentrations. Results showed that the treatments of pear fruits with essential oil and hydrosol extract presented a very interesting protective activity on disease severity of pears caused by P. expansum.     

Chemical Variability of the Essential Oil of Pituranthos scoparius from Algeria

The chemical composition of 93 oil samples from the aerial parts of Pituranthos scoparius, harvested in three regions of Algeria, was investigated by GC‐FID, GC/MS and ¹³C‐NMR. Monoterpene hydrocarbons dominated in association with phenylpropanoids and a chemical variability was found highlighting three clusters. The composition of group I (36 samples) exhibited an atypical composition characterized by a very high contents of 6‐methoxyelemicine (13.0 – 59.6%), followed by sabinene (1.1 – 43.0%) and limonene (6.6 – 39.0%), while the samples of group II (12 samples) contained a high content of limonene (9.2 – 44.0%), followed by myristicine (0.0 – 29.4%) and a lower amount of sabinene (0.8 – 2.3%). Group III (45 samples) could be divided in two subgroups. Subgroup SGIIIA was characterized by a very high content of sabinene (28.0 – 55.6%), followed by elemicine (0.0 – 29.1%), while the samples belonging to SGIIIB were characterized by the lower content of sabinene (6.2 – 35.5%) and a significant content of myristicine (1.5 – 32.4%), α‐pinene (4.2 – 31.0%) and dill apiole (0.1 – 31.4%). Each harvested region was characterized by a different chemical composition.     

Managing diversity: Domestication and gene flow in Stenocereus stellatus Riccob. (Cactaceae) in Mexico

Microsatellite markers (N = 5) were developed for analysis of genetic variation in 15 populations of the columnar cactus Stenocereus stellatus, managed under traditional agriculture practices in central Mexico. Microsatellite diversity was analyzed within and among populations, between geographic regions, and among population management types to provide detailed insight into historical gene flow rates and population dynamics associated with domestication. Our results corroborate a greater diversity in populations managed by farmers compared with wild ones (H_E = 0.64 vs. 0.55), but with regional variation between populations among regions. Although farmers propagated S. stellatus vegetatively in home gardens to diversify their stock, asexual recruitment also occurred naturally in populations where more marginal conditions have limited sexual recruitment, resulting in lower genetic diversity. Therefore, a clear‐cut relationship between the occurrence of asexual recruitment and genetic diversity was not evident. Two managed populations adjacent to towns were identified as major sources of gene movement in each sampled region, with significant migration to distant as well as nearby populations. Coupled with the absence of significant bottlenecks, this suggests a mechanism for promoting genetic diversity in managed populations through long distance gene exchange. Cultivation of S. stellatus in close proximity to wild populations has led to complex patterns of genetic variation across the landscape that reflects the interaction of natural and cultural processes. As molecular markers become available for nontraditional crops and novel analysis techniques allow us to detect and evaluate patterns of genetic diversity, genetic studies provide valuable insights into managing crop genetic resources into the future against a backdrop of global change. Traditional agriculture systems play an important role in maintaining genetic diversity for plant species.     

Chemical Composition and Allelopathic Potential of Essential Oils from Citharexylum spinosum L. Grown in Tunisia

Citharexylum spinosum L. (Verbenaceae) also known as Citharexylum quadrangulare Jacq . or Citharexylum fruticosum L. is an exotic tree introduced many years ago in Tunisia, specially used as a street and park ornamental tree. Essential oils (EOs) were obtained by hydrodistillation of the different parts (roots, stems, leaves, flowers and fruits; drupes) collected from trees grown in the area of Monastir (Tunisia). In total, 84 compounds, representing 90.1 – 98.4% of the whole oil composition, were identified by GC‐FID and GC/MS analyses. The root EO was distinguished by its high content in monoterpene hydrocarbons (α‐phellandrene; 30.8%) whereas that obtained from stems was dominated by sesquiterpene hydrocarbons (cuparene; 16.4%). The leaf oil was rich in an apocarotenoid derivative (hexahydrofarnesylacetone; 26%) and an aliphatic hydrocarbon (nonadecane; 14.5%). Flowers oil was rich in esters (2‐phenylethyl benzoate; 33.5%). Finally, drupes oil was rich in oxygenated sesquiterpenes (β‐eudesmol; 33.1%). Flowers oil showed a significant phytotoxic effect against lettuce seeds germination, it induces a total inhibition when tested at 1 mg/ml. Root and shoot elongation seemed to be more affected than germination. The inhibition of the shoot length varied from 3.6% to 100% and that of the root from 16.1% to 100%. The highest inhibition of 100% was detected for flower oil tested at 1 mg/ml. Our in vitro studies suggest a possible and new alternative use of C. spinosum EOs in herbicidal formulations, further experiments involving field conditions are necessary to confirm its herbicidal potential.