Seasonal Changes of Flavonoid Content in Melittis melissophyllum L. (Lamiaceae)

Melittis melissophyllum L., a medicinal plant currently used in the folk medicine, was analyzed for the content of flavonoid compounds. The plants were collected in two locations in Poland in May and September. MeOH Extracts from the leaves and flowers (separately) were analyzed by HPLC‐DAD. Eight compounds were identified in all the samples and quantitatively analyzed as cinaroside (=luteolin 7‐O‐glucoside), rutin, myricetin, quercitrin, quercetin, luteolin, kaempferol, and apigenin. M. melissophyllum accumulated the highest total amounts of flavonoids in May (flowers: from 258 to 271 mg/100 g dry weight (dw); leaves: from 143 to 155 mg/100 g dw) and significantly lower ones in September (leaves: from 83 to 92 mg/100 g dw). The main compound was cinaroside (May: up to 249 mg/100 g dw; September: up to 43 mg/100 g dw). Advanced multivariate statistical techniques (cluster analysis (CA) and principal component analysis (PCA)) were used to characterize the sample populations and to analyze the data. We report, for the first time, the results of the quantitative analysis of M. melissophyllum flavonoids and seasonal changes in their accumulation. Our results show that the time of harvesting has a significant influence on the flavonoid content in M. melissophyllum, while the geographical location does not have such an effect.     

Biochemical Characterization of Helichrysum italicum (Roth) G.Don subsp. italicum (Asteraceae) from Montenegro: Phytochemical Screening,Chemotaxonomy, and Antioxidant Properties

The chemical composition and antioxidant properties of the essential oil and EtOH extract of immortelle (Helichrysum italicum (Roth) G.Don subsp. italicum, Asteraceae) collected in Montenegro were evaluated. The essential oil was characterized by GC/MS analysis, and the content of total phenolics and flavonoids in the EtOH extract was determined using the Folin? Ciocalteu reagent. The free‐radical‐scavenging capacity (RSC) of both the essential oil and the EtOH extract was assessed with the 2,2‐diphenyl‐1‐pycrylhydrazyl (DPPH) method. Moreover, the inhibition of hydroxyl radical (^.OH) generation by the EtOH extract of immortelle was evaluated for the first time here. Neryl acetate (28.2%) and γ‐curcumene (18.8%) were the main compounds in the essential oil, followed by neryl propionate (9.1%) and ar‐curcumene (8.3%). The chemical composition of the oils of the examined and additional 16 selected Helichrysum italicum taxa described in literature were compared using principal component (PCA) and cluster (CA) analyses. The results of the statistical analyses implied the occurrence of at least four different main and three subchemotypes of essential oils. Considering the antioxidant properties, the EtOH extract of immortelle exhibited similar potential as propyl gallate and quercetin, while the essential oil exhibited relatively weak DPPH^.‐scavenging capacity.     

Antioxidant Activities of Aqueous Extract from Agrimonia pilosa Ledeb and Its Fractions

Agrimonia pilosa Ledeb is used as the tonic for asthenia and fatigue in China. Considering that the energizing effect might be correlated with antioxidant properties, we investigated the antioxidant activities of aqueous extract (AE) from Agrimonia pilosa Ledeb by assessing radical‐scavenging and anti‐lipid‐peroxidation abilities. We found that AE shows a moderate antioxidant activity to scavenge DPPH^., O , and ^.OH and inhibit β‐carotene bleaching with IC₅₀ values of 13.0, 33.2, 351, and 11.9 μg/ml, respectively, while its AcOEt‐soluble fraction (ESF) and BuOH soluble fraction (BSF) exhibit remarkable efficiencies. The ESF's IC₅₀ values of scavenging DPPH^., O , and ^.OH, and inhibiting β‐carotene bleaching are 5.6, 5.8, 171, and 7.6 μg/ml, respectively, and those of BSF are 7.5, 8.4, 82.0, and 6.2 μg/ml, respectively. In addition, we found that there is a significant correlation between total phenol content and the antioxidant activity determined by O and ^.OH scavenging, and β‐carotene‐bleaching assays. Furthermore, HPLC analysis revealed the presence of quercetin, hyperoside, quercitrin, taxifoliol, luteolin‐7‐O‐β‐D ‐glucopyranoside, and rutin in Agrimonia pilosa Ledeb . Thus, we suggest that the extracts from Agrimonia pilosa Ledeb , could be considered as natural antioxidant sources and dietary nutritional supplements to prevent oxidation‐related diseases.     

Variability in the Essential‐Oil Composition of Sideritis scardica Griseb. from Native Bulgarian Populations

The essential‐oil composition of six native populations of Sideritis scardica from Bulgaria was studied by GC‐FID and GC/MS analyses. Altogether, 37 components, representing 73.1 to 79.2% of the total oil content were identified. Among them, α‐pinene (4.4–25.1%), β‐pinene (2.8–18.0%), oct‐1‐en‐3‐ol (2.3–8.0%), phenylacetaldehyde (0.5–9.5%), β‐bisabolene (1.3–11.0%), benzyl benzoate (1.1–14.3%), and m‐camphorene ( 1 ; 0.3–12.4%) were the main compounds. All samples were characterized by low contents of oxygenated mono‐ and sesquiterpenes (≤1.6 and 2.3%, resp.). Principal component analysis (PCA) and cluster analysis (CA) showed a significant variability in the chemical composition of the studied samples as well as a correlation between the oil profiles and the ecological conditions of the natural habitats of S. scardica.     

Chemical Variability and Antioxidant Activity of Limbarda crithmoides L. Essential Oil from Corsica

The chemical compositions of 25 Corsican Limbarda crithmoides ssp. longifolia essential oils were investigated for the first time using GC‐FID, GC/MS, and NMR analyses. Altogether, 65 compounds were identified, accounting for 90.0–99.3% of the total oil compositions. The main components were p‐cymene ( 1 ; 15.1–34.6%), 3‐methoxy‐p‐cymenene ( 4 ; 11.8–28.5%), 2,5‐dimethoxy‐p‐cymenene ( 5 ; 5.9–16.4%), thymol methyl ether ( 6 ; 1.3–14.9%), α‐phellandrene ( 2 ; 0.9–11.9%), and α‐pinene ( 3 ; 0.2–13.4%). The chemical variability of the Corsican oil samples was studied using multivariate statistical analysis, which allowed the discrimination of two main clusters. A direct correlation between the water salinities of the plant locations and the chemical compositions of the L. crithmoides essential oils was evidenced. Indeed, essential oils rich in 1 (30.4–34.6%) were found in samples growing in the wetlands of the southern oriental coast, which exhibit high salinity levels (24.4±0.2–33.9±0.2 ppt), and essential oils with lower contents of 1 (15.1–27.3%) were isolated form samples growing in the wetlands of northern Corsica, which exhibit lower salinity levels (10.90±0.20–15.47±0.15 ppt). The antioxidant potential of L. crithmoides essential oil was also investigated, by assessing the DPPH^.‐ and ABTS^.+‐scavenging activities and the reducing power of ferric ions, and was found to be interesting. Moreover, using bioassay‐guided fractionation of the essential oil, a higher antioxidant activity was obtained for the oxygenated fraction and both ester and alcohol subfractions.     

Environmental and seasonal impacts on the chemical composition of Satureja horvatii Šilić (Lamiaceae) essential oils

The chemical composition of the essential oils of Satureja horvatii Šilić from two natural habitats (Mt. Orjen and Mt. Lovćen in Montenegro) and from cultivated plants (Belgrade, Serbia) were characterized. For the latter, plants from the locus classicus, i.e., Orjenske Lokve (Mt. Orjen), were transferred to Belgrade and, after three years of cultivation, the chemical composition of their essential oils at different phenological stages was analyzed. The essential oils were obtained from the aerial parts of the plants by hydrodistillation and analyzed by GC and GC/MS. The yields and chemical compositions of the S. horvatii oils showed significant differences between the plants collected in the natural habitats and those from cultivation, as well as between the plants at different phenological stages. In the populations from the natural habitats, growing in Mediterranean conditions, the most abundant oil constituents were the phenols thymol (63.7% in the samples from Mt. Orjen) or carvacrol (68.1% in the samples from Mt. Lovćen), while the oils from the cultivated plants (Belgrade), growing in continental conditions, were dominated by linalool (up to 65.8 and 55.9% in average). The basic characteristics of the essential oil from plants at the early phenological stage (before flowering) were high percentages of linalool (37.4%), thymol (27.3%), and carvacrol (12.2%). At the stage of flowering, the percentage of linalool (56.6–57.5%) increased, while those of thymol (15.5–15.8%) and carvacrol (1.4–1.5%) significantly decreased. The essential oil of plants in the full stage of fruiting was characterized by the domination of linalool (58.4 and 65.8%) and lower percentages of thymol (7.6 and 1.3%) and carvacrol (0.7 and 0.1%). In conclusion, the oil composition of S. horvatii was found to depend on the pedoclimatic conditions of the habitat and the phenological stage of the plants.     

Essential Oils and Chemical Diversity of Southeast European Populations of Salvia officinalis L.

The essential oils of 25 populations of Dalmatian sage (Salvia officinalis L.) from nine Balkan countries, including 17 indigenous populations (representing almost the entire native distribution area) and eight non‐indigenous (cultivated or naturalized) populations were analyzed. Their essential‐oil yield ranged from 0.25 to 3.48%. Within the total of 80 detected compounds, ten (β‐pinene, 1,8‐cineole, cis‐thujone, trans‐thujone, camphor, borneol, trans‐caryophyllene, α‐humulene, viridiflorol, and manool) represented 42.60 to 85.70% of the components in the analyzed essential oils. Strong positive correlations were observed between the contents of trans‐caryophyllene and α‐humulene, α‐humulene and viridiflorol, and viridiflorol and manool. Principal component analysis (PCA) on the basis of the contents of the ten main compounds showed that four principal components had an eigenvalue greater than 1 and explained 79.87% of the total variation. Performing cluster analysis (CA), the sage populations could be grouped into four distinct chemotypes (A–D). The essential oils of 14 out of the 25 populations of Dalmatian sage belonged to Chemotype A and were rich in cis‐thujone and camphor, with low contents of trans‐thujone. The correlation between the essential‐oil composition and geographic variables of the indigenous populations was not significant; hence, the similarities in the essential‐oil profile among populations could not be explained by the physical proximity of the populations. Additionally, the southeastern populations tended to have higher EO yields than the northwestern ones.     

Correlations between Genetic,Morphological, and Chemical Diversities in a Germplasm Collection of the Medicinal Plant Origanum vulgare L.

In total, 42 accessions of Origanum vulgare L., mostly originating from Europe, were evaluated, to detect molecular, quantitative morphological, and chemotype polymorphisms and to discover possible correlations between them. Twelve traits related to morphological characteristics were measured. The components in the essential oils were identified by GC/MS analysis, and the oil contents of 18 major compounds were determined. A total of 477 molecular polymorphisms including 214 AFLP (amplified fragment length polymorphism) and 263 SAMPL (selectively amplified microsatellite polymorphic loci) were used for genotyping. Euclidean distances of morphological and chemotypic data and genetic distances (1 – Dice's similarity) of molecular markers were compared by applying Mantel tests to ascertain the congruencies between them. A relatively high correlation between chemotypic patterns and genetic markers was identified, while a lower correlation was found between the morphological and genetic matrices. Pairwise analyses of correlation among all traits showed that the stem diameter was correlated to the essential‐oil yield and the carvacrol content. Cluster analysis, population inference, and principal component analysis revealed a broad genetic and chemical variation among the accessions. The knowledge of these diversities, found in this study, will allow a plant improvement of Origanum vulgare related to pharmaceutical and spice uses.     

Phytochemical and morphological characterization of Satureja khuzistanica Jamzad populations from Iran

Satureja khuzistanica is an endemic herb growing wild in Iran with interesting pharmacological and biological properties. Here, as an initial step of the domestication process, the variability of phytochemical and morphological traits among 69 individuals of eight natural populations of the plant was studied. The investigated characteristics were the essential oil content and composition, the rosmarinic acid (RA) content, and the leaf and flower morphologies. The Abdanan and Kaver populations showed the highest oil contents. The characterization by GC-FID and GC/MS analyses of the oils revealed that all 69 sampled individuals had carvacrol as the main component with very high contents (89.59-95.41%). The content of RA of the MeOH extracts of S. khuzistanica showed a high level of variability (coefficient of variation (CV) 50.0%) ranging from 0.59% (w/w) in the Paalam population to 1.81% (w/w) in the Abdanan population. The peduncle length and the leaf surface area (CVs of 47.39 and 47.21%, resp.) were the most variable morphological characteristics among the examined populations. The high level of phytochemical and morphological variability among the studied populations suggests a breeding approach during the domestication, to gain new, promising, and homogenous cultivars, attractive for the industry and agriculture.     

Essential‐Oil Composition and Chemical Variability of Senecio vulgaris L. from Corsica

The chemical composition of the essential oils isolated from the aerial parts of Senecio vulgaris plants collected in 30 Corsican localities was characterized using GC‐FID and GC/MS analyses. Altogether, 54 components, which accounted for 95.2% of the total oil composition, were identified in the 30 essential‐oil samples. The main compounds were α‐humulene ( 1 ; 57.3%), (E)‐β‐caryophyllene ( 2 ; 5.6%), terpinolene ( 3 ; 5.3%), ar‐curcumene ( 4 ; 4.3%), and geranyl linalool ( 5 ; 3.4%). The chemical composition of the essential oils obtained from separate organs and during the complete vegetative cycle of the plants were also studied, to gain more knowledge about the plant ecology. The production of monoterpene hydrocarbons, especially terpinolene, seems to be implicated in the plant‐flowering process and, indirectly, in the dispersal of this weed species. Comparison of the present results with the literature highlighted the originality of the Corsican S. vulgaris essential oils and indicated that α‐humulene might be used as taxonomical marker for the future classification of the Senecio genus. A study of the chemical variability of the 30 S. vulgaris essential oils using statistical analysis allowed the discrimination of two main clusters according to the soil nature of the sample locations. These results confirmed that there is a relation between the soil nature, the chemical composition of the essential oils, and morphological plant characteristics. Moreover, they are of interest for commercial producers of essential oil in selecting the most appropriate plants.     

Population Variability of Essential Oils of Pinus heldreichii from the Scardo‐Pindic Mountains Ošljak and Galičica

The needle‐terpene profiles of two natural Pinus heldreichii populations from Mts. O?ljak and Gali?ica (Scardo‐Pindic mountain system) were analyzed. Among the 68 detected compounds, 66 were identified. The dominant constituents were germacrene D (28.7%), limonene (27.1%), and α‐pinene (16.2%). β‐Caryophyllene (6.9%), β‐pinene (5.2%), β‐myrcene (2.3%), pimaric acid (2.0%), α‐humulene (1.2%), and seven additional components were found to be present in medium‐to‐high amounts (0.5–10%). Although the general needle‐terpene profile of the population from Gali?ica was similar to those of the populations from Lov?en, Zeletin, Bjelasica, and Zlatibor‐Pe?ter (belonging to the Dinaric Alps), the principle‐component analysis (PCA) of seven terpenes (β‐myrcene, limonene, β‐elemene, β‐caryophyllene, α‐humulene, δ‐cadinene, and germacrene D‐4‐ol) in 121 tree samples suggested a partial divergence in the needle‐terpene profiles between the populations from the Scardo‐Pindic mountain system and the Dinaric Alps. According to previously reported data, the P. heldreichii samples from the Balkan‐Rhodope mountains lack β‐caryophyllene and germacrene D, but contain γ‐muurolene in their terpene profile. Differences in the terpene composition between populations growing in the three above‐mentioned mountain systems were compared and discussed.     

Phylogenetic relationships of the genus Chamaecyparis inferred from leaf essential oil

The species differentiation between Chamaecyparis formosensis, C. obtusa var. formosana, and C. obtusa, based on the composition of the leaf essential oils, was studied. The characterization of the oils by GC-FID and GC/MS analyses showed remarkable differences between these three essential oils. Cluster analysis (CA) and principal-component analysis (PCA) distinguished three groups of essential oils. The C. formosensis oil was dominated by α-pinene while those isolated from C. obtusa var. formosana and C. obtusa were characterized by high levels of (-)-thujopsene and α-terpinyl acetate, respectively. Moreover, the phylogenetic relationships of the genus Chamaecyparis were in agreement with previous findings based on morphological and molecular evidence. In addition, the essential oils from C. obtusa var. formosana could be classified into three chemical types, according to their different characteristic main compounds (β-elemol, (-)-thujopsene, and cis-thujopsenal). The biochemical correlations between the major constituents of the Chamaecyparis species were examined and their relationship is discussed.     

Chemotaxonomic Implications of the n‐Alkane Composition and the Nonacosan‐10‐ol Content in Picea omorika,Pinus heldreichii,and Pinus peuce

The n‐alkane composition and the nonacosan‐10‐ol content in the needle cuticular waxes of Serbian spruce (Picea omorika), Bosnian pine (Pinus heldreichii), and Macedonian pine (Pinus peuce) were compared. The amount of nonacosan‐10‐ol in the needle waxes of P. omorika was higher than those in P. heldreichii and P. peuce. The range of n‐alkanes was also wider in P. omorika (C₁₈–C₃₅) than in P. heldreichii and P. peuce (C₁₈–C₃₃). The dominant n‐alkanes were C₂₉ in the needle waxes of P. omorika, C₂₃, C₂₇, and C₂₅ in those of P. heldreichii, and C₂₉, C₂₅, C₂₇, and C₂₃ in those of P. peuce. The waxes of P. omorika contained higher amounts of n‐alkanes C₂₉, C₃₁, and C₃₃, while those of P. heldreichii and P. peuce had higher contents of n‐alkanes C₂₁, C₂₂, C₂₃, C₂₄, and C₂₆. The principal component analysis of the contents of nine n‐alkanes showed a clear separation of the Serbian spruce populations from those of the two investigated pine species, which partially overlapped. The separation of the species was due to high contents of the n‐alkanes C₂₉ and C₃₁ (P. omorika), C₁₉, C₂₀, C₂₁, C₂₂, C₂₃, and C₂₄ (P. heldreichii), and C₂₈ (P. peuce). Cluster analysis also showed a clear separation between the P. omorika populations on one side and the P. heldreichii and P. peuce populations on the other side. The n‐alkane and terpene compositions are discussed in the light of their usefulness in chemotaxonomy as well as with regard to the biogeography and phylogeny of these rare and endemic conifers.     

Variability of the Root Essential Oils of Seseli rigidum Waldst. & Kit. (Apiaceae) from Different Populations in Serbia

The chemical compositions of the essential oils of seven natural populations of Seseli rigidum were analyzed. The essential‐oil yield ranged from 0.16 to 2.09%. Analysis of variance (ANOVA) revealed that there were no statistically significant differences in the mean essential‐oil yields between the populations, and no significant influence of the climate or soil type on the oil yield was observed. In all 67 analyzed samples, the polyacetylene falcarinol was the main compound, followed by octanal, methyl linoleate, α‐muurolene, 3‐butylphthalide, falcarinone, muurola‐4,10(14)‐dien‐1β‐ol, β‐sesquiphellandrene, salvial‐4(14)‐en‐1‐one, δ‐amorphene, spathulenol, and isospathulenol. The principal component analysis (PCA), the canonical discriminant analysis (CDA), and the cluster analysis (CA) revealed differentiation between the populations based on the climate. Three groups of populations were formed; the first group was composed of samples growing in regions with a humid climate, with oils having high falcarinol and low sesquiterpene contents, and the second and third groups comprised samples exposed to semi‐arid climate, with oils characterized by a lower falcarinol and higher α‐muurolene, δ‐amorphene, β‐sesquiphellandrene, and salvial‐4(14)‐en‐1‐one contents. The semi‐arid populations were divided into two groups, which were distinguished based on the oil contents of sesquiterpenes, falcarinone, and 3‐butylphthalide. On the other hand, no clear separation between populations based on the different soil types could be observed.     

Solid-phase microextraction (SPME) analysis of six Italian populations of Ephedra nebrodensis Tineo ex Guss. subsp. nebrodensis

Headspace solid-phase microextraction (HS-SPME) coupled with GC/FID and GC/MS was applied for the first time in the analysis of the volatile fraction of an Ephedra species. Notably, six Italian populations (Marche, Abruzzo, and Sardinia) of Ephedra nebrodensis subsp. nebrodensis, covering almost the entire Italian area, were investigated to examine the chemical variability and to support the taxonomy of the species. A fiber screening with polymethylsiloxane (PDMS), Carboxen(TM) /polymethylsiloxane (CAR/PDMS), and polymethylsiloxane/divinylbenzene (PDMS/DVB) coatings, together with an optimization of the extraction conditions were carried out before analysis of the six populations. A total of 119 volatiles were identified in the headspace of different samples, accounting for 63.35-100.00% of the total volatiles. A great variability was found in the qualitative composition of different samples, since only 18 components were in common among all populations. The headspace composition was dominated by sesquiterpene hydrocarbons (52.30-88.32%), with β-maaliene (traces-7.49%), β-patchoulene (traces-1.29%), β-panasinsene (traces-6.85%), α-isocomene (traces-31.25%), α-trans-bergamotene (traces-6.95%), alloaromadendrene (traces-33.20%), α-acoradiene (traces-9.41%), and γ-muurolene (0.61-16.33%) being the most abundant constituents. Noteworthy is the occurrence in a sample of two major unknown sesquiterpenes, one hydrocarbon (24.49%, RI: 1396) and one oxygenated compound (10.37%, RI: 1591), whose mass spectra were reported for the first time. Multivariate chemometric techniques, such as cluster analysis (CA) and principal component analysis (PCA), were used to characterize the samples according to the geographical origin.     

Essential‐Oil Composition of the Needles Collected from Natural Populations of Macedonian Pine (Pinus peuce Griseb.) from the Scardo‐Pindic Mountain System

The needle‐terpene profiles of two natural Pinus peuce populations from the Scardo‐Pindic mountain system (Mt. O?ljak and Mt. Pelister) were analyzed. Among the 90 detected compounds, 87 were identified. The dominant constituents were α‐pinene (45.5%), germacrene D (11.1%), β‐pinene (10.8%), and camphene (10.3%). The following eight additional components were found to be present in medium‐to‐high amounts (0.5–10%): bornyl acetate (5.0%), β‐phellandrene (3.4%), β‐caryophyllene (2.9%), β‐myrcene (0.9%), germacrene D‐4‐ol (0.9%), tricyclene (0.7%), (E)‐hex‐2‐enal (0.7%), and bicyclogermacrene (0.6%). Although the general needle‐terpene profiles of the populations from Mt. O?ljak and Mt. Pelister were found to be similar to those of the populations from Zeletin, Sjekirica, and Mokra Gora (Dinaric Alps), principle component analysis (PCA) of eight terpenes (α‐pinene, β‐myrcene, α‐terpinolene, bornyl acetate, α‐terpinyl acetate, β‐caryophyllene, trans‐β‐farnesene, and germacrene D) in 139 tree samples suggested a divergence between the two population groups, i.e., the samples from the Scardo‐Pindic mountain system and those from the Dinaric Alps. Genetic analysis of the β‐pinene content demonstrated a partial divergence between the two geographical groups. The profiles of both population groups differed from those published for populations from the Balkan‐Rhodope mountains system (literature results), which were characterized by high contents of bornyl acetate and citronellol (Greek populations) or δ‐car‐3‐ene (Bulgarian populations).     

Chemical differences in volatiles between Melittis melissophyllum L. subsp. melissophyllum and subsp. albida (Guss) P. W. Ball (Lamiaceae) determined by solid-phase microextraction (SPME) coupled with GC/FID and GC/MS

Melittis melissophyllum (Lamiaceae) is a perennial herb, typical of woody places, occurring in Italy with two subspecies, i.e., melissophyllum and albida. So far, the classification of these two taxa was only based on morphology, i.e., the presence of glandular trichomes, the dimension of the leaves, and the number of teeth on each side as the main discriminant characters. To find marker compounds to chemically discriminate the subsp. melissophyllum with respect to the subsp. albida, a solid-phase microextraction SPME analysis coupled with GC/FID (=flame ionization detector) and GC/MS was carried out. SPME proved to be a chemotaxonomically useful technique that permitted a clearly differentiation of the two subspecies at headspace level. The subsp. melissophyllum was characterized by high amount of the mushroom alcohol oct-1-en-3-ol and the phenolic coumarin, whilst the subsp. albida exhibited a high content in monoterpenes and sesquiterpenes, α-pinene, sabinene, and (E)-caryophyllene being the major compounds. Multivariate chemometric techniques, such as cluster analysis (CA) and principal-component analysis (PCA), were used to support chemical data and characterize the population according to the taxonomy. In addition, the micromorphology and distribution of glandular trichomes of both subspecies were studied by scanning electron microscopy (SEM).     

Variability of n‐Alkanes and Nonacosan‐10‐ol in Natural Populations of Picea omorika

This is the first report of population variability of the contents of n‐alkanes and nonacosan‐10‐ol in the needle epicuticular waxes of Serbian spruce (Picea omorika). The hexane extracts of needle samples originated from three natural populations in Serbia (Vranjak, Zmajeva?ki potok, and Mile?evka Canyon) were investigated by GC and GC/MS analyses. The amount of nonacosan‐10‐ol varied individually from 50.05 to 74.42% (65.74% in average), but the differences between the three investigated populations were not statistically confirmed. The results exhibited variability of the composition of n‐alkanes in the epicuticular waxes with their size ranging from C₁₈ to C₃₅. The most abundant n‐alkanes were C₂₉, C₃₁, and C₂₇ (35.22, 13.77, and 12.28% in average, resp.). The carbon preference index of all the n‐alkanes (CPI_total) of the P. omorika populations (average of populations I–III) ranged from 3.3 to 11.5 (mean of 5.9), while the average chain length (ACL) ranged from 26.6 to 29.2. The principal component and cluster analyses of the contents of nine n‐alkanes showed the greatest difference for the population growing in the Mile?evka Canyon. The obtained results were compared with previous literature data given for other Picea species, and this comparison was briefly discussed.     

Chemosystematic Analyses of Gingidia Volatiles

The leave volatiles of six Gingidia species from New Zealand and Australia and the seed volatiles of G. grisea were characterized by solid‐phase microextraction (SPME)‐GC/MS analysis. This technique, using a small quantity of samples and automated extraction, gave repeatable results, with maximum sensitivity for medium volatility compounds. The major monoterpenes among the volatiles, i.e., β‐phellandrene ( 4 ), limonene ( 6 ), and γ‐terpinene ( 5 ), and phenylpropanoids, i.e., estragole ( 3 ), (E)‐anethole ( 7 ), and myristicin ( 1 ), showed to be useful chemotaxonomic markers. For G. grisea leaves and seeds, similar compositions were detected, characterized by high contents of 4 . As leaves were more readily available for study than seeds, they were used for further investigations. The G. grisea leaf volatiles showed infraspecific variation in the ratio of 4 / 5 between and within sites of collection. The G. montana leaf volatiles also showed infraspecific variation, with high contents of 3 at one site and high contents of 7 at another. The SPME‐GC/MS analysis of G. montana herbarium voucher specimens resulted in the identification of further chemotypes for this species. The volatiles of the G. amphistoma samples were all dominated by 7 and those of the G. haematitica samples were rich in 5 . Moreover, single plants of two Australian Gingidia species were analyzed; the volatiles of G. harveyana showed high concentrations of 5 and 7 , whereas those of G. rupicola were dominated by 5 and 1 .     

Composition of Leaf n‐Alkanes in Three Satureja montana L. Subspecies from the Balkan Peninsula: Ecological and Taxonomic Aspects

The composition of the epicuticular leaf n‐alkanes of eight populations of three Satureja montana subspecies (S. montana L. subsp. pisidica (Wettst.) ?ili? , S. montana L. subsp. montana, and S. montana L. subsp. variegata (Host ) P. W. Ball ), from central and western areas of the Balkan Peninsula was characterized by GC‐FID and GC/MS analyses. In the leaf waxes, 15 n‐alkane homologs with chain‐lengths ranging from C₂₁ to C₃₅ were identified. The main n‐alkane in almost all samples was n‐nonacosane (C₂₉), but differences in the contents of three other dominant n‐alkanes allowed separating the coastal from the continental populations. The diversity and variability of the epicuticular‐leaf‐n‐alkane patterns and their relation to different geographic and bioclimatic parameters were analyzed by several statistical methods (principal component, discriminant, and cluster analyses as well as the Mantel test). All tests showed a high correlation between the leaf n‐alkane pattern and the geographical distribution of the investigated populations, confirming the differentiation between S. montana subsp. pisidica and the other two subspecies. The S. montana subsp. variegata and S. montana subsp. montana populations are geographically closer and their differentiation according to the leaf‐n‐alkane patterns was not clear, even though there was some indication of discrimination between them. Moreover, most of the bioclimatic parameters related to temperature were highly correlated with the differentiation of the coastal and the continental populations.