Isotope ratio analysis by HRGC-MS of monoterpene hydrocarbons from citrus essential oils

The isotope ratio of monoterpene hydrocarbons in citrus essential oils of different origins was measured by ordinary high-resolution gas chromatography-mass spectrometry (HRGC-MS). The isotope ratio (Ir) was determined by the ratio of the isotope peak intensity (m/z 137) to the molecular mass peak intensity (m/z 136) of the monoterpene hydrocarbons. The accuracy of Ir was examined by measuring monoterpene hydrocarbon standards and 13C-labeled compounds. The isotope fingerprints based on the values of monoterpene hydrocarbons from lemon, lime and yuzu essential oils were determined. These citrus essential oils were also discriminated by a principal component analysis of their Ir data. The characteristic vectors showed that alpha-terpinene, beta-pinene and beta-phellandrene were important components for distinguishing between the citrus species. It is suggested that this technique will be applicable to evaluate the quality, genuineness and origin of citrus fruits and their products.     

MS fragment isotope ratio analysis for evaluation of citrus essential oils by HRGC-MS

To evaluate the origin of citrus essential oils, the isotope ratio of fragment peaks on HRGC-MS of the volatile compounds from various citrus oils was measured. The MS fragment ratio was found by the ratio of fragment peak intensity, m+1/m (m/z). This ratio reflects the isotope effect of volatile compounds, that is, it provides information about locality, quality, and species for essential oils. Multivariate analysis based on the MS fragment ratio of monoterpene hydrocarbons clearly distinguished three citrus species, yuzu, lemon, and lime. The carbonyl fractions were also extracted from citrus essential oils by the sodium hydrogensulfite method. The isotope ratio of MS fragments of octanal, nonanal, and decanal was also examined. The results suggest that there was no significant difference in the individual fragment isotope ratios of the three aldehydes.     

Isotope ratio by HRGC-MS of citrus Junos tanaka (yuzu) essential oils: m/z 137/136 of terpene hydrocarbons

The isotope ratios of monoterpene hydrocarbons in Citrus junos Tanaka (yuzu) essential oils from different origins were determined by ordinary high-resolution gas chromatography-mass spectrometry (HRGC-MS). Both intensities of the molecular mass peaks (m/z 136) and of the isotope peaks (m/z 137) of monoterpene hydrocarbons were measured by single-ion monitoring with an MS analysis. The isotope ratios (m/z 137/136) of the ten monoterpene hydrocarbons commonly contained in citrus essential oils, alpha-pinene, beta-pinene, sabinene, myrcene, alpha-phellandrene, alpha-terpinene, limonene, gamma-terpinene, beta-phellandrene and terpinolene, were determined in yuzu samples of the highest commercial quality from 42 different production districts. Statistical treatment of these data by the t-test and sign test revealed significant differences of the isotope effects in each yuzu sample. It is suggested that this technique will be applicable for evaluating the quality, genuineness and origin of citrus fruits and their products. The isotope fingerprints were also demonstrated in several citrus fruits other than the yuzu samples.     

Isotope Ratio by HRGC-MS of Citrus junos Tanaka (Yuzu) Essential Oils: m/z 137/136 of Terpene Hydrocarbons

The isotope ratios of monoterpene hydrocarbons in Citrus junos Tanaka (yuzu) essential oils from different origins were determined by ordinary high-resolution gas chromatography-mass spectrometry (HRGC-MS). Both intensities of the molecular mass peaks (m/z 136) and of the isotope peaks (m/z 137) of monoterpene hydrocarbons were measured by single-ion monitoring with an MS analysis. The isotope ratios (m/z 137/136) of the ten monoterpene hydrocarbons commonly contained in citrus essential oils, α-pinene, β-pinene, sabinene, myrcene, α-phellandrene, α-terpinene, limonene, γ-terpinene, β-phellandrene and terpinolene, were determined in yuzu samples of the highest commercial quality from 42 different production districts. Statistical treatment of these data by the t-test and sign test revealed significant differences of the isotope effects in each yuzu sample. It is suggested that this technique will be applicable for evaluating the quality, genuineness and origin of citrus fruits and their products. The isotope fingerprints were also demonstrated in several citrus fruits other than the yuzu samples.     

Effects of several variable factors on the isotope ratio by HRGC-MS

In the isotope ratio (Ir) analysis using GC-MS, several variable factors in sampling incidental to any food analysis were investigated for yuzu fruit. The Irs of ten monoterpene hydrocarbons in yuzu essential oils from each of six fruiting positions of three trees were measured. The sign test following t-test of all the Ir values demonstrated that there was no significant difference between both sampling years of 2001 and 2002. There was also no significant variation in the Ir values among the three trees and six fruiting positions in the individual two years.     

A comparative study of Scalesia species (Asteraceae) by gas chromatographic analysis of the monoterpene hydrocarbons of their essential oils

ADSERSEN, A. & SVENDSEN, A. B., 1986. A comparative study of Scalesia species (Asteraceae) by gas chromatographic analysis of the monoterpene hydrocarbons of their essential oils. The monoterpene hydrocarbons of the essential oils of 13 species of Scalesia , Asteraceae, endemic to the Galapagos Islands, were analysed by gas chromatography to determine whether the composition of these compounds could show any characteristics for the various species. Marked differences in the monoterpene hydrocarbon patterns of the Scalesia species investigated were observed. Whereas a-pinene was usually found in amounts of 30–50%, Scalesia villosa contained only 12%. Sabinene usually occurred in amounts of 10-1.5%, but S. divisa and S. affinis contained about twice as much. The limonene content was usually less than 1%, but S. microcephala, S. baurii and S. affinis contained amounts varying from about 2% to 9.5%.     

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.     

In-vitro antimicrobial activity and chemical composition of Sardinian Thymus essential oils

Essential oils and their components are becoming increasingly popular as naturally occurring antimicrobial agents. In this work the chemical composition and the antimicrobial properties of Thymus essential oils and of their main components were determined. Three essential oils obtained from different species of Thymus growing wild in Sardinia and a commercial sample of Thymus capitatus oil were analysed. The essential oil components were identified by GC/MS analysis. The antimicrobial activity of the oils and components was determined against a panel of standard reference strains and multiple strains of food-derived spoilage and pathogenic bacteria, using a broth microdilution method. The GC/MS analysis showed that the major constituents of the oils were monoterpene hydrocarbons and phenolic monoterpenes, but the concentration of these compounds varied greatly among the oils examined. The results of the antimicrobial assay showed that essential oils extracted from Sardinian Thymus species have an antimicrobial activity comparable to the one observed in other thyme oils. It seems also confirmed that the antimicrobial properties of thyme essential oils are mainly related to their high phenolic content. Among the single compounds tested carvacrol and thymol turned out to be the most efficient against both reference strains and food-derived bacteria. The results of this study confirmed the possibility of using thyme essential oils or some of their components in food systems to prevent the growth of foodborne bacteria and extend the shelf-life of processed foods.     

Chemotaxonomic significance of the essential oils of 18 Ferula species (Apiaceae) from Iran

To evaluate the chemotaxonomic significance of the essential oils of 23 populations of 18 Iranian Ferula species, the chemical composition of the oils was investigated by GC/FID and GC/MS. Altogether, 84 constituents, representing 81.3-99.7% of the total composition of the oils, have been identified. The composition of six species of the genus, i.e., F. oopoda, F. foetida, F. behboudiana, F. diversivittata, F. galbaniflua, and F. hezarlalehzarica, has been reported for the first time. The main constituents identified were α-terpinyl acetate (73.3%), 2,3,4-trimethylthiophene (2; 49.0%), sabinene (75.3%), verbenone (5; 69.4%), β-pinene (59.0-66.3%), and (Z)-β-ocimene (41.7%). Cluster analysis (CA) of the percentage content of the essential oil components of the Ferula species resulted in the characterization of four groups, i.e., taxa containing either i) monoterpene hydrocarbons, ii) oxygenated monoterpenes, iii) organosulfur compounds, or iv) monoterpene, sesquiterpene, and aliphatic hydrocarbons as the principal classes of compounds. Based on the results obtained, the chemical independence of F. hirtella from F. szowitsiana and of F. galbaniflua from F. gummosa at the specific level was concluded and their positions as distinct species were confirmed. The chemotaxonomic relationships among the representatives of the genus Ferula have been discussed in detail.     

Essential oil of Daucus glaber Forssk

The composition of the essential oil of the fruits, leaves and stems of Daucus glaber Forssk has been studied by GC/MS. It was found that, the essential oil of the fruits consists of monoterpene hydrocarbons (limonene and sylvestrene are the majors) and phenylpropanoids (elemicin is the major). Sylvestrene has never been reported before in the essential oil of any Daucus species. The study of the essential oil of the leaves revealed the presence of monoterpene hydrocarbons; limonene and gamma-terpinene are the majors and a small amount of sylvestrene. The essential oil of stems consists of monoterpene hydrocarbons (gamma-terpinene is the major), terpene alcohols (mainly 4-terpineol) and phenylpropanoids (myristicin and elemicin are the majors). It is interesting that, the essential oil of the fruits is free from any oxygenated terpenes while that of the stems is free from limonene and sylvestrene which are present in the essential oil of the fruits and leaves in fairly large amounts The essential oil of the fruits, leaves and stems shows broad antimicrobial activities against both gram positive and gram negative bacteria. In addition, the volatile oil of the stem, particularly, show activities against Candida albicans (yeast). Also, the prepared oils have variable cytotoxic activities with LC50 21.52, 36.01 and 42.34 microg/ml, respectively.     

Composition of the essential oils in various organs at different developmental stages of Ammi visnaga (L.) Lam. from Tunisia

The composition of the essential oils isolated by hydrodistillation from various organs at different development stages of Ammi visnaga (L.) Lam. growing in Tunisia was determined by GC/MS analysis. In particular, the oil profiles of the leaves, stems, flower buds, roots, umbels, and fruits have been examined during the whole life cycle. The oil from the flowering aerial parts was characterized by a high content of isoamyl 2-methylbutanoate. After flowering and during desiccation and fructification, the umbels and fruits expressed a high content of linalool. The oils, extracted from the roots collected in the vegetatif, buds floral, and floral stages, were rich in monoterpene aldehydes, oxygenated monoterpenes, and monoterpene hydrocarbons. The highest level of non-terpene hydrocarbons was found at the flower-bud stage, represented by 61.3% of nonane. Among the monoterpenes, sabinene (12.5%) and β-pinene (8.5%) were identified in the flower buds.     

Insecticidal activity of essential oils from eleven Eucalyptus spp. and two hybrids: lethal and sublethal effects of their major components on Blattella germanica

The aim of the current study was to evaluate the fumigant activity of the essential oils from 11 species of the genus Eucalyptus and two of their hybrids on first instar of Blattella germanica L. The fumigant activity and repellence of the four major monoterpene components of these essential oils also were tested. Fumigant activity was evaluated by exposing nymphs to the vapors emitted by 50 microl of essential oil or monoterpene in a closed container. The lowest knockdown time 50% (KT50) values, expressed in minutes, were elicited by the essential oils of the Eucalyptus grandis X Eucalyptus tereticornis (57.9) hybrid, Eucalyptus sideroxylon A. Cunn (62.0), E. grandis X Eucalyptus camaldulensis (63.8) hybrid, Eucalyptus viminalis Labill (64.1), Eucalyptus dunnii Maiden (64.5), and Eucalyptus grandis (Hill) ex Maiden (68.7). The KT50 values for the remaining essential oils ranged between 74.5 (E. saligna Smith) and 161.4 min (E. tereticornis Smith). The essential oil from the hybrid E. grandis X E. tereticornis was 3.7 times less toxic than dichlorvos (positive control). The KT50 values of monoterpenes were 38.8 for alpha-pinene, 55.3 for 1,8-cineole, 175.6 for p-cymene, and 178.3 for gamma-terpinene. Alpha-pinene was 2.5 times less toxic than dichlorvos. There was a strong positive correlation between the fumigant activity of essential oils and their corresponding 1,8-cineole and alpha-pinene concentration. Repellency was quantified using a video tracking system. Two concentrations of monoterpenes were studied (7 and 70 microg/cm2). All compounds produced a light repellent effect but only when applied at 70 microg/cm2. In all cases, the repellent effect was less than that produced by the broad-spectrum insect repellent N,N-diethyl-3-methylbenzamide (positive control).     

Composition and chemical polymorphism of the essential oils from Piper lanceaefolium

Essential oils obtained by hydrodistillation from leaves and spikes of Piper lanceaefolium H.B.K. of Costa Rica were analysed by GC-FID, GC-MS and 13C-NMR methods. Main constituents found in the oil from leaves were sesquiterpene hydrocarbons - especially beta-caryophyllene and germacrene D - and phenylpropanoids, of which elemicin and parsley apiol were the major ones. The volatile oil from spikes showed monoterpene hydrocarbons, namely alpha- and beta-pinene, and the same phenylpropanoids as in the oil from leaves as the major constituents. Results obtained in the analysis by GC-FID and GC-MS of the essential oils from individual plants of different geographic origin were submitted to chemometric cluster analysis and principal component analysis, showing the presence of three different types of oils (i) parsley apiol/elemicin, (ii) elemicin/parsley apiol/dill apiol, and (iii) parsley apiol/dill apiol.     

Toxicity of citrus essential oils against Ceratitis capitata (Diptera: Tephritidae) larvae

Citrus peel essential oils are considered to constitute the most important resistance factor of citrus fruits against fruit flies. Essential oils were obtained from three sweet orange varieties, one bitter orange and one lemon variety. Yield, chemical composition and toxicity against neonates of the Mediterranean fruit fly were determined. Based on chemical analysis, the toxicity of commercially purchased major and minor components (monoterpenes and sesquiterpenes) of essential oils was determined. In addition, fractions were prepared to evaluate the role of minor components in the toxicity of crude essential oils. Limonene was by far the most abundant ingredient (96.2–97.4%) in all sweet orange varieties and in bitter orange, while the concentration of limonene was much lower in lemon essential oils (74.3%). Orange and bitter orange essential oils were more toxic than lemon essential oils. The toxicity of orange and bitter orange essential oils was similar to that of their major component limonene. In tests of commercially purchased chemicals, the oxygenated components of essential oils were more toxic than hydrocarbons but their low concentration in citrus essential oils could not affect the toxic activity of essential oils. The presence of α-pinene and β-pinene seems to account for the lower toxicity of lemon essential oils in relation to other citrus essential oils. The importance of understanding the toxicity of essential oils in relation to their composition and their role regarding the resistance of citrus fruits to Ceratitis capitata infestation is discussed.     

Characterization of gamma-terpinene synthase from Citrus unshiu (Satsuma mandarin)

Gamma-terpinene is a monoterpene and a major component of essential oils made from citrus fruits and shows strong antioxidant activity in various assay systems. Plant gamma-terpinene synthase is a member of the monoterpene cyclase family, which produces a specific monoterpene through cyclization of geranyl diphosphate (GPP), but the monoterpene cyclases have not been fully characterized. It is necessary to prepare large amounts of gamma-terpinene synthase from Citrus unshiu (Satsuma mandarin) for the characterization, on this purpose we expressed the protein in Escherichia coli (E. coli) cells. As most monoterpene synthases have plastid-targeting signals, a gene lacking these signals was prepared and functionally expressed in E. coli cells harboring extra copies of the argU gene. The purified enzyme was incubated with GPP and the main product was confirmed to be gamma-terpinene by GC/MS.     

Autooxidation of a mixture of lemon essential oils,methyl linolenoate,and methyl oleinate

Stability of components of a mixture of methyl linolenoate and methyl oleinate with two lemon (Citrus limon L.) essential oils in hexane during their autooxidation in light was studied by gas chromatography. The essential oils differed by their quantitative ratio of components: the single-fold (1x) oil contained approximately 90% monoterpene hydrocarbons and 1.47% citral, whereas the proportions of hydrocarbons and citral in the tenfold (10x) oil were approximately 60 and 18.32%, respectively. The concentration and composition of essential oils influence the rates of fatty-acid oxidation and fatty-acid peroxide cleavage. The 1x lemon oil inhibited the oxidation of methyl linolenoate and methyl oleinate, whereas the 10x oil accelerated these processes. The distinctions in the resistance of the major components of lemon essential oil to oxidation, which are determined by their composition and antioxidant properties of unsaturated fatty acids, were revealed.     

Geographic variation in the essential oil monoterpenes of Liriodendron tulipifera L

The monoterpene component of Liriodendron tulipifera L. (Magnoliaceae) essential oils was quantitatively analysed in branch samples collected from an Ohio provenance test of 11 geographic seed sources representing the natural range of the species. Mean monoterpene composition of the essential oil is described and significant differences in the monoterpene composition among geographic sources are discussed.     

Chemical Composition and Biological Activities of Tunisian Cupressus arizonica Greene Essential Oils

The chemical composition of the essential oils obtained by hydrodistillation of leaves, stems, and female cones of Cupressus arizonica Greene , grown in Tunisia, was studied by GC‐FID and GC/MS analyses. Altogether, 62 compounds were identified, 62 in the leaf oil, 19 in the cone oil, and 24 in the stem oil. The cone and stem oils were mainly composed by monoterpene hydrocarbons (96.6 and 85.2%, resp.). In the leaf oil, the total sesquiterpene fraction constituted 36.1% and that of the monoterpene hydrocarbons 33.8% of the total oil composition. The three oils were evaluated for their in vitro herbicidal activity by determining their influence on the germination and the shoot and root growth of the four weed species Sinapis arvensis L., Lolium rigidum Gaudin , Trifolium campestre Schreb ., and Phalaris canariensis L. At the highest doses tested (0.8 and 1.0 mg/ml), the leaf essential oil inhibited either totally or almost completely the seed germination and the shoot and root growth of S. arvensis and T. campestre. The oils were also tested for their antifungal activity; however, their effects on the fungal growth were statistically not significant.     

Essential‐Oil Composition of Helichrysum italicum (Roth) G.Don ssp. italicum from Elba Island (Tuscany,Italy)

The composition of 21 essential‐oil samples isolated from Helichrysum italicum collected in seven locations of Elba Island (Tuscany, Italy), characterized by different soil types, during three different periods (January, May, and October 2010) was determined by GC‐FID and GC/EI‐MS analyses. In total, 115 components were identified, representing 96.8–99.8% of the oil composition. The oils were characterized by a high content of oxygenated monoterpenes (38.6–62.7%), while monoterpene and sesquiterpene hydrocarbons accounted for 2.3–41.9 and 5.1–20.1% of the identified constituents, respectively. The main oxygenated derivatives were nerol (2.8–12.8%) and its ester derivative neryl acetate (5.6–45.9%). To compare the chemical variability of the species within Elba Island and between the island and other localities within the Mediterranean area, studied previously, multivariate statistical analysis was performed. The results obtained showed a difference in the composition of the essential oils of H. italicum from Elba Island, mainly due to the environment where the plant grows, and, in particular, to the soil type. These hypotheses were further confirmed by the comparison of these oils with essential oils obtained from H. italicum collected on other islands of the Tuscan archipelago.     

A comparative study of ×Cupressocyparis leylandii clones (Cupressaceae) by gas chromatographic analysis of their leaf oils*

The essential oils isolated from foliage of various clones of x Cupressocyparis leylandzi were analyzed by gas chromatography to determine whether the oils could be used as aids in clonal identification. Several botanical gardens supplied foliage which was submitted to hydrodistillation. The oils were isolated under standardized conditions and then analyzed. Comparison of the oil components (mostly monoterpene hydrocarbons) revealed that it is possible to identify, on the basis of the composition of the monoterpene hydrocarbon mixture, clones 1–6 as a group and clones 10 and 11 individually. The composition of the oils from foliage of the widely propagated clones 1,2, 10 and 11 is given in more detail.