TERPENE

Terpenes and sterols are complex molecules synthesized by equally complex biosynthetic pathways. Recent progress in using the tools of genetics, molecular genetics and genetic engineering to dissect triterpene metabolism in the cytosol, and terpene metabolism in the ...

Tropical medicinal plant spices have remained underutilized in commercial food processing because of the inadequate chemical characterization of their terpene compositions. The activity of medicinal plants used in flavoring is due to their terpene contents. Terpene contents of Aframomum danielli seeds, Xylopia aethiopica fruits, Syzygium aromaticum leaves, Piper guineense seeds, and Monodora myristica seeds were determined using headspace solid-phase microextraction combined with gas chromatography-mass spectrometry. P. guineense is rich in terpinene and terpinolene (2.5 g per 100 g), S. aromaticum has caryophyllene (0.68 g per 100 g); A. danielli is rich in ocimene (1.65 g per 100 g); X. aethiopica is rich in ocimene (2.94 g per 100 g); M. mystristica is rich in delta carene (0.49 g per 100 g). Combinations of the five medicinal plant spices are being applied as flavorings in packaged tropical food.

Transgenic tomato (Solanum lycopersicum) plants that overexpress the Prosystemin gene (35S::PS) and plants with a mutation in the JA biosynthetic pathway (def1) are known to exhibit a constitutive or reduced wound response, respectively. Here it is demonstrated that several independent 35S::PS lines emit high levels of specific volatiles in addition to increased accumulation of proteinase inhibitors (PIs). Furthermore, the temporal dynamics of systemically induced volatile compounds including green-leaf volatiles, terpenes, and shikimic acid-derivatives from 35S::PS and def1 plants in response to herbivore wounding and treatment with jasmonic acid (JA) are described. Application of JA induced defense protein accumulation and volatile emissions in wild type plants, but did not further increase systemic volatile emissions from 35S::PS plants. Wounding by Manduca sexta larvae induced synthesis of defense proteins and emission of volatiles in wild type plants, but not in def1 plants. Application of jasmonic acid restored the local and systemic accumulation of defense proteins in def1, as well as enhanced herbivore-induced volatile emissions. These results provide strong support for the role of prosysteminand JA-signaling in the regulation of volatile emissions in tomato plants.

The barks of Drimys winteri are used in folk medicine as a remedy to treat several diseases, including dolorous processes. Previous pre-clinical experiments carried out in our laboratories revealed that the hydroalcoholic extract of this plant showed anti-allergenic, anti-inflammatory and antinociceptive properties. Such promising results led us to determine the analgesic compounds present in D. winteri. Through conventional chromatographic procedures with fractions of CH 2 Cl 2 and EtOAc obtained from methanolic extract, it was found that polygodial (1), 1-i-(p-methoxycynnamyl) polygodial (2), taxifolin (3) and astilbin (4), are the main components of these fractions. Compounds 1 and 2 exhibited marked antinociceptive action by intraperitoneal and oral routes against acetic acid-induced abdominal constrictions in mice, suggesting that they are responsible, at least partially, for the antinociceptive effects of this plant. In addition, both compounds were notably more potent than aspirin and acetaminophen, two well-known drugs used here as comparison.

The aroma of sweet Fiano wine, the most representative non-aromatic white wine variety in southern Italy, was evaluated by sensory and instrumental analysis to determine the influence of grape overripeness, drying and Botrytis cinerea infection. Sensory descriptive analysis was used to evaluate the sensory properties of wines. Gas chromatography/mass spectrometry (GC/MS), gas chromatography (GC) and gas chromatography/olfactometry (GC/O) techniques were used to identify and to determine the content of free and bound volatile compounds and their odour impact. A sensory descriptive analysis had revealed that the sensory descriptors such as citrus jam, dried apricot, dried figs, prune, honey and coconut, occur in sweet Fiano wine more than in base Fiano wine. Thirty five free volatile compounds had higher levels in sweet Fiano wine than in base Fiano wine, these components were mostly terpenes, b-damascenone, lactones, aldehydes and ketones. Moreover higher levels of terpenes, b-damascenone, benzyl alcohol and 2-phenylethanol were found in the bound volatile fraction of sweet Fiano wine. The main odour impact compounds in sweet Fiano wine were nerol, geraniol and linalool (orange flowers), vitispirane (camphor), lactones such as c-nonalactone (coconut), d-decalactone and c-decalactone (apricot) and 1-octen-3-ol (mushroom). These differences appear to be due to the fact that grape overripeness and the drying process allows concentration of varietal aromatic compounds and an easier transfer of them from skins to must during vinification.

Four model drugs were selected based on their lipophilicity denoted as log P (nicardipine hydrochloride log P −0.9990.1, hydrocortisone log P 1.439 0.47, carbamazepine log P 2.679 0.38, and tamoxifen log P 7.879 0.75). The enhancing activities of four terpene enhancers (fenchone log P 2.1390.30, thymol log P 3.289 0.20, D-limonene log P 4.58 9 0.23, and nerolidol log P 5.3690.38) were tested in vitro across full thickness hairless mouse skin with each of the evaluated drugs formulated in hydroxypropyl cellulose gel formulations. The relationships between lipophilicity (log P) of the terpene enhancers and model drugs and efficacy (represented by the enhancement ratio of flux ER flux) of the drugs when coadministered with the enhancers were examined using linear regression. Terpene enhancers had significant effect on the percutaneous permeation of the model drugs. Nerolidol (highest lipophilicity) provided the highest increase in the flux of the evaluated model drugs. The flux of nicardipine hydrochloride increased by approximately 135-fold, hydrocortisone by 33-fold, carbamazepine 8-fold, and tamoxifen 2-fold. The lowest increase in the flux was observed with fenchone. Linear relationships were generated between the ER flux of nicardipine hydrochloride, hydrocortisone, carbamazepine, and tamoxifen and the log P of the terpene enhancers [r =0.951, (P =0.049), r= 0.977, (P= 0.023), r=0.942, (P= 0.057), and r= 0.874, (P= 0.126), respectively]. Furthermore, the four terpene enhancers produced linear relationships, indicating that they were more effective at enhancing the penetration of hydrophilic drugs rather than lipophilic drugs r = −0.824 (P=0.176) for fenchone, r = −0.891 (P= 0.109) for thymol, r= −0.846 (P= 0.154) for limonene, and r= −0.769 (P =0.232) for nerolidol.

Background. Linalool is a common fragrance terpene that, in pure form, is not allergenic or is a very weak allergen. However, linalool autoxidizes on air exposure, and the oxidation products can cause contact allergy. In a Swedish study, oxidized linalool 6.0% in petrolatum (pet.) gave 5% positive patch test reactions in 2500 dermatitis patients. Objectives. To investigate whether oxidized linalool 6%, with a stable concentration of the main haptens, the linalool hydroperoxides (Lin-OOHs) in pet., could be a useful tool for the detection of contact allergy in an international setting. Methods. Oxidized linalool 6.0% (Lin-OOHs 1%) pet. was tested in 2900 consecutive dermatitis patients in Denmark, the United Kingdom, Singapore, Spain, Sweden, and Australia. Results. Overall, 6.9% (range 3-13%) of the patients showed positive patch test reactions to oxidized linalool. Doubtful reactions were found in 9.2% of the patients (range 0-36%). Few irritant reactions were seen. Conclusions. In an international setting, oxidized linalool has been shown to be a common allergen. Oxidized linalool 6.0% (Lin-OOHs 1%) pet. is a useful, standardized and stable tool for the detection of contact allergy in dermatitis patients. Many patients showing positive patch test reactions to oxidized linalool would not have been informed of their fragrance allergy if this specific test had not been performed.

Building occupants, including cleaning personnel, are exposed to a wide variety of airborne chemicals when cleaning agents and air fresheners are used in buildings. Certain of these chemicals are listed by the state of California as toxic air contaminants (TACs) and a subset of these are regulated by the US federal government as hazardous air pollutants (HAPs). California's Proposition 65 list of species recognized as carcinogens or reproductive toxicants also includes constituents of certain cleaning products and air fresheners. In addition, many cleaning agents and air fresheners contain chemicals that can react with other air contaminants to yield potentially harmful secondary products. For example, terpenes can react rapidly with ozone in indoor air generating many secondary pollutants, including TACs such as formaldehyde. Furthermore, ozone-terpene reactions produce the hydroxyl radical, which reacts rapidly with organics, leading to the formation of other potentially toxic air pollutants. Indoor reactive chemistry involving the nitrate radical and cleaning-product constituents is also of concern, since it produces organic nitrates as well as some of the same oxidation products generated by ozone and hydroxyl radicals.

HS-SPME analysis of terpenes does usually have inherent quantification problems when working with complex samples, especially due to the matrix effect of the substrate or the calibration solution. Three different terpene carrier matrices were compared: methanol, synthetic oil and milk fat obtained by centrifugation from milk cream. Considerable differences in calibration sensitivity parameters were observed depending on the matrix used and on the type of terpene standard analysed. For milk sample quantification purposes internal standard method was preferred using milk fat as calibration matrix. Linearity range, repeatability, recovery and limits of detection and quantification were determined. Validation parameters were different depending on the concentration and molecular structure of each terpene analysed, particularly between mono-and sesquiterpenes. The method was useful to determine in an accurate manner the terpene content in milk samples from pasture fed animals, and it will help to establish objective terpene levels to differentiate milks from specific production systems.

Volatile compounds were isolated from Costa Rican guava fruit by simultaneous steam distillationsolvent extraction according to the Likens-Nickerson method. Compounds were identified by capillary GC-MS and sensorially characterized by sniffing-GC. One hundred and seventy-three components were identified in the aroma concentrate, from which (E)--caryophyllene, R-terpineol, R-pinene, R-selinene, -selinene, δ-cadinene, 4,11-selinadiene, and R-copaene were found to be the major constituents. The amounts of aliphatic esters and terpenic compounds were thought to contribute to the unique flavor of this fruit.

Oleoresin biomass production by trees of Pinus elliottii, which is a source of terpenes for the chemical and pharmaceutical industries, was investigated. Trees were individually analyzed for oleoresin yield using the bark streak method of wounding for stimulation of resin flow. Controls included plain wounding and wounding followed by application of commercial stimulant paste, based on sulfuric acid and synthetic precursors of the phytohormone ethylene. Metal cofactors of terpene synthases and ethylene receptors applied locally as adjuvants of the oleoresin stimulant paste on wounded bark tissue improved resin yields. The use of potassium, copper, and iron did not affect significantly the composition of semiochemical monoterpenes, which are involved with pine - bark beetle interactions, and are important as oleoresin-derived products. Some adjuvants, particularly potassium, were capable of supporting oleoresin yields equivalent to those obtained with current commercial stimulant pas...

A headspace solid-phase microextraction (HS-SPME) method coupled to gas chromatography-ion trap mass spectrometry (GC-ITMS) has been developed and applied for profiling of volatile compounds released from five Ocimum basilicum L. cultivars grown under both organic and conventional conditions. Comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GCÂGC-TOFMS) was employed for confirmation of identity of volatiles extracted from the basil headspace by SPME.

Volatile oil composition of hydro-distilled (HD) and supercritical carbon dioxide (SC-CO2) essential oil of freshly collected aerial parts of Heracleum thomsonii (Umbeliferae) from the western Himalayas was studied by GC-FID and GC-MS. Results revealed qualitative and quantitative dissimilarity in the composition of hydro-distilled and SC-CO2 extracted oils. Nineteen constituents, which accounted for 89.32% of total constituents in HD oil, represented

The Uncaria genus is an important source of medicinal natural products, particularly alkaloids and triterpenes. The collected information is an attempt to cover the more recent developments in the ethnobotany, pharmacology and phytochemistry of this genus. During the past 20 years, alkaloids, terpenes, quinovic acid glycosides, flavonoids and coumarins have been isolated from Uncaria. Fifty-three novel structures are reported in this review. The species in which the largest number of compounds has been identified is the Peruvian Uncaria tomentosa or ÔcatÕs claw.Õ Pharmacological studies are described according to cytotoxicity, anti-inflammatory, antiviral, immunostimulation, antioxidant, CNS-related response, vascular, hypotensive, mutagenicity and antibacterial properties. The potential for development of leads from Uncaria continues to grow, particularly in the area of immunomodulatory, anti-inflammatory and vascular-related conditions. The information summarized here is intended to serve as a reference tool to practitioners in the fields of ethnopharmacology and natural products chemistry. (G.B. Hammond). www.elsevier.com/locate/phytochem Phytochemistry 66 (2005) 5-29 PHYTOCHEMISTRY 6 M.E. Heitzman et al. / Phytochemistry 66 (2005) 5-29

2 Extracting and trapping biogenic 3 volatile organic compounds stored 4 in plant species Elena Ormeñ o, Allen Goldstein, Ü lo Niinemets 7 Biogenic volatile organic compounds (BVOCs), released by practically all plants, have important atmospheric and ecological 8 consequences. Because BVOC-emission measurements are especially tedious, complex and extremely variable between species, 9 two approaches have been used in scientific studies to try to estimate BVOC-emission types and rates from plant species. The first, 10 which has known little success, involves grouping species according to plant-taxonomy criteria (typically, genus and family). The 11 second involves studying the correlation between BVOC content and emission (i.e. how leaf content could be used to estimate 12 emissions). The latter strategy has provided controversial results, partly because BVOCs are amazingly chemically diverse, and, as 13 a result, techniques used to study plant BVOC content, which we review, cannot be equally adequate for all analytes. 14 In order to choose an adequate technique, two patterns must be distinguished. Specifically stored compounds -mainly 15 monoterpenes and sesquiterpenes that dominate the essential oil obtained from a plant -are permanently and massively present in 16 specific storage structures (e.g., secretory cavities, trichomes) of the order of lg/g-mg/g and usually allow emissions to occur 17 during stress periods when terpenes are weakly synthesized. These BVOCs can be studied directly through traditional extraction 18 techniques (e.g., hydrodistillation) and novel techniques (e.g., application of microwaves and ultrasound), and indirectly by 19 trapping techniques involving the collection, within adsorbent material, of BVOCs present in the headspace of a plant. 20

Background: Linalool is a widely used fragrance terpene. Pure linalool is not allergenic or a very weak allergen, but autoxidizes on air exposure and the oxidation products can cause contact allergy. Oxidized (ox.) linalool has previously been patch tested at a concentration of 2.0% in petrolatum (pet.) in 1511 patients, and 1.3% positive patch test reactions were observed.

Natural plant products such as essential oils have gained interest for use in pest control in place of synthetic pesticides because of their low environmental impact. Essential oils can be effective in controlling parasitic mites that infest honeybee colonies, but effective encapsulants are needed to provide a sustained and targeted delivery that minimizes the amount of active ingredient used. The present study reports the encapsulation of essential oils in porous microspheres that are within the size range of pollen grains and can be easily dispersed. The microspheres were made by pumping an 8% aqueous high-amylose starch gelatinous melt through an atomizing nozzle. The atomized starch droplets were air-classified into two fractions and collected in ethanol. The size range for each fraction was measured using a particle size analyzer. The mean particle size for the largest fraction was approximately 100 μm with a range from 5 μm to over 300 μm. Part of the reason for the large particle size was attributed to the merging of smaller particles that impinged upon each other before they solidified. The smaller fraction of spheres had a mean particle size of approximately 5 μm. The starchbased porous microspheres were loaded with 16.7% (w/w) essential oils including thymol (5-methyl-2isopropylphenol), clove, origanum, and camphor white oil. The essential oils appeared to be largely sequestered within the pore structure, since the spheres remained a free-flowing powder and exhibited little if any agglomeration in spite of the high loading rate. Furthermore, SEM micrographs verified that the pore structure was stable, as evidenced by the persistence of pores in spheres that had first been loaded with essential oils and then had the oil removed by solvent extraction. Thermal gravimetric analyses were consistent with a loading rate at predicted levels.

The successful air sampling and detection of cocaine, methylenedioxymethylamphetamine (MDMA), and marijuana using SPME-IMS achieved by targeting their volatile markers (methyl benzoate, piperonal, and terpenes, respectively) is presented. Conventional methods of direct air sampling for drugs are ineffective because the parent compounds of these drugs have very low vapor pressures, making them unavailable for headspace sampling. Instead of targeting the parent drugs, IMS was set at the optimal operating conditions (determined in previous work) in order to detect their volatile chemical markers. SPME is an effective and rapid air sampling technique for the preconcentration of analytes which is especially useful in confined spaces such as cargo containers, where the volatile marker compounds of drugs can be found in sufficient concentrations. By sampling the air using a 100 μm polydimethyl siloxane (PDMS) SPME fiber for as little as one minute, enough mass of the targeted volatile markers in the headspace of a quart-sized metal paint can (gallon, ∼1101 cm 3 ) which contained sub-gram quantities of the drug samples was recovered for IMS detection. Additionally, several potentially interfering compounds found in goods commonly shipped in cargo containers were tested individually as well as in mixtures with the drugs. No peak interferences were observed for MDMA or marijuana, and minimal peak interferences were found for cocaine.

The novel bromotriterpene polyethers aplysqualenol A (1) and B (2) have been isolated from the Caribbean sea slug Aplysia dactylomela, collected in Puerto Rico, and their structures and relative configurational assignments established from spectroscopic data aided by quantum mechanical calculations of NMR chemical shifts. Although both these compounds may be conceived as polyoxacyclic derivatives of the C 30 squalene skeleton, it is notable that 1 and 2 each

The aerial part of Artemisia arborescens L. (Asteraceae) was extracted with water and methanol, and both extracts were fractionated using n-hexane, chloroform, ethyl acetate and n-butanol. The potential phytotoxicity of both crude extracts and their fractions were assayed in vitro on seed germination and root growth of lettuce (Lactuca sativa L.), a sensible species largely employed in the allelopathy studies. The inhibitory activity was analyzed by dose-response curves and the ED 50 was estimated. Both crude extracts strongly inhibited either germination and root growth processes. The fraction-bioassay indicated the following hierarchy of phytotoxicity for both physiological processes: ethyl acetate ≥ n-hexane > chloroform ≥ n-butanol. On the n-hexane fraction, GC-MS analyses were carried out to characterize and quantify some of the potential allelochemicals. Twentyone compounds were identified and three of them, camphor, trans-caryophyllene and pulegone were quantified.

The aroma of sweet Fiano wine, the most representative non-aromatic white wine variety in southern Italy, was evaluated by sensory and instrumental analysis to determine the influence of grape overripeness, drying and Botrytis cinerea infection. Sensory descriptive analysis was used to evaluate the sensory properties of wines. Gas chromatography/mass spectrometry (GC/MS), gas chromatography (GC) and gas chromatography/olfactometry (GC/O) techniques were used to identify and to determine the content of free and bound volatile compounds and their odour impact. A sensory descriptive analysis had revealed that the sensory descriptors such as citrus jam, dried apricot, dried figs, prune, honey and coconut, occur in sweet Fiano wine more than in base Fiano wine. Thirty five free volatile compounds had higher levels in sweet Fiano wine than in base Fiano wine, these components were mostly terpenes, β-damascenone, lactones, aldehydes and ketones. Moreover higher levels of terpenes, β-damascenone, benzyl alcohol and 2-phenylethanol were found in the bound volatile fraction of sweet Fiano wine. The main odour impact compounds in sweet Fiano wine were nerol, geraniol and linalool (orange flowers), vitispirane (camphor), lactones such as γ-nonalactone (coconut), δ-decalactone and γ-decalactone (apricot) and 1-octen-3-ol (mushroom). These differences appear to be due to the fact that grape overripeness and the drying process allows concentration of varietal aromatic compounds and an easier transfer of them from skins to must during vinification.

This paper discusses the development of catalytic (Pt, Pd) hydrogenation in high-pressure CO 2 to convert limonene into valuable chemicals. It was noticed that varying the catalyst results in product regioselectivity. Platinum as catalyst favours the formation of p-menthane stereoisomers in equimolar quantities whereas palladium as catalyst furnishes trans-p-menthane and cis-p-menthane in 2:1 ratio. These results are in agreement with hydrogenation mechanisms for Pt-and Pd-catalysed reactions. Platinum is a more active catalyst than palladium, but higher activity results in lower chemical stability of the catalyst. Palladium as catalyst usually catalyses limonene isomerisation in the first stage of the process. Pressure tuning of the processes affects termination of the reaction. The flow rate of the reaction mixture through the stationary catalyst bed affects the composition of products; partially hydrogenated limonene is obtained at low flow rates and completely hydrogenated products at high flow rates.

The family Asteraceae comprises aromatic plants with a strong distinctive fragrance. Artemisia species belonging to this family are known to contain essential oils. Fresh aerial parts of Artemisia spicigera C. Koch and A. scoparia Waldst. et Kit. were subjected to headspace-SPME (HS-SPME) and the same plants were hydrodistilled (HD) after drying to obtain the profiles of volatiles. Headspace volatiles were analysed by GC–MS, whereas the essential oils were analysed by both GC and GC–MS. Camphor (37.5% and 43.9%) was found as the main constituent in both oils and in SPME samples of A. spicigera. However, in the oil sample of A. scoparia, capillene (1-phenylhexa-2,4-diyne) (53.1%) was found as the main constituent, while β-pinene (20.8%), β-caryophyllene (16.4%), (Z)-β-ocimene (16.4%), myrcene (12.8%), and limonene (11.0 %) were the main components in the headspace-SPME, respectively. Copyright © 2005 John Wiley & Sons, Ltd.

When attacked by insects, plants release mixtures of volatile compounds that are beneficial for direct or indirect defense. Natural variation of volatile emissions frequently occurs between and within plant species, but knowledge of the underlying molecular mechanisms is limited. We investigated intraspecific differences of volatile emissions induced from rosette leaves of 27 accessions of Arabidopsis (Arabidopsis thaliana) upon treatment with coronalon, a jasmonate mimic eliciting responses similar to those caused by insect feeding. Quantitative variation was found for the emission of the monoterpene (E)-b-ocimene, the sesquiterpene (E,E)-a-farnesene, the irregular homoterpene 4,8,12-trimethyltridecatetra-1,3,7,11-ene, and the benzenoid compound methyl salicylate. Differences in the relative emissions of (E)-b-ocimene and (E,E)-a-farnesene from accession Wassilewskija (Ws), a high-(E)-b-ocimene emitter, and accession Columbia (Col-0), a trace-(E)-b-ocimene emitter, were attributed to allelic variation of two closely related, tandem-duplicated terpene synthase genes, TPS02 and TPS03. The Ws genome contains a functional allele of TPS02 but not of TPS03, while the opposite is the case for Col-0. Recombinant proteins of the functional Ws TPS02 and Col-0 TPS03 genes both showed (E)-b-ocimene and (E,E)-a-farnesene synthase activities. However, differential subcellular compartmentalization of the two enzymes in plastids and the cytosol was found to be responsible for the ecotype-specific differences in (E)-b-ocimene/(E,E)-a-farnesene emission. Expression of the functional TPS02 and TPS03 alleles is induced in leaves by elicitor and insect treatment and occurs constitutively in floral tissues. Our studies show that both pseudogenization in the TPS family and subcellular segregation of functional TPS enzymes control the variation and plasticity of induced volatile emissions in wild plant species.

Despite the fact that Arabidopsis is largely self-pollinating, its flowers emit a complex mixture of terpene volatiles consisting predominantly of a large group of over 20 sesquiterpenes. Here we report that only two terpene synthases, encoded by the florally expressed genes At5g23960 and At5g44630, are responsible for the formation of virtually all sesquiterpenes found in the Arabidopsis floral volatile blend. Two independent mutant lines with T-DNA insertions in the previously identified At5g23960 gene lacked the emission of three sesquiterpenes, including the main sesquiterpene volatile (E)-b-caryophyllene, confirming the previous in vitro functional assignment for this gene. Flowers of a mutant line carrying a T-DNA insertion in gene At5g44630 emitted these three sesquiterpenes, but did not emit any of the remaining sesquiterpene volatiles. An At5g44630 cDNA was expressed in Escherichia coli and the produced protein catalyzed the conversion of farnesyl diphosphate into over 15 sesquiterpenes in similar proportions to those found in the floral volatile blend. At5g23960 and At5g44630 promoter-b-glucuronidase (GUS) fusion experiments demonstrated that both genes are expressed in several parts of the Arabidopsis flower, with strong At5g23960 promoter-GUS activity in the stigma and strong expression of At5g44630 in intrafloral nectaries. Given the previously reported antimicrobial activity of terpenes, their production in stigmas and nectaries may serve to inhibit microbial infection at these vulnerable sites. A survey of 37 Arabidopsis thaliana ecotypes revealed quantitative, but almost no qualitative, variations of floral monoterpene and sesquiterpene emissions suggesting that floral terpene volatiles must play some significant role in the life of the Arabidopsis plant.

. Tea tree oil, or the essential oil of Melaleuca alternfoolia, is becoming increasingly popular as a naturally occurring antimicrobial agent. T h e antimicrobial activity of eight components of tea tree oil was evaluated using disc diffusion and broth microdilution methods. Attempts were also made to overcome methodological problems encountered with testing compounds which have limited solubility in aqueous media. After assessing media with and without solubilizing agents, the disc diffusion method was used to determine the susceptibility of a range of microorganisms to 1,8-cineole, 1-terpinen-4-ol, p-cymene, linalool, a-terpinene, y-terpinene, a-terpineol and terpinolene. While the disc diffusion method lacked reproducibility, it was considered useful as a procedure for screening for antimicrobial activity. Terpinen-4-01 was active against all the test organisms while p-cymene demonstrated no antimicrobial activity. Linalool and a-terpineol were active against all organisms with the exception of Pseudomonas aeruginosa. Minimum inhibitory and minimum cidal concentrations of each component against Candida albicans, Escherichia coli and Staphylococcus aureus were determined using a broth microdilution method. Modifications to this method overcame solubility and turbidity problems associated with the oil components and allowed the antimicrobial activity of each of the components to be quantified reproducibly. There was reasonable agreement between minimum inhibitory concentrations and zones of inhibition. These results may have significant implications for the future development of tea tree oil as an antimicrobial agent.

The present article reports the antimicrobial efficacy of four monoterpenes (thymol, carvacrol, p-cymene, and γ-terpinene) against the Gram-positive bacterium Staphylococcus aureus and the Gram-negative bacterium Escherichia coli. For a better understanding of their mechanism of action, the damage caused by these four monoterpenes on biomembranes was evaluated by monitoring the release, following exposure to the compounds under study, of the water-soluble fluorescent marker carboxyfluorescein (CF) from large unilamellar vesicles (LUVs) with different lipidic composition (phosphatidylcholine, PC, phosphatidylcholine/phosphatidylserine, PC/PS, 9:1; phosphatidylcholine/ stearylamine, PC/SA, 9:1). Furthermore, the interaction of these terpenes with dimyristoylphosphatidylcholine multilamellar vesicles as model membranes was monitored by means of differential scanning calorimetry (DSC) technique. Finally, the results were related also with the relative lipophilicity and water solubility of the compounds examined. We observed that thymol is considerably more toxic against S. aureus than the other three terpenes, while carvacrol and p-cymene are the most inhibitory against E. coli. Thymol and carvacrol, but not γ-terpinene and p-cymene, caused a concentrationdependent CF leakage from all kinds of LUVs employed; in particular, thymol was more effective on PC and PC/SA LUVS than on PC/PS vesicles, while carvacrol challenge evoked a CF leakage from PC/PS LUVs similar to that induced from PC/SA LUVs, and lower than that measured with PC vesicles. Concerning DSC experiments, these four terpenes caused a decrease in T m and (especially carvacrol and p-cymene) ∆H values, very likely acting as substitutional impurities. Taken together, our findings lead us to speculate that the antimicrobial effect of thymol, carvacrol, p-cymene, and γ-terpinene may result, partially at least, from a gross perturbation of the lipidic fraction of the plasmic membrane of the microorganism. In addition to being related to the physicochemical characteristics of the compounds (such as lipophilicity and water solubility), this effect seems to be dependent on the lipidic composition and net surface charge of the microbic membranes. Furthermore, the compounds might cross the cell membranes, thus penetrating into the interior of the cell and interacting with intracellular sites critical for antibacterial activity.

Terpenes contribute to some white wines aroma, especially these produced from Muscat grapes and others aromatic ones of high terpene contents (Gewürtztramminer, Traminer, Huxel, Sylvaner). Terpenes are present in wine in free and bound (in a form of glycosides) forms. Analyses of bound terpenes are usually performed using solid phase extraction after hydrolysis of glycosides. A new method for determination of terpenes from wine, focused on determination of terpenes released after acidic hydrolysis, based on solid phase extraction (SPE) followed by solid phase microextraction (SPME) was developed. Non-polar (free) and polar (bound terpenes) fractions were separated on 500 mg C18 cartridges. Bound terpenes were sampled using SPME immediately after acidic hydrolysis in non-equilibrium conditions. Application of combined SPE-SPME approach allowed quantification of selected terpenes in lower concentrations than in SPE approach and added a selectivity to the method, which enabled detection of compounds nondetectable in SPE extracts. Results obtained by SPE and SPE-SPME approach were correlated for free terpenes and those released after acid hydrolysis 20 white wines obtained from different grape varieties (R 2 = 0.923). Although developed for wine terpenes analysis, SPE followed by SPME approach has a great potential in analysis of other bound wine flavor compounds, especially those potent odorants present in trace amounts.

Despite the fact that Arabidopsis is largely self-pollinating, its flowers emit a complex mixture of terpene volatiles consisting predominantly of a large group of over 20 sesquiterpenes. Here we report that only two terpene synthases, encoded by the florally expressed genes At5g23960 and At5g44630, are responsible for the formation of virtually all sesquiterpenes found in the Arabidopsis floral volatile blend. Two independent mutant lines with T-DNA insertions in the previously identified At5g23960 gene lacked the emission of three sesquiterpenes, including the main sesquiterpene volatile (E)-β-caryophyllene, confirming the previous in vitro functional assignment for this gene. Flowers of a mutant line carrying a T-DNA insertion in gene At5g44630 emitted these three sesquiterpenes, but did not emit any of the remaining sesquiterpene volatiles. An At5g44630 cDNA was expressed in Escherichia coli and the produced protein catalyzed the conversion of farnesyl diphosphate into over 15 sesquiterpenes in similar proportions to those found in the floral volatile blend. At5g23960 and At5g44630 promoter-β-glucuronidase (GUS) fusion experiments demonstrated that both genes are expressed in several parts of the Arabidopsis flower, with strong At5g23960 promoter-GUS activity in the stigma and strong expression of At5g44630 in intrafloral nectaries. Given the previously reported antimicrobial activity of terpenes, their production in stigmas and nectaries may serve to inhibit microbial infection at these vulnerable sites. A survey of 37 Arabidopsis thaliana ecotypes revealed quantitative, but almost no qualitative, variations of floral monoterpene and sesquiterpene emissions suggesting that floral terpene volatiles must play some significant role in the life of the Arabidopsis plant.

Mimusops caffra E. Mey. ex A.DC and Mimusops obtusifolia Lam (both members of the Sapotaceae family), and Hypoxis colchicifolia Bak (family Hypoxidaceae) are used by traditional healers in Zululand to manage malaria. Anti-plasmodial investigation of the crude extracts and some triterpenes isolated from the plants showed activity against a chloroquine sensitive (CQS) strain of Plasmodium falciparum (D10). Among the crude extracts the leaves of M. caffra exhibited the highest activity, with an IC 50 of 2.14 μg/mL. The pentacyclic tritepenoid ursolic acid (1), isolated from the leaves of M. caffra was the most active compound (IC 50 6.8 μg/mL) as compared to taraxerol (2) and sawamilletin (3) isolated from the stem bark of M. obtusifolia (IC 50 > 100). Chemical modification of the ursolic acid (1) to 3β-acetylursolic acid (4) greatly enhanced its anti-plasmodial activity. Compound 4 reduced parasitaemia against Plasmodium berghei by 94.01% in in vivo studies in mice. The cytotoxicity of 3β-acetylursolic acid (IC 50) to two human cell lines (HEK293 and HepG2) was 366.00 μg/mL and 566.09 μg/mL, respectively. The results validate the use of these plants in folk medicine.

HS-SPME analysis of terpenes does usually have inherent quantification problems when working with complex samples, especially due to the matrix effect of the substrate or the calibration solution. Three different terpene carrier matrices were compared: methanol, ...

This work was conducted to study a new separation and evaluation approach for the chemical fingerprinting of petroleum biomarkers in biota samples. The final aim of this work was to study the correlation between the observed effects in the shore habitats (mussels and limpets) and one pollution source: the oil spill of the Prestige tanker. The method combined a clean-up step of the biota extracts (mussels and limpets), the retention-time locking of the gas chromatographic set up, and the multivariate data analysis of the chromatograms. For clean-up, solid-phase extraction and gel permeation chromatography were compared, and 5 g Florisil cartridges assured the lack of interfering compounds in the last extracts. In order to assure reproducible retention times and to avoid the realignment of the chromatograms, the retention-time locking feature of our gas chromatography–mass spectrometry (GC–MS) set up was used. Finally, in the case of multivariate analysis, the GC–MS chromatograms were treated, essentially by derivatization and by normalization, and all the chromatograms at m/z 191 (terpenes), m/z 217–218 (steranes and diasteranes) and m/z 231 (triaromatic steranes) were treated by means of principal component analysis. Furthermore, slightly different four oil samples from the Prestige oil spill were analyzed following the Nordtest method, and the GC–MS chromatograms were considered as the reference chemical fingerprints of the sources. In this sense, the correlation between the studied samples, including sediments and biota samples, and the source candidate was completed by means of a supervised pattern recognition method. As a result, the method proposed in this work was useful to identify the Prestige oil spill as the source of many of the analyzed samples.

The allylic oxidation of ␣-pinene 1 was investigated using various catalytic systems. By a proper choice of the latter, the reaction can be directed toward the selective synthesis of verbenol 2 or verbenone 3. High yield into verbenone was achieved under mild conditions with copper salts as catalysts and t-butyl hydroperoxide (TBHP) as oxidant. On the other hand, when Pd(acac) 2 was used with hydrogen peroxide, verbenol was obtained as the main product.

Two new glycosides, the triterpenoid glycoside 19~-hydroxyurs-12-ene-24,28-dioate 3-O-fl-D-xylopyranoside and the phenol glycoside 2-isoprenyl-5-isopropylphenol 4-O-fl-D-xylopyranoside have been isolated from the whole plant of Blumea lacera and their structures elucidated by means of spectral and chemical studies.

A mixture of N-acetyl-cysteine + caVeic acid, at 10 + 30 mg/L respectively, was tested as inhibitor of the decrease of volatile esters and terpenes during storage of Debina white wine. Moreover, the inhibition of the decrease of isoamyl acetate, ethyl hexanoate and linalool in a model wine medium by the mixture, at 1.25 + 3.75 to 10 + 30 mg/L, was tested. Several volatiles, such as isoamyl acetate, ethyl hexanoate, ethyl octanoate, ethyl decanoate and linalool, decreased during wine storage. The mixture of N-acetyl-cysteine + caVeic acid restricted the decrease of these volatiles to a large extent. In the model medium, the mixture inhibited the decrease of the three volatiles in a dose dependent manner. It was active at any concentration tested. The present results indicate that mixture of N-acetyl-cysteine + caVeic acid, containing each compound at low concentration, may be taken into account as an eVective inhibitor of the disappearance of volatile aroma in wines.

Steam distillation (SD) extraction-solid-phase microextraction coupled to GC-MS was developed for the determination of terpenes and Bligh-Dyer extraction-derivatization coupled with GC for the determination of fatty acids such as ethyl esters were used. It was found that the three different Agave species have the same profile of fatty acids; the quantity of these compounds is different in each Agave variety. On the other hand, different terpenes were identified in the three Agave plants studied: nine in A. salmiana, eight in A. angustifolia and 32 in A. tequilana Weber var. azul.

In Morocco, the region covered with argan trees is named the argan grove. Its long-term preservation depends on the discovery of new and economically rewarding markets to sell argan tree produces. At the present time, the argan oil appears to be the best candidate to fulfill this task. The scientific results that have allowed the emergence of argan oil on the international edible and cosmetic oil markets are reported together with recent analytic results. Alternative approaches, not based on argan oil marketing but also aimed at safeguarding the argan grove, are also reported.

Volatile organic compounds (VOCs) emitted by damaged plants convey information to undamaged neighbouring plants, and previous research has shown that these signals are effective over short distances in nature. Many herbivore-induced VOCs react with ozone, which is the most important tropospheric air pollutant in rural areas. We used extrafloral nectar (EFN) secretion as a phenotypic indicator of between-plant communication in Phaseolus lunatus L. (Lima bean) and show that an ozone-rich (80 ppb) atmosphere reduces the distance over which signalling occurs. We found that ozone degrades several herbivore-induced VOCs, a likely mechanism reducing communication distances. Direct exposure to 80-ppb ozone did not affect the VOC emissions from P. lunatus. In addition, we demonstrated that high ozone concentrations, 120 and 160 ppb, induced EFN secretion in exposed plants, whereas more moderate concentrations, 80 and 100 ppb, did not. This suggests that ozone can play a complex role in the indirect defence of P. lunatus.