Jenny Fäldt

Volatiles of bracket fungi Fomitopsis pinicola and Fomes fomentarius and their functions as insect attractants

Volatiles released from fruiting bodies of the polypores Fomitopsis pinicola and Fomes fomentarius (Polyporaceae) were collected by entrainment and SPME techniques and analyzed by GC-MS. The most significant difference between the two species was found in the terpene fraction. F. fomentarius emitted a more complex blend, with more than 10 sesquiterpenes. During the sporulating phase, the release of (R)- and (S)-oct-1-en-3-ol, octan-3-one, and some sesquiterpene hydrocarbons (mainly β-barbatene) increased in F. pinicola, whereas in F. fomentarius the release of octan-3-one, cis-furanoid linalool oxide, β-phellandrene, β-myrcene, and several sesquiterpene hydrocarbons increased. β-Barbatene was identified for the first time in fungi. Chopping of the fruiting bodies altered the odor composition more in F. fomentarius than in F. pinicola. Five volatiles giving a typical fungal odor (rac-oct-1-en-3-ol, nonan-1-ol, rac-octan-3-ol, octan-1-ol, and octan-3-one) were tested for insect attraction in the field. Females of the three wood-living generalist beetles Malthodes fuscus, Anaspis marginicollis, and A. rufilabris and both sexes of the moth Epinotia tedella were attracted to rac-oct-1-en-3-ol. The generalist predator on fungus-insects Lordithon lunulatus was attracted to rac-oct-1-en-3-ol and octan-3-one in combination. Previous results regarding the host-specific attraction of beetles to the odors emanating from chopped fruiting bodies of F. pinicola and F. fomentarius are discussed in the light of this investigation.

Enzymatic properties of native and deglycosylated hybrid aspen (Populus tremula×tremuloides) xyloglucan endotransglycosylase 16A expressed in Pichia pastoris

The cDNA encoding a xyloglucan endotransglycosylase, PttXET16A, from hybrid aspen (Populus tremulaxtremuloides) has been isolated from an expressed sequence tag library and expressed in the methylotrophic yeast Pichia pastoris. Sequence analysis indicated a high degree of similarity with other proteins in the XTH (xyloglucan transglycosylase/hydrolase) gene subfamily of GH16 (glycoside hydrolase family 16). In addition to the conserved GH16 catalytic sequence motif, PttXET16A contains a conserved N-glycosylation site situated proximal to the predicted catalytic residues. MS analysis indicated that the recombinant PttXET16A expressed in P. pastoris is heterogeneous due to the presence of variable N-glycosylation and incomplete cleavage of the alpha-factor secretion signal peptide. Removal of the N-glycan by endoglycosidase H treatment did not influence the catalytic activity significantly. Similarly, site-directed mutagenesis of Asn93 to serine to remove the N-glycosylation site resulted in an enzyme which was comparable with the wild-type enzyme in specific activity and thermal stability but had clearly reduced solubility. Hydrolytic activity was detected neither in wild-type PttXET16A before or after enzymatic deglycosylation nor in PttXET16A N93S (Asn93-->Ser) mutant.

Determination of hydrocarbons in old creosote contaminated soil using headspace solid phase microextraction and GC–MS

Headspace solid phase microextraction (HS-SPME) has been used together with GC-MS to analyze organic substances directly in a soil, heavily contaminated with PAHs/creosote (approximately 300 mg/kg soil), from an old gaswork site in Stockholm, Sweden. The HS-SPME results, both qualitative and quantitative, were compared with traditional liquid extraction using ethyl acetate/hexane (20:80). It was shown that the concentrations determined with HS-SPME at 60 degrees C correlated well, for compounds containing up to two and three aromatic rings (naphthalenes, acenaphthene, acenaphthylene and fluorenes, while a lower concentration was obtained for phenanthrene, anthracene, fluoranthene and pyrene. The total concentrations for each compound determined with HS-SPME ranged from 2 to 25 microg/g soil. Quantification was done using standard addition of compounds directly to the soil samples. The bioavailable fraction of the compounds in the contaminated soil at 20 degrees C was analyzed using external calibration by spiking sterile uncontaminated sand (same texture and particle size as the contaminated soil but without a heavily sorbed organic fraction) with hydrocarbon standards in different concentrations. Storage of exposed fibers at 20 degrees C showed that analysis should be done within two days to make qualitative measurements and earlier (as soon as possible) for quantitative determinations.

Occurrence and correlations of monoterpene hydrocarbon enantiomers in Pinus sylvestris and Picea abies.

The relative amounts and enantiomeric compositions of monoterpene hydrocarbons in branch and trunk xylem, in needles, and in resin from apical buds in 18 Pinus sylvestris trees have been determined and compared with the terpene content in branch xylem and needles of Picea abies. Besides the high amount of (+)-3-carene, an excess of (+)-α-pinene has been found in P. sylvestris, whereas in P. abies (−)-α-pinene dominates over (+)-α-pinene. In P. sylvestris, clear positive correlations were found between (+)-α-pinene and (+)-camphene in the four tissues analyzed. Good positive correlations were also observed between (−)-α-pinene and (−)-camphene in the two types of xylem, between (+)-α-pinene and (+)-β-pinene in the resin, and between tricyclene and (−)-camphene in resin and needles. In P. abies, positive correlations were found between (+)-α-pinene and (+)-camphene in the branch xylem and between tricyclene and (−)-camphene as well as between (−)-α-pinene and (−)-camphene in the needles. Complex relationships between (−)-α-pinene and (−)-β-pinene were found both in the P. abies and in the P. sylvestris tissues. The importance of the enantiomeric composition of α-pinene for the host selection of Ips typographus, Tomicus piniperda, and Hylobius abietis is discussed.

Biological degradation of diesel fuel in water and soil monitored with solid-phase micro-extraction and GC-MS

Abstract Solid-phase micro-extraction (SPME) was used for monitoring degradation of hydrocarbons in diesel-fuel-contaminated (1% v/v) water and soil. Natural soil bacteria with and without external addition of inoculum were used. Directly after a 10-s exposure of the sample, the polydimethylsiloxane fibre was injected into the GC-MS. This method strongly reduced the time of analysis compared to a conventional liquid/liquid extraction. A comparison of SPME and pentane extraction of diesel oil was made and found to be consistent. The degradation of diesel fuel in water was monitored for 10 weeks using SPME. After 5 weeks all hydrocarbons were degraded except for the decahydronaphthalenes. These compounds were approximately 3% of the total hydrocarbons in the diesel oil used and remained undegraded throughout the study although none of the chemical or physical parameters was limiting. In the soil study the degradation of diesel fuel in normal soil was completed after 3 weeks, when the only remaining substances were decahydronaphthalenes. All samples were compared to sterile references to make up for evaporation losses. SPME proved to be a fast and reliable method to monitor changes in concentrations of semi-volatile organic compounds.

Correlations between selected monoterpene hydrocarbons in the xylem of six Pinus (pinaceae) species

Summary. The compositions of 23 monoterpene hydrocarbons of six pine species (Pinus sylvestris,P. yunnanensis, P. armandii, P. tropicalis, P. cubensis and P. caribaea) were compared, using multivariate data analysis. Four of the six species were clearly different from the other species in a Principal Components Analysis (PCA) model, based on the relative amounts (selective normalization) of the monoterpenes. The correlation coefficients between constituents were determined separately for each species and the strongest correlations were found between (+)-α-pinene and (+)-camphene and between the corresponding (—)-enantiomers, in all species. This pattern, i.e. a good correlation in all species, was neither shown by the correlation of the structurally more similar (+)-α/β-pinenes, nor by the (—)-α/β-pinenes or within the enantiomeric pairs of α-pinene and β-pinene. For these pairs of monoterpenes, good correlations were found in some species. None of the species showed good correlations in all the investigated monoterpene pairs presented here. Correlations between monoterpenes in insect-attacked trees (P. cubensis and P. caribaea, attacked by Dioryctria horneana, and P. yunnanensis, attacked by Tomicus piniperda) were also determined. The results are discussed from chemotaxonomic and biosynthetic points of view.

Comparison of headspace techniques for sampling volatile natural products in a dynamic system.

Abstract Commonly used dynamic sorption techniques for collecting biologically active volatile compounds have been compared. Solid phase microextraction (SPME) using two types of fibers (polydimethylsiloxane, PDMS, 100 μm, and carbowax/divinylbenzene, CW/DVB, 65 μm) was compared to purge and trap methods (Porapak Q, Tenax TA and charcoal) and a technique based on absorption in methanol in a cooling bath. Sampling was done in a stream of purified air (20 ml/min) in a closed and temperature-regulated (27 °C) glass tube, passing over a capillary tube containing a hexane solution of tridecane, heptadecane, 1-octen-3-ol, 1- hexadecanol, ethyl tetradecanoate, α-pinene, linalool, terpinen-4-ol, cis-verbenol, verbenone, β-caryophyllene, E,E-farnesol, and geranylgeraniol. With all of the methods, the sampling was performed for a period of 30 min before extraction and analysis was done on a GC-FID system. In general, SPME gave a higher response for all compounds except for a-pinene, which was only extracted by the CW/DVB fiber. Purge and trap methods and methanol absorption gave the same response for all substances extracted. None of the methods extracted hexadecanol and geranylgeraniol under the conditions used. However, the SPME equipped with the PDMS coating extracted heptadecane, E,E-farnesol and ethyl tetradecanoate. Our results show that SPME, when selecting the fibers to fit the polarity and volatility of the compounds, is an outstanding extraction method compared to purge and trap and methanol absorption, especially for a qualitative analysis. The best conditions for storing fibers exposed to compounds of high volatility were at low temperatures (6 °C) in sealed vials, while the worst way was to leave the exposed fiber unprotected at room temperature (22 °C). The dynamic sampling system was effectively tested on a fruiting body of a polypore fungus ( Ganoderma applanatum) emitting 1-octen-3-ol, and again SPME showed to be the most sensitive technique.

Host selection in Tomicus piniperda L.: composition of monoterpene hydrocarbons in relation to attack frequency in the shoot feeding phase.

The aim of this study was to investigate the host selection capacity of the pine shoot beetle, Tomicus piniperda, in the shoot-feeding phase and analyze the chiral and non-chiral host volatiles by means of GC-MS and 2D-GC in five Pinus species originating from France (Pinus sylvestris, P. halepensis, P. nigra laricio, P. pinaster maritima, P. pinaster mesogeensis). Dominating monoterpenes were (-)-α-pinene, (+)-α-pinene, (-)-β-pinene and (+)-3-carene. The amounts of the enantiomers varied considerably within and among the species. In a principal component analysis-plot, based on the absolute amounts of 18 monoterpene hydrocarbons, separation of the pine species into two groups was obtained. P. halepensis and P. sylvestris were grouped according to the amount of (+)-α-pinene and (+)-3-carene, while P. nigra laricio, P. pinaster maritima and P. pinaster mesogeensis were grouped according to (-)-α-pinene and (D)-β-pinene. P. nigra laricio was the species most attacked and P. halepensis the one least attacked by T. piniperda.