Combined perceptual and chemical analyses show no compelling evidence for ovulatory cycle shifts in women's axillary odour.

Although men's attraction to women's body odour has been suggested to vary over the ovulatory cycle, peaking around the fertile window, we still lack methodologically robust evidence corroborating this effect. Further, the chemical underpinnings of male preference for the odour of ovulating women remain unknown. Here, we combined perceptual and chemical analyses to investigate the axillary odour of naturally cycling women over 10 days, covering the gradual change in fertility across the ovulatory cycle with a focus on fertile days. The fertile state was confirmed by urinary ovulation tests as well as salivary oestradiol and progesterone levels. Men rated the scent of unfamiliar women, resembling a first encounter. We used multivariate analyses to relate variation in both odour ratings and chemical composition to female conception probability, temporal distance to ovulation and ovarian hormone levels. Our results provide no evidence that males prefer the odour of fertile women. Furthermore, the volatile analysis indicated no link between axillary odour composition and current fertility status. Together, our results showed no convincing support for a chemical fertility cue in women's axillary odour, questioning the presence of olfactory fertility information that is recognizable during first encounters in modern humans.

Trends and applications in plant volatile sampling and analysis.

Volatile organic compounds (VOCs) released by plants serve as information and defense chemicals in mutualistic and antagonistic interactions and mitigate effects of abiotic stress. Passive and dynamic sampling techniques combined with gas chromatography-mass spectrometry analysis have become routine tools to measure emissions of VOCs and determine their various functions. More recently, knowledge of the roles of plant VOCs in the aboveground environment has led to the exploration of similar functions in the soil and rhizosphere. Moreover, VOC patterns have been recognized as sensitive and time-dependent markers of biotic and abiotic stress. This focused review addresses these developments by presenting recent progress in VOC sampling and analysis. We show advances in the use of small, inexpensive sampling devices and describe methods to monitor plant VOC emissions in the belowground environment. We further address latest trends in real-time measurements of volatilomes in plant phenotyping and most recent developments of small portable devices and VOC sensors for non-invasive VOC fingerprinting of plant disease. These technologies allow for innovative approaches to study plant VOC biology and application in agriculture.

Optimized Solid-Phase Mesh-Enhanced Sorption from Headspace (SPMESH) for Rapid Sub-ng/kg Measurements of 3-Isobutyl-2-methoxypyrazine (IBMP) in Grapes.

Parallel extraction of headspace volatiles from multiwell plates using sorbent sheets (HS-SPMESH) followed by direct analysis in real-time high-resolution mass spectrometry (DART-HRMS) can be used as a rapid alternative to solid-phase micro-extraction (SPME) gas-chromatography mass-spectrometry (GC-MS) for trace level volatile analyses. However, an earlier validation study of SPMESH-DART-MS using 3-isobutyl-2-methoxypyrazine (IBMP) in grape juice showed poor correlation between SPMESH-DART-MS and a gold standard SPME-GC-MS around the compound's odor detection threshold (<10 ng/kg) in grape juice, and lacked sufficient sensitivity to detect IBMP at this concentration in grape homogenate. In this work, we report on the development and validation of an improved SPMESH extraction approach that lowers the limit of detection (LOD < 0.5 ng/kg), and regulates crosstalk between wells (<0.5%) over a calibration range of 0.5−100 ng/kg. The optimized SPMESH-DART-MS method was validated using Cabernet Sauvignon and Merlot grape samples harvested from commercial vineyards in the central valley of California (n = 302) and achieved good correlation and agreement with SPME-GC-MS (R2 = 0.84) over the native range of IBMP (<0.5−20 ng/kg). Coupling of SPMESH to a lower resolution triple quadrupole (QqQ)-MS via a new JumpShot-HTS DART source also achieved low ng/kg detection limits, and throughput was improved through positioning stage optimizations which reduced time spent on intra-well SPMESH areas.

Non-invasive quantification of ethylene in attached fruit headspace at 1 p.p.b. by gas chromatography-mass spectrometry.

Ethylene is a gaseous plant hormone involved in defense, adaptations to environmental stress and fruit ripening. Its relevance to the latter makes its detection highly useful for physiologists interested in the onset of ripening. Produced as a sharp peak during the respiratory burst, ethylene is biologically active at tens of nl L-1 . Reliable quantification at such concentrations generally requires specialized instrumentation. Here we present a rapid, high-sensitivity method for detecting ethylene in attached fruit using a conventional gas chromatography-mass spectrometry (GC-MS) system and in situ headspace collection chambers. We apply this method to melon (Cucumis melo L.), a unique species consisting of climacteric and non-climacteric varieties, with a high variation in the climacteric phenotype among climacteric types. Using a population of recombinant inbred lines (RILs) derived from highly climacteric ('Védrantais', cantalupensis type) and non-climacteric ('Piel de Sapo', inodorus type) parental lines, we observed a significant variation for the intensity, onset and duration of the ethylene burst during fruit ripening. Our method does not require concentration, sampling times over 1 h or fruit harvest. We achieved a limit of detection of 0.41 ± 0.04 nl L-1 and a limit of quantification of 1.37 ± 0.13 nl L-1 with an analysis time per sample of 2.6 min. Validation of the analytical method indicated that linearity (>98%), precision (coefficient of variation ≤2%) and sensitivity compared favorably with dedicated optical sensors. This study adds to evidence of the characteristic climacteric ethylene burst as a complex trait whose intensity in our RIL population lies along a continuum in addition to two extremes.

Volatile and non-volatile compounds in green tea affected in harvesting time and their correlation to consumer preference.

Current study was designed to find out how tea harvesting time affects the volatile and non-volatile compounds profiles of green tea. In addition, correlation of instrumental volatile and non-volatile compounds analyses to consumer perception were analyzed. Overall, earlier harvested green tea had stronger antioxidant capacity (~61.0%) due to the polyphenolic compounds from catechin (23,164 mg/L), in comparison to later harvested green teas (11,961 mg/L). However, high catechin content in green tea influenced negatively the consumer likings of green tea, due to high bitterness (27.6%) and astringency (13.4%). Volatile compounds drive consumer liking of green tea products were also identified, that included linalool, 2,3-methyl butanal, 2-heptanone, (E,E)-3,5-Octadien-2-one. Finding from current study are useful for green tea industry as it provide the difference in physiochemical properties of green tea harvested at different intervals.

Decolorization of Cheddar cheese whey by activated carbon.

Colored Cheddar whey is a source for whey protein recovery and is decolorized conventionally by bleaching, which affects whey protein quality. Two activated carbons were studied in the present work as physical means of removing annatto (norbixin) in Cheddar cheese whey. The color and residual norbixin content of Cheddar whey were reduced by a higher level of activated carbon at a higher temperature between 25 and 55°C and a longer time. Activated carbon applied at 40g/L for 2h at 30°C was more effective than bleaching by 500mg/L of hydrogen peroxide at 68°C. The lowered temperature in activated-carbon treatments had less effect on protein structure as investigated for fluorescence spectroscopy and volatile compounds, particularly oxidation products, based on gas chromatography-mass spectrometry. Activated carbon was also reusable, removing more than 50% norbixin even after 10 times of regeneration, which showed great potential for decolorizing cheese whey.

Correlating wine quality indicators to chemical and sensory measurements.

Twenty-seven commercial Californian Cabernet Sauvignon wines of different quality categories were analyzed with sensory and chemical methods. Correlations between five quality proxies-points awarded during a wine competition, wine expert scores, retail price, vintage, and wine region-were correlated to sensory attributes, volatile compounds, and elemental composition. Wine quality is a multi-faceted construct, incorporating many different layers. Depending on the quality proxy studied, significant correlations between quality and attributes, volatiles and elements were found, some of them previously reported in the literature.

Influence of the Drying Process on the Volatile Profile of Different Capsicum Species.

Chili is a globally significant spice used fresh or dried for culinary, condiment, and medicinal purposes. Growing concerns about food safety have increased the demand for high-quality products and non-invasive tools for quality control like origin tracing and safety assurance. Volatile analysis offers a rapid, comprehensive, and safe method for characterizing various food products. Thus, this study aims to assess the impact of the drying process on the aromatic composition of various Capsicum species and to identify key compounds driving the aromatic complexity of each genetic makeup. To accomplish these objectives, the aroma was examined in fruits collected from 19 different pepper accessions (Capsicum sp.) belonging to four species: one ancestral (C. chacoense) and three domesticated pepper species (C. annuum, C. baccatum and C. chinense). Fresh and dried samples were analyzed using a headspace PTR-TOF-MS platform. Our findings reveal significant changes in the composition and concentration of volatile organic compounds (VOCs) from fresh to dried Capsicum. Notably, chili peppers of the species C. chinense consistently exhibited higher emission intensity and a more complex aroma compared to other species (both fresh and dried). Overall, the data clearly demonstrate that the drying process generally leads to a reduction in the intensity and complexity of the aromatic compounds emitted. Specifically, fresh peppers showed higher volatile organic compounds content compared to dried ones, except for the two sweet peppers studied, which exhibited the opposite behavior. Our analysis underscores the variability in the effect of drying on volatile compound composition among different pepper species and even among different cultivars, highlighting key compounds that could facilitate species classification in dried powder. This research serves as a preliminary guide for promoting the utilization of various pepper species and cultivars as powder, enhancing product valorization.

Impact of feeding regimes and lactation stage on sensory attributes of Cheddar cheese.

This study investigated the effects of diet and stage of lactation (SOL) on sensory profiles, texture, volatile profiles, and colour of Cheddar cheese. Cheddar cheese was manufactured from early-, mid-, and late-lactation milk obtained from seasonally calved cows (n = 54). Cows were assigned a diet; group 1: perennial ryegrass (GRS), group 2: total mixed ration (TMR), and group 3: partial mixed ration (PMR). Instrumental analysis was performed at 270 days (mature Cheddar). Sensory evaluation took place after 548 days (extra mature Cheddar). Toluene was the only volatile compound that was significantly influenced by diet. The trained panel rated early-lactation cheese as stronger than mid- and late- for cowy/barny flavour and late-lactation cheese as sweeter than early- and mid-lactation cheese. Mid-lactation cheese was liked least overall. Early-lactation cheeses were rated higher for 'crumbly' texture than mid- and late. Diet affected consumer ratings, with GRS and PMR cheese rated as more intense than TMR for flavour, aftertaste, and saltiness. Consumers reported that TMR cheese was lighter in colour compared to GRS cheese, which was supported by instrumental analysis. Consumers perceived GRS as more springy and less crumbly than TMR and PMR, while Texture Profile Analysis indicated that TMR was harder than GRS. Consumer segmentation was observed with two clear preference groups, one preferring GRS and one preferring TMR. For both groups, 'taste' seemed to be the main driver of liking, highlighting that consumer preference is most impacted by individual taste preferences.

Unravelling the origin of isoprene in the human body-a forty year Odyssey.

In the breath research community's search for volatile organic compounds that can act as non-invasive biomarkers for various diseases, hundreds of endogenous volatiles have been discovered. Whilst these systemic chemicals result from normal and abnormal metabolic activities or pathological disorders, to date very few are of any use for the development of clinical breath tests that could be used for disease diagnosis or to monitor therapeutic treatments. The reasons for this lack of application are manifold and complex, and these complications either limit or ultimately inhibit the analytical application of endogenous volatiles for use in the medical sciences. One such complication is a lack of knowledge on the biological origins of the endogenous volatiles. A major exception to this is isoprene. Since 1984, i.e. for 40 years, it has been generally accepted that the pathway to the production of human isoprene, and hence the origin of isoprene in exhaled breath, is through cholesterol biosynthesis via the mevalonate (MVA) pathway within the liver. However, various studies between 2001 and 2012 provide compelling evidence that human isoprene is produced in skeletal muscle tissue. A recent multi-omic investigation of genes and metabolites has revealed that this proposal is correct by showing that human isoprene predominantly results from muscular lipolytic cholesterol metabolism. Despite the overwhelming proof for a muscular pathway to isoprene production in the human body, breath research papers still reference the hepatic MVA pathway. The major aim of this perspective is to review the evidence that leads to a correct interpretation for the origins of human isoprene, so that the major pathway to human isoprene production is understood and appropriately disseminated. This is important, because an accurate attribution to the endogenous origins of isoprene is needed if exhaled isoprene levels are to be correctly interpreted and for assessing isoprene as a clinical biomarker.

The Impact of Drying and Rehydration on the Structural Properties and Quality Attributes of Pre-Cooked Dried Beans.

Fresh common beans can be made 'instant' to produce fast-cooking beans by first soaking and cooking the beans before drying to create a shelf-stable product that can be rehydrated at the time of use. This study investigated the interplay between the drying process (air, vacuum and freeze drying), the microstructure and functional attributes of rehydrated pre-cooked beans. The microscopic study revealed that the three different drying techniques resulted in distinctly different microstructures, with the freeze drying process resulting in highly porous materials, while the air- and vacuum-dried samples underwent shrinkage. Additionally, the rehydration behavior (modeled using empirical and diffusion models) demonstrates that the high rehydration rate of freeze-dried beans is due to capillarity, while rehydration, in the case of air- and vacuum-dried beans, is primarily diffusion-controlled. Irrespective of the drying technique, the high rehydration capacity supports little to no structural collapse or damage to the cell walls. The color and texture of the rehydrated beans did not differ greatly from those of freshly cooked beans. The total peak area of the volatiles of rehydrated beans was significantly reduced by the drying process, but volatiles characteristic of the cooked bean aroma were retained. This new understanding is beneficial in tailoring the functional properties of pre-cooked dry convenient beans requiring short preparation times.

Rapid Analysis of Volatile Phenols from Grape Juice by Immersive Sorbent Sheet Extraction Prior to Direct Analysis in Real-Time Mass Spectrometry (DART-MS).

Poly(dimethylsiloxane)-based thin-film sorbent sheets (SPMESH) have previously been used for parallel headspace (HS) extraction prior to direct analysis in real-time mass spectrometry (DART-MS) for rapid quantitation of odorants in complex matrices. However, HS-SPMESH extraction is poorly suited for less volatile odorants, e.g., volatile phenols. This report describes modifications to the previous SPMESH extraction device, which make it amenable to parallel extraction of low-volatility analytes from multiwell plates under direct immersion (DI) conditions. Optimization and validation of the DI-SPMESH-DART-MS approach were performed on four volatile phenols (4-ethylphenol, 4-ethylguaiacol, 4-methylguaiacol, and guaiacol) of relevance to the quality of grape juices. Negative-ion mode DART-MS spectra showed a series of oxygenated adducts [M + nO - H]- for all analytes, but isobaric interferences could be limited for three of the four analytes by selecting an appropriate MS/MS transition. Signal suppression from nonvolatiles (sugars, acids) could be overcome by a rinse step. DI-SPMESH-DART-MS analysis of 24 samples could be performed in ∼45 min (30 min extraction, 16 min DART analysis) with 0.5-3 µg/L detection limits in aqueous and model juice solutions. In real grape juices (n = 5 cultivars), good accuracy (72-137%) could be achieved for two of the four volatile phenols initially investigated, 4-ethylphenol and 4-ethylguaiacol. However, poor accuracy was observed for guaiacol in some cultivars, and 4-methylguaiacol could not be quantitated due to interferences with other volatile phenols. Despite these limitations, DI-SPMESH-DART-MS/MS may be useful for prescreening a large number of samples prior to more selective conventional analyses.

Analysis of volatile compounds in rooibos tea (Aspalathus linearis) using different extraction methods and their relationship with human sensory perception.

This study aimed to comparatively analyze the volatile flavor of rooibos tea (Aspalathus linearis) obtained by two commonly used flavor extraction methods, simultaneous distillation-extraction (SDE) and steam distillation under reduced pressure (DRP). The tea obtained by the two extraction methods, were analyzed by gas chromatography-mass spectrometry to identify volatile aroma-related compounds. Descriptive sensory analysis of the extracted rooibos tea flavor was carried out by a trained panel (n = 7). Fifty volatile compounds were identified, including 26 and 25 aroma-active compounds by SDE (45.9 µg/g) and DRP (37.5 µg/g), respectively. SDE recovered larger quantities of alcohols, acids, and esters, whereas DRP was useful for analyzing thermally unstable volatile compounds, including various alcohols, aldehydes, and hydrocarbons. Descriptive sensory analysis revealed that ketones and phenolic compounds may be responsible for the sensory attributes woody and grassy green, whereas the aldehydes and acidic compounds may contribute to floral and fruity.

A cross-cultural sensory analysis of skim powdered milk produced from pasture and non-pasture diets.

Understanding potential cross-cultural sensory differences in the perception of Irish dairy products is important for key markets such as the USA and China. As most Irish dairy products are produced from pasture derived milk, this study investigated the impact of pasture and non-pasture diets on the cross cultural sensory perception of skim milk powder (SMP) in Ireland, USA and China. SMP was produced from cows fed outdoors on ryegrass (GRS), ryegrass/white clover (CLV), and indoors on trial mixed rations (TMR). SMP samples were evaluated by Irish (n = 78), USA (n = 100) and Chinese (n = 106) consumers using an identical hedonic sensory acceptance test in Ireland, USA and China. Optimized Descriptive Profiling (ODP) was performed using trained assessors familiar with dairy products in Ireland (n = 25) and China (n = 22), and traditional descriptive analysis was undertaken by a trained panel (n = 7) in the USA. Volatile analysis was undertaken on each SMP sample. Hedonic assessment found that USA consumers preferred SMP derived from TMR, and Irish consumers preferred SMP from either GRS or CLV. Chinese consumers perceived SMP samples differently to the USA and Irish consumers, but preference was not influenced by diet. Both Irish and Chinese trained assessors found it more difficult to discern differences between GRS or CLV SMP, but could differentiate TMR SMP. Irish assessors preferred GRS and CLV SMP. Chinese and Irish assessors had different preferences for many attributes. Trained USA panelists found significant differences, exclusively associating pasture based diets with "cowy/barny" and "cardboard/wet paper" attributes and more intense "grassy/hay" attributes than in TMR SMP. The abundance of ten volatile compounds differed significantly based on diet with acetoin derived from carbohydrate metabolism at much greater abundance in TMR SMP. This study found that sensory perception and volatile profiles of SMP were influenced by diet and differences in sensory perception existed between the three cultural groups. Irish and USA sensory responses aligned with familiarity of dairy products derived from pasture and non-pasture diets, respectively, and Chinese sensory responses differed to Irish and USA responses likely reflecting their lack of familiarity with dairy products.

Polymeric Sorbent Sheets Coupled to Direct Analysis in Real Time Mass Spectrometry for Trace-Level Volatile Analysis-A Multi-Vineyard Evaluation Study.

Etched polymeric sorbent sheets (solid-phase mesh-enhanced sorption from headspace (SPMESH) sheets) were recently described as an alternative to solid-phase microextraction (SPME) for rapid, parallel, multi-sample extraction and pre-concentration of headspace volatiles. In this report, a workflow was evaluated based on SPMESH sheet extraction followed by direct analysis in real time-mass spectrometry (DART-MS) using grape samples harvested from multiple commercial vineyards at different maturities. SPMESH sheet-DART-MS(-MS) was performed on two grape-derived odorants related to wine quality: 3-isobutyl-2-methoxypyrazine (IBMP) in Cabernet Sauvignon and Merlot grape homogenate (n = 86 samples) and linalool in Muscat-type grape juice samples (n = 18 samples). As part of the optimization process, an MS-MS method was developed for IBMP and an equilibration procedure prior to extraction was established for homogenate samples. Following optimization, we achieved good correlation between SPMESH sheet-DART-MS and SPME-GC-MS for both IBMP (range by GC-MS = < 2 ng/L to 28 ng/L, R2 = 0.70) and linalool (range by GC-MS = 135 to 415 µg/L, R2 = 0.66). The results indicate SPMESH sheet-DART-MS is suitable for rapid measurements of trace level volatiles in grapes.

Application of a Novel Instantized Glycerol Monooleate Ingredient in a Protein-Stabilized Oil-In-Water Emulsion.

Glycerol monooleate (GMO), casein and whey proteins are surfactants that can stabilize emulsion systems. This study investigates the impact of instantized GMO powders on creaming stability and oxidative stability in protein-stabilized emulsions. Model emulsions with bulk GMO, two instantized GMO powders, and two controls (without GMO) were produced by microfluidization. The droplet size, ζ-potential, viscosity, and creaming index of the emulsions were measured, while oxidative stability was evaluated by analysis of volatile compounds during storage (28 days, 45 °C) using gas chromatography mass spectrometry. Emulsions with GMO produced smaller average droplet sizes (180.0 nm) with a narrower distribution (polydispersity index of 0.161) compared to the controls (197.6 nm, 0.194). The emulsion stability of instantized emulsions was as good as bulk GMO, which were both better than controls. Based on the relative abundance of 3-octen-2-one, 2,4-heptadienal isomer 2, and 3,5-octadien-2-one isomer 1, the oxidative stability of the instantized emulsions was not significantly different from controls; however, bulk GMO emulsion showed significantly lower stability than controls. Instantized GMO powders can successfully produce physically stable protein-stabilized emulsions with good oxidative stability in a convenient powdered format.

Spatially Resolved Headspace Extractions of Trace-Level Volatiles from Planar Surfaces for High-Throughput Quantitation and Mass Spectral Imaging.

The use of headspace thin-film microextraction devices (SPMESH) for parallel extraction of trace-level volatiles prior to direct analysis in real-time mass spectrometry (DART-MS) has been reported previously, in which volatiles were extracted from samples in multi-well plates. In this report, we demonstrate that headspace extraction of volatiles by SPMESH sheets can be performed directly from planar surfaces. When coupled with DART-MS, this approach yields volatile mass spectral images with at least 4 mm resolution. When samples were spotted onto general-purpose silica gel thin-layer chromatography (TLC) plates, the SPMESH extraction could reach equilibrium within 2-4 min and 48 samples could be extracted and analyzed in 14 min. Because volatilization of analytes from TLC plates was very rapid, SPMESH extraction was delayed by the addition of 5% polyethylene glycol. Good linearity was achieved in the microgram per liter to milligram per liter range for four odorants (3-isobutyl-2-methoxypyrazine, linalool, methyl anthranilate, and o-aminoacetophenone) in several matrices (water, 10% ethanol, juice, and grape macerate) using 5 µL sample sizes. Detection limits as low as 50 pg/spot (10 µg/L in grape macerate) could be achieved. In contrast to many reports on headspace solid-phase microextraction, negligible matrix effects were observed for ethanol and grape macerates compared to water. SPMESH can preserve volatile images from planar surfaces, and SPMESH-DART-MS from TLC plates is well-suited for rapid trace volatile analysis, especially with small sample sizes.

Instrumental volatile flavor analysis of omija (Schisandra chinesis Baillon) using headspace stir-bar sorptive extraction-gas chromatography-mass spectrometry and its relationship to human sensory perceptions.

Omija (Schisandra chinesis Baillon) is a cluster of berries, mainly cultivated in East Asian countries. The objective of this study was to conduct an instrumental volatile flavor analysis and descriptive analysis of omija samples using headspace stir-bar sorptive extraction gas chromatography-mass spectrometry. Three omija fruits were included and pre-treated using three different methods: frozen omija (FO), frozen-blended omija (FBO), and freeze-dried omija (FDO). A total of 28 volatiles were identified, and significant differences in concentrations found among omija samples were noted, especially for previously identified aroma-active compounds such as α-pinene, α-terpinene, and (E)-ß-ocimene (p < .05). In terms of sensory perception, significant differences were observed in three sensory attributes that included, ginger, pine needle, and wet grassy aromatics among the samples (p < 0.05). While similarities and dissimilarities were observed among the three samples in the instrumental flavor analysis results, a principal component analysis revealed that FBO and FDO shared similar flavor characteristics while FO had distinctive flavor characteristics.

Nerolidol production in agroinfiltrated tobacco: Impact of protein stability and membrane targeting of strawberry (Fragraria ananassa) NEROLIDOL SYNTHASE1.

The sesquiterpene alcohol nerolidol, synthesized from farnesyl diphosphate (FDP), mediates plant-insect interactions across multiple trophic levels with major implications for pest management in agriculture. We compared nerolidol engineering strategies in tobacco using agroinfiltration to transiently express strawberry (Fragraria ananassa) linalool/nerolidol synthase (FaNES1) either at the endoplasmic reticulum (ER) or in the cytosol as a soluble protein. Using solid phase microextraction and gas chromatography-mass spectrometry (SPME-GCMS), we have determined that FaNES1 directed to the ER via fusion to the transmembrane domain of squalene synthase or hydroxymethylglutaryl - CoA reductase displayed significant improvements in terms of transcript levels, protein accumulation, and volatile production when compared to its cytosolic form. However, the highest levels of nerolidol production were observed when FaNES1 was fused to GFP and expressed in the cytosol. This SPME-GCMS method afforded a limit of detection and quantification of 1.54 and 5.13 pg, respectively. Nerolidol production levels, which ranged from 0.5 to 3.0 µg/g F.W., correlated more strongly to the accumulation of recombinant protein than transcript level, the former being highest in FaNES-GFP transfected plants. These results indicate that while the ER may represent an enriched source of FDP that can be exploited in metabolic engineering, protein accumulation is a better predictor of sesquiterpene production.

A subset of chemosensory genes differs between two populations of a specialized leaf beetle after host plant shift.

Due to its fundamental role in shaping host selection behavior, we have analyzed the chemosensory repertoire of Chrysomela lapponica. This specialized leaf beetle evolved distinct populations which shifted from the ancestral host plant, willow (Salix sp., Salicaceae), to birch (Betula rotundifolia, Betulaceae). We identified 114 chemosensory candidate genes in adult C. lapponica: 41 olfactory receptors (ORs), eight gustatory receptors, 17 ionotropic receptors, four sensory neuron membrane proteins, 32 odorant binding proteins (OBPs), and 12 chemosensory proteins (CSP) by RNA-seq. Differential expression analyses in the antennae revealed significant upregulation of one minus-C OBP (Clap OBP27) and one CSP (Clap CSP12) in the willow feeders. In contrast, one OR (Clap OR17), four minus-C OBPs (Clap OBP02, 07, 13, 20), and one plus-C OBP (Clap OBP32) were significantly upregulated in birch feeders. The differential expression pattern in the legs was more complex. To narrow down putative ligands acting as cues for host discrimination, the relative abundance and diversity of volatiles of the two host plant species were analyzed. In addition to salicylaldehyde (willow-specific), both plant species differed mainly in their emission rate of terpenoids such as (E,E)-α-farnesene (high in willow) or 4,8-dimethylnona-1,3,7-triene (high in birch). Qualitatively, the volatiles were similar between willow and birch leaves constituting an "olfactory bridge" for the beetles. Subsequent structural modeling of the three most differentially expressed OBPs and docking studies using 22 host volatiles indicated that ligands bind with varying affinity. We suggest that the evolution of particularly minus-C OBPs and ORs in C. lapponica facilitated its host plant shift via chemosensation of the phytochemicals from birch as novel host plant.