Identification of Salicylates in Willow Bark (Salix Cortex) for Targeting Peripheral Inflammation.

Salix cortex-containing medicine is used against pain conditions, fever, headaches, and inflammation, which are partly mediated via arachidonic acid-derived prostaglandins (PGs). We used an activity-guided fractionation strategy, followed by structure elucidation experiments using LC-MS/MS, CD-spectroscopy, and 1D/2D NMR techniques, to identify the compounds relevant for the inhibition of PGE2 release from activated human peripheral blood mononuclear cells. Subsequent compound purification by means of preparative and semipreparative HPLC revealed 2'-O-acetylsalicortin (1), 3'-O-acetylsalicortin (2), 2'-O-acetylsalicin (3), 2',6'-O-diacetylsalicortin (4), lasiandrin (5), tremulacin (6), and cinnamrutinose A (7). In contrast to 3 and 7, compounds 1, 2, 4, 5, and 6 showed inhibitory activity against PGE2 release with different potencies. Polyphenols were not relevant for the bioactivity of the Salix extract but salicylates, which degrade to, e.g., catechol, salicylic acid, salicin, and/or 1-hydroxy-6-oxo-2-cycohexenecarboxylate. Inflammation presents an important therapeutic target for pharmacological interventions; thus, the identification of relevant key drugs in Salix could provide new prospects for the improvement and standardization of existing clinical medicine.

Comparative Anti-Inflammatory Effects of Salix Cortex Extracts and Acetylsalicylic Acid in SARS-CoV-2 Peptide and LPS-Activated Human In Vitro Systems.

The usefulness of anti-inflammatory drugs as an adjunct therapy to improve outcomes in COVID-19 patients is intensely discussed in this paper. Willow bark (Salix cortex) has been used for centuries to relieve pain, inflammation, and fever. Its main active ingredient, salicin, is metabolized in the human body into salicylic acid, the precursor of the commonly used pain drug acetylsalicylic acid (ASA). Here, we report on the in vitro anti-inflammatory efficacy of two methanolic Salix extracts, standardized to phenolic compounds, in comparison to ASA in the context of a SARS-CoV-2 peptide challenge. Using SARS-CoV-2 peptide/IL-1ß- or LPS-activated human PBMCs and an inflammatory intestinal Caco-2/HT29-MTX co-culture, Salix extracts, and ASA concentration-dependently suppressed prostaglandin E2 (PGE2), a principal mediator of inflammation. The inhibition of COX-2 enzyme activity, but not protein expression was observed for ASA and one Salix extract. In activated PBMCs, the suppression of relevant cytokines (i.e., IL-6, IL-1ß, and IL-10) was seen for both Salix extracts. The anti-inflammatory capacity of Salix extracts was still retained after transepithelial passage and liver cell metabolism in an advanced co-culture model system consisting of intestinal Caco-2/HT29-MTX cells and differentiated hepatocyte-like HepaRG cells. Taken together, our in vitro data suggest that Salix extracts might present an additional anti-inflammatory treatment option in the context of SARS-CoV-2 peptides challenge; however, more confirmatory data are needed.

Management of the poultry red mite, Dermanyssus gallinae, using silica-based acaricides.

Four silica-based acaricides were examined in laboratory tests for their effectiveness against poultry red mite, Dermanyssus gallinae. All acaricides resulted in 100% mite mortality. Two groups of active ingredients could be differentiated. The products Silicosec® and Ewazid®, based on naturally occurring diatomaceous earth (DE), killed 100% of adult D. gallinae within 48 h exposure time. The time to kill 50% of the mites (LT50) was calculated to be 31.7 and 34.9 h, respectively. The other two products, containing aggregates and agglomerates of pyrogenic synthetic amorphous silicon dioxide as active ingredients, killed the mites in a significantly shorter time: LT50 was 6.3 h for the liquid product Fossil Shield® Instant White and 11.8 h for the powdery product Fossil Shield 90.0 White. This is more remarkable as the quantities of active ingredients used for the DE treatments were several folds higher. The effectiveness of all tested products was also shown in practical tests. A professional company treated five chicken houses on one farm in the Berlin-Brandenburg region with the test products, three houses with Fossil Shield Instant White and one each with Ewazid and Silicosec. Over a period of 46 weeks after stocking, the mite development in the houses was assessed. Only in one of the houses, treated with Fossil Shield Instant White, the mite population remained permanently low. In two houses treated with Fossil Shield Instant White, small mite colonies appeared in week 36, which were controlled by a follow-up spot treatment in week 41. In the houses treated with DE, the first mite colonies appeared 12 weeks after stocking. The number increased continuously over the experimental period and in week 31 after stocking there were clearly visible colonies (2-3 cm diameter) and the first mites could also be detected on the chicken eggs. At this time both houses were treated again with a follow-up spot-treatment, which only led to a slight improvement in one house and to a stabilization of the infestation in the other house. In week 41, large mite colonies were detected in both houses. A spot treatment at this point was ineffective in reducing the infestation. The tests showed faster acaricidal action of the products with the synthetic active ingredients compared to the natural DE-based products. This matches the shorter killing times under laboratory conditions. The experiments in a commercial chicken farm showed that it is possible to control the mite population for a period of 46 weeks by using physically effective SiO2-based products. These products are therefore an effective alternative to the use of chemical acaricides.

1-Methoxy-3-indolylmethyl DNA adducts in six tissues, and blood protein adducts, in mice under pak choi diet: time course and persistence.

We previously showed that purified 1-methoxy-3-indolylmethyl (1-MIM) glucosinolate, a secondary plant metabolite in Brassica species, is mutagenic in various in vitro systems and forms DNA and protein adducts in mouse models. In the present study, we administered 1-MIM glucosinolate in a natural matrix to mice, by feeding a diet containing pak choi powder and extract. Groups of animals were killed after 1, 2, 4 and 8 days of pak choi diet, directly or, in the case of the 8-day treatment, after 0, 8 and 16 days of recovery with pak choi-free diet. DNA adducts [N2-(1-MIM)-dG, N6-(1-MIM)-dA] in six tissues, as well as protein adducts [τN-(1-MIM)-His] in serum albumin (SA) and hemoglobin (Hb) were determined using UPLC-MS/MS with isotopically labeled internal standards. None of the samples from the 12 control animals under standard diet contained any 1-MIM adducts. All groups receiving pak choi diet showed DNA adducts in all six tissues (exception: lung of mice treated for a single day) as well as SA and Hb adducts. During the feeding period, all adduct levels continuously increased until day 8 (in the jejunum until day 4). During the 14-day recovery period, N2-(1-MIM)-dG in liver, kidney, lung, jejunum, cecum and colon decreased to 52, 41, 59, 11, 7 and 2%, respectively, of the peak level. The time course of N6-(1-MIM)-dA was similar. Immunohistochemical analyses indicated that cell turnover is a major mechanism of DNA adduct elimination in the intestine. In the same recovery period, protein adducts decreased more rapidly in SA than in Hb, to 0.7 and 37%, respectively, of the peak level, consistent with the differential turnover of these proteins. In conclusion, the pak choi diet lead to the formation of high levels of adducts in mice. Cell and protein turnover was a major mechanism of adduct elimination, at least in gut and blood.

Plant growth-promoting bacteria Kosakonia radicincitans mediate anti-herbivore defense in Arabidopsis thaliana.

MAIN CONCLUSION: This study demonstrates that the application of the PGPB strain, Kosakonia radicincitans enhances a plant's resistance against phloem-feeding and chewing insects in Arabidopsis thaliana. The plant growth-promoting bacterial strain K. radicincitans DSM 16656 applied to A. thaliana reduced the number of phloem-feeding insects of both the specialist Brevicoryne brassicae and the generalist Myzus persicae. While weight gain of the generalist chewing insect Spodoptera exigua was reduced by 30% on A. thaliana plants treated with K. radicincitans, growth of the specialist caterpillar Pieris brassicae was not affected when compared with caterpillars from control plants. Since generalist and specialist chewing insects responded differentially to PGPB application, the implication of signaling pathways in PGPB mediated changes in plant defense was studied using two signaling pathway mutants impaired in their salicylic acid (npr1-1 mutant) or jasmonic acid (coi1-1 mutant) pathway. We found that the jasmonic acid pathway is relevant for upregulation of aliphatic glucosinolates and suppression of the chewing generalist S. exigua larval growth. Chewing from generalist P. brassicae increased glucosinolate content in A. thaliana leaves mediated via both signaling pathways. However, only in the npr1-1 mutant, which contains the highest aliphatic glucosinolate content, the P. brassicae induced further enrichment of glucosinolates, resulting in a reduction of larval growth. Effects of K. radicincitans on plant resistance could not be explained by changes in glucosinolate contents or composition. Our results demonstrate the distinct role played by K. radicincitans in suppressing insect performance in A. thaliana.

Phyllotreta striolata flea beetles use host plant defense compounds to create their own glucosinolate-myrosinase system.

The ability of a specialized herbivore to overcome the chemical defense of a particular plant taxon not only makes it accessible as a food source but may also provide metabolites to be exploited for communication or chemical defense. Phyllotreta flea beetles are adapted to crucifer plants (Brassicales) that are defended by the glucosinolate-myrosinase system, the so-called "mustard-oil bomb." Tissue damage caused by insect feeding brings glucosinolates into contact with the plant enzyme myrosinase, which hydrolyzes them to form toxic compounds, such as isothiocyanates. However, we previously observed that Phyllotreta striolata beetles themselves produce volatile glucosinolate hydrolysis products. Here, we show that P. striolata adults selectively accumulate glucosinolates from their food plants to up to 1.75% of their body weight and express their own myrosinase. By combining proteomics and transcriptomics, a gene responsible for myrosinase activity in P. striolata was identified. The major substrates of the heterologously expressed myrosinase were aliphatic glucosinolates, which were hydrolyzed with at least fourfold higher efficiency than aromatic and indolic glucosinolates, and ß-O-glucosides. The identified beetle myrosinase belongs to the glycoside hydrolase family 1 and has up to 76% sequence similarity to other ß-glucosidases. Phylogenetic analyses suggest species-specific diversification of this gene family in insects and an independent evolution of the beetle myrosinase from other insect ß-glucosidases.

The Aggregation Pheromone of Phyllotreta striolata (Coleoptera: Chrysomelidae) Revisited.

Aggregations of the striped flea beetle Phyllotreta striolata on their crucifer host plants are mediated by volatiles emitted from feeding males. The male-specific sesquiterpene, (6R,7S)-himachala-9,11-diene (compound A), was shown previously to be physiologically and behaviorally active, but compound A was attractive only when combined with unnaturally high doses of the host plant volatile allyl isothiocyanate (AITC) in field trapping experiments. This indicated that our understanding of the chemical communication in this species is incomplete. Another male-specific sesquiterpenoid, (3S,9R,9aS)-3-hydroxy-3,5,5,9-tetramethyl-5,6,7,8,9,9a-hexahydro-1H-benzo[7]annulen-2(3H)-one (compound G), has been reported from an American P. striolata population. We confirmed the presence of compound G, and investigated its interaction with compound A and AITC in a P. striolata population in Taiwan. Compound G was attractive to Taiwanese P. striolata in laboratory bioassays, but significantly more beetles were attracted to a blend of compounds A and G. Under the same conditions, P. striolata showed no preference for the blend of A and G combined with a range of doses of AITC over the sesquiterpenoid blend alone. The sesquiterpenoid blend was tested further in field trapping experiments and attracted significantly more beetles than traps baited with compound A and ecologically relevant amounts of AITC. We conclude that A and G are components of the male-specific aggregation pheromone of P. striolata in Taiwan, and that the attractiveness of the pheromone is not reliant on the presence of AITC. Our results further indicate that the male-specific sesquiterpenoid blends differ qualitatively between the Taiwanese and American populations of P. striolata.

Pheromone Blend Analysis and Cross-Attraction among Populations of Maruca vitrata from Asia and West Africa.

The legume pod borer, Maruca vitrata, is a pantropical pest on leguminous crops. (E,E)-10,12-Hexadecadienal, (E,E)-10,12-hexadecadienol, and (E)-10-hexadecenal were described previously as sex pheromone components for this nocturnal moth. A blend of these components in a ratio of 100:5:5 attracted males in field trapping experiments in Benin, but not in Taiwan, Thailand, or Vietnam. This finding suggests geographic variation in the pheromone blend between Asian and West African populations of M. vitrata. We, therefore, determined the pheromone compositions of single pheromone glands of females from the three Asian regions and from Benin by gas chromatography-mass spectrometry. Additionally, we compared the responses of males from Taiwan and Benin to calling females and to gland extracts of females from both regions in laboratory no-choice and two-choice assays. Chemical analysis revealed the presence of (E,E)-10,12-hexadecadienal and (E,E)-10,12-hexadecadienol, as well as the absence of (E)-10-hexadecenal in all four populations. The relative amounts of the detected compounds did not vary significantly among the insect populations. The behavioral bioassays showed that Taiwanese and Beninese males were similarly attracted to females from both regions, as well as to their gland extracts. As a result, we did not find geographic variation in the sexual communication system of M. vitrata between West African and Asian insect populations.

A secondary metabolite of Brassicales, 1-methoxy-3-indolylmethyl glucosinolate, as well as its degradation product, 1-methoxy-3-indolylmethyl alcohol, forms DNA adducts in the mouse, but in varying tissues and cells.

1-Methoxy-3-indolylmethyl (1-MIM) glucosinolate, a secondary metabolite of Brassicales species, and its breakdown product 1-MIM alcohol are mutagenic in cells in culture after activation by plant ß-thioglucosidase and human sulphotransferase, respectively. In the present study, we administered these compounds orally to mice to study time course, dose dependence, tissue distribution and cellular localization of the 1-MIM DNA adducts formed. We used isotope-dilution ultra-performance liquid chromatography-tandem mass spectrometry to quantify the adducts and raised an antiserum for their immunohistochemical localization. Both compounds formed the same adducts, N(2)-(1-MIM)-2'-deoxyguanosine and N(6)-(1-MIM)-2'-deoxyadenosine, approximately in a 3.3:1 ratio. 1-MIM glucosinolate primarily formed these adducts in the large intestine, with a luminal-basal gradient, probably due to activation by thioglucosidase from intestinal bacteria. 1-MIM alcohol formed higher levels of adduct than the glucosinolate. Unlike after treatment with the glucosinolate, luminal and basal enterocytes were similarly affected in caecum, and liver and stomach were additional important target tissues. Maximal adduct levels were reached 8 h after the administration of both compounds. The hepatic DNA adducts persisted for the entire observation period (48 h), whereas those in large intestine rapidly declined due to cell turnover, as verified by immunohistochemistry. Hepatic adduct formation was focused on the periportal hepatocytes with concomitant depletion of glycogen, p53 activation and p21 induction. Adduct formation in caecum was associated with massive apoptosis, p53 activation and p21 induction, in particular after treatment with 1-MIM alcohol. It remains to be studied whether similar effects occur in humans after the consumption of Brassicales species.

Characterization, mode of action, and efficacy of twelve silica-based acaricides against poultry red mite (Dermanyssus gallinae) in vitro.

Poultry red mite infestation still is an unsolved problem in poultry farms. Legal regulations, residue risks, and resistances limit chemical control of mites. Alternatives to chemical acaricides for control of poultry red mite are silica-based products, which have as a main constituent silicon dioxide. The acaricidal effect is attributed to sorptive properties of the particles, which result in the mite's death by desiccation. In the present study, the acaricidal efficacy of 12 products containing natural or synthetic silica, 9 in powder form, and 3 for liquid application was tested under laboratory conditions. Mite mortality was measured at several intervals and the mean lethal time (LT50) determined by Probit analysis after Abbott's correction. The LT50 values of the products significantly differed (Tukey's HSD p < 0.05). LT50 values of powdery formulations ranged from 5.1 to 18.7 h and overlapped with those of the fluid ones which ranged from 5.5 to 12.7 h. In order to explain the differences in efficacy of the tested silica products, further characterizations were carried out. X-ray fluorescence, specific surface, cation exchange capacity (CEC), and water absorption capacity (WAC) were measured. Furthermore, electron microscopy was conducted and different products compared. Silicon dioxide content (ranging from 65 to 89% for powders and 57 to 80% for fluids) had no significant impact on efficacy, while specific surface and CEC (2.4-23.2 mEq 100(-1) g(-1) for powders and 18-30.8 mEq 100(-1) g(-1)) were positively and WAC (1.3-4.4 wt% for powders and 3.3-4.8 wt% for fluids) negatively related to the acaricidal efficacy. Influence of these parameters on acaricidal efficacy was significant according to the results of a stepwise regression analysis (p < 0.01).

Arbuscular mycorrhizal fungi affect glucosinolate and mineral element composition in leaves of Moringa oleifera.

Moringa is a mycorrhizal crop cultivated in the tropics and subtropics and appreciated for its nutritive and health-promoting value. As well as improving plant mineral nutrition, arbuscular mycorrhizal fungi (AMF) can affect plant synthesis of compounds bioactive against chronic diseases in humans. Rhizophagus intraradices and Funneliformis mosseae were used in a full factorial experiment to investigate the impact of AMF on the accumulation of glucosinolates, flavonoids, phenolic acids, carotenoids, and mineral elements in moringa leaves. Levels of glucosinolates were enhanced, flavonoids and phenolic acids were not affected, levels of carotenoids (including provitamin A) were species-specifically reduced, and mineral elements were affected differently, with only Cu and Zn being increased by the AMF. This study presents novel results on AMF effects on glucosinolates in leaves and supports conclusions that the impacts of these fungi on microelement concentrations in edible plants are species dependent. The nonspecific positive effects on glucosinolates and the species-specific negative effects on carotenoids encourage research on other AMF species to achieve general benefits on bioactive compounds in moringa.

Impact of the PGPB Enterobacter radicincitans DSM 16656 on growth, glucosinolate profile, and immune responses of Arabidopsis thaliana.

Plant growth-promoting bacteria (PGPB) affect plant cellular processes in various ways. The endophytic bacterial strain Enterobacter radicincitans DSM 16656 has been shown to improve plant growth and yield in various agricultural and vegetable crops. Besides its ability to fix atmospheric nitrogen, produce phytohormones, and solubilize phosphate compounds, the strain is highly competitive against native endophytic organisms and colonizes the endorhizosphere in high numbers. Here, we show that E. radicincitans inoculation of the noncrop plant Arabidopsis thaliana promotes plant growth. Furthermore, high performance liquid chromatography (HPLC) analysis revealed that bacterial inoculation slightly decreased amounts of aliphatic glucosinolates in plant leaves in a fast-growing stage but increased these compounds in an older phase where growth is mostly completed. This effect seems to correlate with developmental stage and depends on the nitrogen requirement. Additionally, nitrogen deficiency studies with seedlings grown on medium containing different nitrogen concentrations suggest that plant nitrogen demand can influence the intensity of plant growth enhancement by E. radicincitans. This endophyte seems not to activate stress-inducible mitogen-activated protein kinases (MAPKs). Analyzing transcription of the defense-related genes PR1, PR2, PR5, and PDF1.2 by quantitative real time polymerase chain reaction (qPCR) revealed that E. radicincitans DSM 16656 is able to induce priming via salicylic acid (SA) or jasmonate (JA)/ethylene (ET) signaling pathways to protect plants against potential pathogen attack.

Determination of benzyl isothiocyanate metabolites in human plasma and urine by LC-ESI-MS/MS after ingestion of nasturtium (Tropaeolum majus L.).

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated to determine the concentration of benzyl isothiocyanate (BITC) metabolites in human plasma and urine. In this study, the following BITC metabolites have been considered: BITC-glutathione, BITC-cysteinylglycine, BITC-cysteine, and BITC-N-acetyl-L-cysteine. The assay development included: (1) synthesis of BITC conjugates acting as reference substances; (2) sample preparation based on protein precipitation and solid-phase extraction; (3) development of a quantitative LC-MS/MS method working in the multiple-reaction monitoring mode; (4) validation of the assay; (5) investigation of the stability and the reactivity of BITC conjugates in vitro; (6) application of the method to samples from a human intervention study. The lower limits of quantification were in the range of 21-183 nM depending on analyte and matrix, whereas the average recovery rates from spiked plasma and urine were approximately 85 and 75 %, respectively. BITC conjugates were found to be not stable in alkaline buffered solutions. After consumption of nasturtium, containing 1,000 µM glucotropaeolin, the primary source of BITC, quantifiable levels of BITC-NAC, BITC-Cys, and BITC-CysGly were found in human urine samples. Maximum levels in urine were determined 4 h after the ingestion of nasturtium. With regard to the human plasma samples, all metabolites were determined including individual distributions. The work presented provides a validated LC-MS/MS method for the determination of BITC metabolites and its successful application for the analysis of samples collected in a human intervention study.

Hemp Waste as a Substrate for Hermetia illucens (L.) (Diptera: Stratiomyidae) and Tenebrio molitor L. (Coleoptera: Tenebrionidae) Rearing.

The proper treatment of cannabis agricultural wastes can reduce the environmental impact of its cultivation and generate valuable products. This study aimed to test the potential of cannabis agricultural wastes as a substrate for the rearing of black soldier fly larvae (BSFL) and yellow mealworms (MW). In the case of BSFL, replacing the fibre component (straw) in the substrate with the hemp waste can increase the nutritional value of the substrate and led to bigger larvae. The bigger larvae had lower P and Mg, and higher Fe and Ca. Crude protein also varied based on the size of larvae and/or the content of protein in the initial substrate, which was boosted by replacing straw with hemp material. No other cannabinoids than cannabidiolic acid (CBDA), cannabigerolic acid (CBGA), and cannabidiol (CBD) were found in significant amounts in the larvae. In the case of MW, the larvae grew less on the hemp material in comparison to wheat bran. Replacing wheat bran with the hemp material led to smaller larvae with higher Ca, Fe, K, and crude protein content, but lower Mg and P values. No cannabinoids were detected in the MW fed with the hemp material.

Physical Properties of Substrates as a Driver for Hermetia illucens (L.) (Diptera: Stratiomyidae) Larvae Growth.

The growth and nutritional profile of the black soldier fly larvae (BSFL) is usually investigated and compared when the larvae feed on substrates that differ in the chemical composition as well as physical properties. This study compares BSFL growth on substrates that differ primarily in physical properties. This was achieved by using various fibres in the substrates. In the first experiment, two substrates with 20% or 14% chicken feed were mixed with three fibres (cellulose, lignocellulose, or straw). In the second experiment, the growth of BSFL was compared with a 17% chicken feed substrate that additionally contained straw with different particle sizes. We show that the substrate texture properties values did not influence the BSFL growth, but the bulk density of the fibre component did. The substrate mixed with cellulose led to higher larvae growth over time in comparison to substrates with higher bulk density fibres. BSFL grown on the substrate mixed with cellulose reached their maximum weight in 6 days instead of 7. Neither the fibres nor the nutrient level changed the crude protein content of BSFL and the values ranged between 33.5% and 38.3%, but an interaction between the fibre and nutrient level was observed. The size of straw particles in the substrates influenced the BSFL growth and led to a 26.78% difference in Ca concentration, a 12.04% difference in Mg concentration, and a 35.34% difference in P concentration. Our findings indicate that the BSFL-rearing substrates can be optimised by changing the fibre component or its particle size. This can improve the survival rate, reduce the cultivation time needed to reach the maximum weight, and alter the chemical composition of BSFL.

UV-B irradiation changes specifically the secondary metabolite profile in broccoli sprouts: induced signaling overlaps with defense response to biotic stressors.

Only a few environmental factors have such a pronounced effect on plant growth and development as ultraviolet light (UV). Concerns have arisen due to increased UV-B radiation reaching the Earth's surface as a result of stratospheric ozone depletion. Ecologically relevant low to moderate UV-B doses (0.3-1 kJ m(-2) d(-1)) were applied to sprouts of the important vegetable crop Brassica oleracea var. italica (broccoli), and eco-physiological responses such as accumulation of non-volatile secondary metabolites were related to transcriptional responses with Agilent One-Color Gene Expression Microarray analysis using the 2×204 k format Brassica microarray. UV-B radiation effects have usually been linked to increases in phenolic compounds. As expected, the flavonoids kaempferol and quercetin accumulated in broccoli sprouts (the aerial part of the seedlings) 24 h after UV-B treatment. A new finding is the specific UV-B-mediated induction of glucosinolates (GS), especially of 4-methylsulfinylbutyl GS and 4-methoxy-indol-3-ylmethyl GS, while carotenoids and Chl levels remained unaffected. Accumulation of defensive GS metabolites was accompanied by increased expression of genes associated with salicylate and jasmonic acid signaling defense pathways and up-regulation of genes responsive to fungal and bacterial pathogens. Concomitantly, plant pre-exposure to moderate UV-B doses had negative effects on the performance of the caterpillar Pieris brassicae (L.) and on the population growth of the aphid Myzus persicae (Sulzer). Moreover, insect-specific induction of GS in broccoli sprouts was affected by UV-B pre-treatment.

Male Phyllotreta striolata (F.) produce an aggregation pheromone: identification of male-specific compounds and interaction with host plant volatiles.

The chrysomelid beetle Phyllotreta striolata is an important pest of Brassicaceae in Southeast Asia and North America. Here, we identified the aggregation pheromone of a population of P. striolata from Taiwan, and host plant volatiles that interact with the pheromone. Volatiles emitted by feeding male P. striolata attracted males and females in the field. Headspace volatile analyses revealed that six sesquiterpenes were emitted specifically by feeding males. Only one of these, however, elicited an electrophysiological response from antennae of both sexes. A number of host plant volatiles, e.g., 1-hexanol, (Z)-3-hexen-1-ol, and the glucosinolate hydrolysis products allyl isothiocyanate (AITC), 3-butenyl isothiocyanate, and 4-pentenyl isothiocyanate also elicited clear responses from the antenna. The active male-specific compound was identified as (+)-(6R,7S)-himachala-9,11-diene by chiral stationary phase gas-chromatography with coupled mass spectrometry, and by comparison with reference samples from Abies nordmanniana, which is known to produce the corresponding enantiomer. The pheromone compound was synthesized starting from (-)-α-himachalene isolated from Cedrus atlantica. Under field conditions, the activity of the synthetic pheromone required concomitant presence of the host plant volatile allyl isothiocyanate. However, both synthetic (+)-(6R,7S)-himachala-9,11-diene alone and in combination with AITC were attractive in a two-choice laboratory assay devoid of other natural olfactory stimuli. We hypothesize that P. striolata adults respond to the pheromone only if specific host volatiles are present. In the same laboratory set up, more beetles were attracted by feeding males than by the synthetic stimuli. Thus, further research will be necessary to reveal the components of a more complex blend of host or male-produced semiochemicals that might enhance trap attractiveness in the field.

Drug-Drug Interaction Potential, Cytotoxicity, and Reactive Oxygen Species Production of Salix Cortex Extracts Using Human Hepatocyte-Like HepaRG Cells.

Herbal preparations of willow bark (Salix cortex) are available in many countries as non-prescription medicines for pain and inflammation, and also as dietary supplements. Currently only little information on toxicity and drug interaction potential of the extracts is available. This study now evaluated the effects of two Salix cortex extracts on human hepatocyte-like HepaRG cells, in view of clinically relevant CYP450 enzyme activity modulation, cytotoxicity and production of reactive oxygen species (ROS). Drug metabolism via the CYP450 enzyme system is considered an important parameter for the occurrence of drug-drug interactions, which can lead to toxicity, decreased pharmacological activity, and adverse drug reactions. We evaluated two different bark extracts standardized to 10 mg/ml phenolic content. Herein, extract S6 (S. pentandra, containing 8.15 mg/ml total salicylates and 0.08 mg/ml salicin) and extract B (industrial reference, containing 5.35 mg/ml total salicylates and 2.26 mg/ml salicin) were tested. Both Salix cortex extracts showed no relevant reduction in cell viability or increase in ROS production in hepatocyte-like HepaRG cells. However, they reduced CYP1A2 and CYP3A4 enzyme activity after 48 h at ≥25 µg/ml, this was statistically significant only for S6. CYP2C19 activity inhibition (0.5 h) was also observed at ≥25 µg/ml, mRNA expression inhibition by 48 h treatment with S6 at 25 µg/ml. In conclusion, at higher concentrations, the tested Salix cortex extracts showed a drug interaction potential, but with different potency. Given the high prevalence of polypharmacy, particularly in the elderly with chronic pain, further systematic studies of Salix species of medical interest should be conducted in the future to more accurately determine the risk of potential drug interactions.

Chemoprofiling as Breeding Tool for Pharmaceutical Use of Salix.

Willow bark is traditionally used for pharmaceutical purposes. Evaluation is so far based on the salicylate content, however, health promoting effects of extracts might be attributed to the interaction of those salicylates with other compounds, which support and complement their action. So far, only S. purpurea, S. daphnoides, and S. fragilis are included in pharmaceutical extracts. Crossing with other species could result in a more diverse secondary metabolite profile with higher pharmacological value. With the help of targeted inter- and intraspecific crossing, new chemotypes were generated, whereby nine different Salix genotypes (S. alba, S. daphnoides, S. humboldtiana, S. lasiandra, S. nigra, S. pentandra, S. purpurea, S. x rubens, S. viminalis) were included in the study. Based on substances known for their health promoting potential and characteristic for Salix (selected phenolic compounds including salicylates), a targeted metabolomics analysis and clustering of 92 generated Salix clones was performed revealing four different cluster/chemoprofiles. In more specific, one group is formed by S. daphnoides clones and inter- and intraspecific hybrids, a second group by S. viminalis clones and inter- and intraspecific hybrids, a third group generally formed by S. alba, S. pentandra, S. x rubens, and S. lasiandra clones and hybrids, and a fourth group by S. purpurea clones and inter- and intraspecific hybrids. Clustering on the basis of the selected phenolic compounds can be used for identifying Salix clones with a different compound profile. New combinations of secondary plant metabolites offer the chance to identify Salix crosses with improved effects on human health.

Comparison of different silicas of natural origin as possible insecticides.

Many insect-pests have developed resistances to pesticides. Therefore, there is always a need for new plant protection substances. For example the physically active insecticide diatomaceous earth (DE) gained much attention as an alternative insecticide in stored products. DE is a naturally occurring silica, which acts by destroying the insect's cuticle by absorbing the protective wax layer. This results in body water loss and ultimately the insect's death by desiccation. The silica-based materials tested were the commercial DE product Fossil Shield 90.0s, Advasan, and a formulation newly developed by the Urban Horticultural Department at Humboldt University, called Al-06. The trials were performed in small covered plastic boxes. Test substances were either dusted onto the surface of the boxes (E. vigintioctopunctata, S. litura) or mixed into rice medium (S. oryzae). The mortality was observed after 1, 2, 4, 7, 14, and 28 days. Untreated insects served as control. The first test series showed that some AL-06-formulations and FS90.0s were very effective against adults of S. oryzae and S. litura and larvae of E. vigintioctopunctata. For adult Epilachna beetles, we could not detect any differences between the treatments. The highest mortality rate in S. oryzae trials occurred with FS90.0s (100%) after 21 days. The same efficiency was achieved after 2 days with some AL-06 formulations against S. litura and E. vigintioctopunctata. The results of this study indicate that silica dusts can effectively control insect pests from different orders. At higher dosages, all materials resulted in higher insect mortality rates. It was also found that some substances did not perform well under higher rel. humidity; therefore, the conclusion was drawn that hydrophilic substances saturate with water from the surrounding air and lose their insecticidal efficacy. Earlier studies have proven that particles with a larger surface area are more effective than particles with smaller surfaces. As a result, the most effective substances in the field trials were the ones containing the small particles, since there is a larger surface area available to interact with the insects' cuticles. Further studies will be conducted to analyse the relevance of water saturation of substances in order to examine their effectiveness under greenhouse conditions. Greenhouse experiments are generally considered to study practicability of silica dusts in horticulture. Perhaps the silica dusts will show phytotoxic side effects.