Encapsulation of Fennel and Basil Essential Oils in ß-Cyclodextrin for Novel Biopesticide Formulation.

ß-cyclodextrin (ß-CD) is a good host for the encapsulation of fennel and basil essential oils (FEO and BEO, respectively) and the formation of inclusion complexes (ICs) using the co-precipitation method. According to the results of the GC/MS analysis conducted in this study, monoterpenes and monoterpenoids were the dominant chemical groups in total FEO, while in BEO, these two groups occurred along with sesquiterpenes and sesquiterpenoids. The presence of dominant compounds from both EOs was validated using the FT-IR spectra of ICs, which indicated successful complexation. Analyses conducted using SPME/GC-MS showed the continuous emission of volatiles over 24 h from both ICs. Under SEM, particles of both ICs appeared to have a rectangular or rhomboid morphology and few aggregates. The insecticidal properties of EOs and ICs with ß-CD were tested on the Colorado potato beetle (CPB) as a model pest. The inclusion complex of ß-CD with FEO altered the developmental dynamic and body mass of the CPB. The initial increase in the proteolytic activity of CPB larvae fed with potato plants sprayed with ICs was not maintained for long, and the proteolytic efficacy of treated larvae remained in line with that of the control larvae. Future investigations will focus on manipulating the volume of EOs used and the treatment duration for optimal efficacy and potential application.

Solanum dulcamara L. Berries: A Convenient Model System to Study Redox Processes in Relation to Fruit Ripening.

The present study provides, for the first time, a physicochemical and biochemical characterization of the redox processes associated with the ripening of Solanum dulcamara L. (bittersweet) berries. Electron Paramagnetic Resonance Spectroscopy (EPRS) and Imaging (EPRI) measurements of reactive oxygen species (ROS) were performed in parallel with the tissue-specific metabolic profiling of major antioxidants and assessment of antioxidant enzymes activity. Fruit transition from the mature green (MG) to ripe red (RR) stage involved changes in the qualitative and quantitative content of antioxidants and the associated cellular oxidation and peroxidation processes. The skin of bittersweet berries, which was the major source of antioxidants, exhibited the highest antioxidant potential against DPPH radicals and nitroxyl spin probe 3CP. The efficient enzymatic antioxidant system played a critical protective role against the deleterious effects of progressive oxidative stress during ripening. Here, we present the EPRI methodology to assess the redox status of fruits and to discriminate between the redox states of different tissues. Interestingly, the intracellular reoxidation of cell-permeable nitroxide probe 3CP was observed for the first time in fruits or any other plant tissue, and its intensity is herein proposed as a reliable indicator of oxidative stress during ripening. The described noninvasive EPRI technique has the potential to have broader application in the study of redox processes associated with the development, senescence, and postharvest storage of fruits, as well as other circumstances in which oxidative stress is implicated.

Comprehensive Phytohormone Profiling of Kohlrabi during In Vitro Growth and Regeneration: The Interplay with Cytokinin and Sucrose.

The establishment of an efficient protocol for in vitro growth and regeneration of kohlrabi (Brassica oleracea var. gongylodes) allowed us to closely examine the phytohormone profiles of kohlrabi seedlings at four growth stages (T1-T4), additionally including the effects of cytokinins (CKs)-trans-zeatin (transZ) and thidiazuron (TDZ)-and high sucrose concentrations (6% and 9%). Resulting phytohormone profiles showed complex time-course patterns. At the T2 stage of control kohlrabi plantlets (with two emerged true leaves), levels of endogenous CK free bases and gibberellin GA20 increased, while increases in jasmonic acid (JA), JA-isoleucine (JA-Ile), indole-3-acetic acid (IAA) and indole-3-acetamide (IAM) peaked later, at T3. At the same time, the content of most of the analyzed IAA metabolites decreased. Supplementing growth media with CK induced de novo formation of shoots, while both CK and sucrose treatments caused important changes in most of the phytohormone groups at each developmental stage, compared to control. Principal component analysis (PCA) showed that sucrose treatment, especially at 9%, had a stronger effect on the content of endogenous hormones than CK treatments. Correlation analysis showed that the dynamic balance between the levels of certain bioactive phytohormone forms and some of their metabolites could be lost or reversed at particular growth stages and under certain CK or sucrose treatments, with correlation values changing between strongly positive and strongly negative. Our results indicate that the kohlrabi phytohormonome is a highly dynamic system that changes greatly along the developmental time scale and also during de novo shoot formation, depending on exogenous factors such as the presence of growth regulators and different sucrose concentrations in the growth media, and that it interacts intensively with these factors to facilitate certain responses.

UHPLC-OrbiTrap MS Characterization of Phenolic Profiles in French Marigold Extracts and Analysis of Their Antifeedant Activity against Colorado Potato Beetle.

French marigold is an aromatic plant rich in polyphenolic secondary metabolites, which pesticidal potential was examined in this study. Ultra-high-performance liquid chromatography (UHPLC) connected with OrbiTrap mass spectrometer (MS) identified 113 phenolics and revealed the most detailed phytochemistry of French marigold published so far. Depending on plant material (flowers or leaves) and solvents used for extraction (water, methanol, dichloromethane), the phenolic composition varied. Methanol extract of flowers, with 89 identified phenolics and high antioxidant activity statistically comparable with positive control Trolox, was chosen for testing of antifeedant potential against the 3rd and 4th instars of Colorado potato beetle (CPB). A significant reduction in final body mass of 4th larval stage fed with potato leaves coated with methanol extract of flowers in the concentration of 10 mg/mL was observed (157.67 mg vs. 182.26 mg of controls fed with non-treated leaves). This caused delayed molting since treated larvae reached the maximal mass a day after controls and this delay persisted during the entire larval development. Continuous feeding caused a 25% decline in digestive proteolytic activity of the 4th instar in comparison to controls. The results suggest that French marigold methanol extract of flowers could be proposed as a promising antifeedant for CPB management, with an impact on the reduction in the environmental footprint associated with synthetic pesticide application.

Transcriptome Profiling of the Potato Exposed to French Marigold Essential Oil with a Special Emphasis on Leaf Starch Metabolism and Defense against Colorado Potato Beetle.

Flower strips of French Marigold are commonly used pest repellents in potato fields. However, the effect of French Marigold volatiles on potato metabolism, physiology and induced defense is unknown. Thus, a microarray transcriptome analysis was performed to study the effects of French Marigold essential oil (EO) on laboratory-grown potato. After 8 h of exposure to EO, with gas chromatography/mass spectrometry (GC/MS)-detected terpinolene and limonene as dominant compounds, 2796 transcripts were differentially expressed with fold change >2 compared to expression in controls. A slightly higher number of transcripts had suppressed expression (1493 down- vs. 1303 up-regulated). Since transcripts, annotated to different photosynthesis-related processes, were mostly down-regulated, we selected a set of 10 genes involved in the leaf starch metabolism pathway, and validated microarray patterns using quantitative reverse transcription polymerase chain reaction (RT-qPCR). Except for decreased synthesis and induced decomposition of starch granule in leaves, 8 h long EO exposure slightly elevated the accumulation of sucrose compared to glucose and fructose in subjected potato plants. An in vitro feeding bioassay with Colorado potato beetle showed that EO-induced alternations on transcriptional level and in the sugars' metabolism caused the enhancement of feeding behavior and overall development of the tested larvae. Results of comprehensive analysis of transcriptional responses in potato exposed to French Marigold EO provide a basis for further elucidation of molecular mechanisms underlying eco-physiological interactions in companion planting cropping systems.

In vitro cultivation of tansy (Tanacetum vulgare L.): a tool for the production of potent pharmaceutical agents.

In this study, tansy (Tanacetum vulgare L.) in vitro culture was established from seeds collected from natural populations. The multiplication of plantlets was conducted through shoot tips that exhibited potent apical growth and regeneration capacities on basal medium (BM), without the addition of any plant growth regulators (PGRs). PGRs were also omitted for the establishment and cultivation of tansy root cultures. Both abaxial and adaxial leaf surfaces of in vitro micropropagated plantlets were covered with glandular biseriate trichomes. Histochemical staining showed that glandular secretions were rich in lipid and terpene compounds, confirmed by GC-MS analysis of essential oil (EO). In the total EO, similar portions of oxygenated monoterpenes (38.5% m/m) and oxygenated sesquiterpenes (22.6% m/m) were detected. Chemical profiles of methanol extracts of in vitro cultured tansy shoots and roots varied in quantity and quality from those obtained from wild-growingtansy. HPLC analysis indicated that the methanol extracts of in vitro cultured roots were the richest in 3,5-O-dicaffeoylquinic acid (3,5-O-DCQA), in which the concentration was 6 times higher (10.220 mg/g DW) than that in the extract obtained from roots of wild-growing tansy (1.684 mg/g DW). This result is noticeable in the manner of industrial production of biologically active 3,5-O-DCQA that has been shown to have antioxidant, hepatoprotective, antiviral, antimutagenic, and immunomodulatory activity. Biotechnological interventions on secondary metabolite production taking place in trichomes could further enhance the production of some important tansy metabolites and further investigation will be directed toward the elucidation of the pharmaceutical potential of tansy in vitro obtained metabolites, as mixtures or single moieties.

Evaluation of tansy essential oil as a potential "green" alternative for gypsy moth control.

The development of "green" alternatives to chemical pesticides could play a crucial role in integrated pest management (IPM). Their use is considered either as a substitution for or in addition to hazardous synthetic products. We analysed the influence of three concentrations of tansy (Tanacetum vulgare L.) essential oil (EO), previously characterised by GC-MS, on the survival and moulting of the 2nd instar and the nutritional indices of the 4th instar gypsy moth (Lymantria dispar L.) larvae. In a residual contact toxicity assessment, the exposure to tansy EO caused low mortality (< 10%) while larval development was significantly slowed down, i.e., the percentage of larvae that moulted into the 3rd instar was reduced. On the other hand, when tansy EO was incorporated into the diet (digestive toxicity assay), high mortality and a lack of moulting after 120 h of eating were recorded for the highest applied concentration of EO. During 48 h of feeding on EO-supplemented food at concentrations of 0.5 and 1% (v/v), the relative growth rate (RGR) of the 4th instar larvae significantly decreased, which can be explained by a significant reduction of the relative consumption rate (RCR) and significantly or marginally significantly lower efficiency of conversion of ingested food into insect biomass (ECI). Although the RCR was also reduced with the lowest applied EO concentration (0.1%), the ECI was not affected which meant the RGR was as high as it was for the control larvae. ECI changes, when two higher EO concentrations were applied, were due to a reduction in the efficiency of conversion of digested food into biomass (ECD), while approximate digestibility was unaffected by the presence of EO in the food. Our results on the significant negative effects of tansy EO on gypsy moth larval survival, development time, and nutritional physiology suggest that it could be considered in future designs for botanical insecticides for gypsy moth control.