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Karen C dos Santos,1 Maria Fatima GF da Silva,1 Edenir R Pereira-Filho,1 Joao B Fernandes,1 Igor Polikarpov,2 Moacir R Forim11Department of Chemistry, Federal University of Sao Carlos, Sao Carlos, 2Physics Institute of Sao Carlos, University of Sao Paulo, Sao Carlos, Sao Paulo, BrazilAbstract: This present investigation deals with the development and optimization of polymeric nanoparticle systems loaded with 3,5,3´-triiodothyroacetic acid (Triac). A 211–6 fractional factorial design and another 22 factorial design were used to study the contrasts on particle size distribution, morphology, surface charge, drug content, entrapment efficiency, and in vitro drug release profiles. The independent variables were the concentration of Triac, type and quantity of both polymer and oil, quantity of Span™ 60 and Tween® 80, volume of solvent and water, and velocity of both magnetic stirring and the transfer of the organic phase into the aqueous solution. The results of optimized formulations showed a narrow size distribution with a polydispersity index lower than 0.200. The particle sizes were on average 159.6 nm and 285.6 nm for nanospheres and nanocapsules, respectively. The zeta potential was higher than 20 mV (in module) and the entrapment efficiency was nearly 100%. A high-performance liquid chromatography method was developed, validated, and efficiently applied to Triac quantification in colloidal suspension. The main independent variables were the type and quantity of the polymer and oil. In vitro drug release profile depicted several features to sustain Triac release. Different formulations showed various release rates indicating an interaction between Triac and other formulation compounds such as polymer and/or oil quantity. Two different models were identified (biexponential and monoexponential) that allowed the control of both the release rate and Triac concentration. Thus, the prepared nanoparticles described here may be of clinical importance in delivering Triac for thyroid treatment.Keywords: Triac, nanoparticles, optimization, factorial design of experiments, HPLC analytical method

The contamination of water bodies by synthetic organic compounds coupled with climate change and the growing demand for water supply calls for new approaches to water management and treatment. To tackle the decontamination issue, the activation of peroxymonosulfate (PMS) using copper magnetic ferrite (CuMF) nanoparticles prepared under distinct synthesis conditions was assessed to oxidize imidacloprid (IMD) insecticide. After optimization of some operational variables, such as CuMF load (62.5-250 mg L -1 ), PMS concentration (250-1000 μM), and solution pH (3-10), IMD was completely oxidized in 2 h without interferences from leached metal ions. Such performance was also achieved when using tap water but was inhibited by a simulated municipal wastewater due to scavenging effects promoted by inorganic and organic species. Although there was evidence of the presence of sulfate radicals and singlet oxygen oxidizing species, only four intermediate compounds were detected by liquid chromatography coupled to mass spectrometry analysis, mainly due to hydroxyl addition reactions. Concerning the changes in surface properties of CuMF after use, no morphological or structural changes were observed except a small increase in the charge transfer resistance. Based on the changes of terminal surface groups, PMS activation occurred on Fe sites., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

This study investigated the chemical composition and the antifungal activity of the essential oil extracted from the Hymenaea stigonocarpa fruit peel (HSFPEO) against Botrytis cinerea , Sclerotinia sclerotiorum , Aspergillus flavus and Colletotrichum truncatum . The HSFPEO obtained through hydrodistillation was analyzed with gas chromatography coupled to mass spectrometry. The antifungal activity was determined by the mean mycelial growth inhibition of the fungus treated with essential oils and growth control. The major constituents of HSFPEO were spathulenol (25.19%) and caryophyllene oxide (13.33%). HSFPEO demonstrated antifungal activity against all fungi tested in all concentrations evaluated in a dose-dependent behavior. The best results were observed against B. cinerea and A. flavus , in which the lowest concentration tested inhibited more than 70% of mycelial growth. Based on the current knowledge, this study describes for the first time the chemical composition and the antifungal activity of HSFPEO against the phytopathogenic fungi B. cinerea and C. truncatum .

We report on a new iron (iii)-cyamelurate-based coordination polymer. The new material based on a heptazine derivative was prepared in aqueous medium and characterized by a variety of techniques including TGA, FTIR, XRD, HRTEM, and STEM. Due to the high structural stability of the complex in aqueous media, its heterogeneous Fenton-like catalytic activity was evaluated using a model molecule. The results obtained showed a high catalytic activity in both in basic and acid media. The pseudo-first-order rate constants normalized by iron(III) concentrations was approximately 1000 times higher than the result obtained for traditional heterogeneous catalysts based on iron(III) oxyhydroxides. The best observed catalytic activities were attributed to the increase in the binding sites of Fe 3+ ions, in parallel with the increased exposure of the catalytic sites, leading to a higher atomic efficiency of the reaction.

Diabrotica speciosa is an important pest of several crops in South America, including soybeans. Adults cause severe defoliation in soybean plants, and damage is significant when cotyledons are attacked. This study evaluated feeding non-preference to D. speciosa adults using 10 soybean genotypes, testing (i) 15-day-old whole plants and (ii) leaf disks of 60-day-old plants, through assessments of soybean attractiveness and leaf area consumed (LAC). Foliar contents of flavonoids and nutrients, and leaf trichome density were quantified for potential correlations with soybean resistance to adult of D. speciosa . In the whole young-plant experiment, under free-choice conditions, the lowest LAC was observed in IAC 100 and PI 227687. In no-choice, PI 227687 and IGRA RA 626 RR showed lower LAC than the other genotypes. In the leaf disk test, in free-choice, the genotypes IAC 100, PI 274454, PI 227687, DM 339, and BR 16 were the least preferred by adult of D. speciosa . In no-choice, PI 274454 was one of the least preferred, similarly to IGRA RA 626 RR, Dowling, and PI 227687. In the whole plant experiment, a high rutin content and low amounts of zinc, calcium, sulfur and manganese were associated with less consumption of D. speciosa on leaves of resistant genotypes. In contrast, in the leaf disk test there was a significant influence of trichomes in soybean resistance to the pest. In conclusion, the PI lines herein assessed are also promising sources for developing cultivars resistant to D. speciosa .

Lignan dinitrohinokinin displays important biological activities, which led to the preparation of its poly-ε-caprolactone nanoparticles. Kinetics analysis revealed initially slow drug release followed by a prolonged, moderate release 6 h later due to DNHK diffusion through the polymeric matrix. Molecular dynamics simulations show that DNHK molecules that interact stronger with other DNHK molecules near the PCL/DNHK surface are more difficult to dissociate from the nanoparticle. The smaller diameter nanocapsules with negative surface charge conferred good colloidal stability. The formulations showed a size distribution with monodisperse systems formation. In vivo evaluation of schistosomicidal activity against Schistosoma mansoni showed that DNHK, when incorporated into nanoparticles, caused egg number reduction of 4.2% and 28.1% at 40 mg/kg and 94.2% and 84.4% at 400 mg/kg in the liver and the spleen, respectively. The PCL nanoparticles were stable in aqueous dispersion and could be optimized to be used as a promising lignan release agent.

Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) Middle-East Asia Minor 1 is a major pest of agricultural production systems. It is controlled by synthetic insecticides. Essential oils are promising eco-friendly alternatives. This study developed and characterized nanoparticles loaded with essential oils of Zanthoxylum riedelianum Engl. (Rutaceae) leaves and evaluated their potential for B. tabaci management. The essential oil exhibited an average yield of 0.02% (w w -1 ) and showed as major components γ-elemene (24.81%), phytol (18.16%), bicyclogermacrene (16.18%), cis-nerolidol (8.26%), and D-germacrene (6.52%). Characterization of the nanoparticles showed a pH between 4.5 and 6.7, a zeta potential of approximately - 25 mV, particle-size distribution ranging from 450 to 550 nm, and encapsulation efficiency close to 98%. The nanoencapsulation was an efficient process that provided photostability against photodegradation. Bioassays with crude and nanoencapsulated essential oils significantly reduced the number of nymphs and eggs of B. tabaci, with the best results observed at concentrations of 5 and 2% (v v -1 ). Our results demonstrated that essential oils from Z. riedelianum can be nanoformulated resulting in a stable product while maintaining their biological activity against B. tabaci Middle-East Asia Minor 1., (© 2022. Sociedade Entomológica do Brasil.)

Schistosomiasis mansoni is an infectious parasitic disease caused by worms of the genus Schistosoma, and praziquantel (PZQ) is the medication available for the treatment of schistosomiasis. However, the existence of resistant strains reinforces the need to develop new schistosomicidal drugs safely and effectively. Thus, the (±)-licarin A neolignan incorporated into poly-Ɛ-caprolactone (PCL) nanoparticles and not incorporated were evaluated for their in vivo schistosomicidal activity. The (±)-licarin A -loaded poly(ε-caprolactone) nanoparticles and the pure (±)-licarin A showed a reduction in the number of worm eggs present in spleens of mice infected with Schistosoma mansoni. In addition, the (±)-licarin A incorporated in the concentration of 20 mg/kg and 200 mg/kg reduced the number of worms, presenting percentages of 56.3% and 41.7%, respectively., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Soybean crop is regulated by abiotic and biotic stresses with great potential in reducing grain yield and quality. The selection of resistant cultivars is a promising approach for mitigating these damages. We evaluated the chemical profile of Glycine max leaves from different cultivars in order to explore their defense mechanisms against Spodoptera cosmioides caterpillars. We optimized solid-liquid extraction techniques using ultrasound bath and static headspace extraction. Additionally, we developed an approach based on liquid and gas chromatography for analyzing the chemical profile of G. max cultivars. The principal component analysis allowed the classification of transgenic cultivars, which are classified as susceptible to S. cosmioides , from those obtained by genetic improvement and resistant to the insect. Differences were observed in the abundance of phenolic glycosides, lipids, aldehydes, and alcohols. More specifically, S. cosmioides resistant cultivars presented molecules related to the jasmonic and salicylic acid pathways. Such data can contribute to a molecular understanding of phenotypic diversity in soybean cultivars, from plant quality to resistance mechanisms and adaptation, to environmental stress and herbivory., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Aguiar, Mueller de Freitas, de Freitas, Boiça Júnior, Carneiro, da Silva, Fernandes and Forim.)

Encapsulation via nanotechnology offers a potential method to overcome limited thermal and photo-stability of botanical pesticides. In this study, nanospheres of essential oils (NSEO) derived from Zanthoxylum rhoifolium Lam. fruit were characterized and evaluated for their photostability and insecticidal activity against Bemisia tabaci. Three major compounds of Z. rhoifolium fruits were detected by CG-MS: β-phellandrene (76.8%), β-myrcene (9.6%), and germacrene D (8.3%). The nanoprecipitation method was used to obtain homogeneous spherical NSEO, with ≥98% encapsulation efficiency. Tests with UV/Vis spectrophotometry showed significantly reduced photodegradation from exposed NSEO samples when compared with essential oil (EO) controls. Whitefly screenhouses bioassays with bean plants treated with 0.25, 0.5, 1 and 1.5% suspensions showed EO treatments in both free and nanoencapsulated forms reduced adult whitefly oviposition by up to 71%. In further tests, applications at 1.5% caused ≥64% mortality of second instar nymphs. When the test was conducted under high temperature and light radiation conditions, the insecticidal effect of NSEO treatments was improved (i.e., 84.3% mortality) when compared to the free form (64.8%). Our results indicate the insecticidal potential of EO-derived from Z. rhoifolium fruits with further formulation as nanospheres providing greater photostability and enhanced insecticidal activity against B. tabaci under adverse environmental conditions.

The phytopathogenic fungi Sclerotinia sclerotiorum and Rhizopus stolonifer infect different crop plants. Essential oils have been used as an alternative to chemical control methods. Therefore, the objective of this study was to analyze the essential oil chemical composition of the stem bark of Zanthoxylum riedelianum (ZREO) and evaluate its antifungal potential. The ZREO obtained by hydrodistillation was analyzed using gas chromatography coupled to mass spectrometry and evaluated for in vitro antifungal activity. The major components identified in the chemical analysis were E -nerolidol (67.21%), α-selinene (14.94%), and β-selinene (7.41%). The antifungal potential of ZREO against S. sclerotiorum and R. stolonifer was detected at all concentrations evaluated in a dose-dependent manner. The best results were against R. stolonifer , with the concentration of 150 µLmL -1 inhibiting more than 80% of mycelial growth. On the basis of the current knowledge, this study describes for the first time the chemical composition and their antifungal activity.

Garlic is a health promoter that has important bioactive compounds. The bioactive extraction is an important step in the analysis of constituents present in plant preparations. The purpose of this study is to optimize the extraction with the best proportion of solvents to obtain total phenolic compounds (TPC) and thiosulfinates (TS) from dried garlic powder, and evaluate the antioxidant activities of the optimized extracts. A statistical mixture simplex axial design was used to evaluate the effect of solvents (water, ethanol, and acetone), as well as mixtures of these solvents, after two ultrasound extraction cycles of 15 min. Results showed that solvent mixtures with a high portion of water and pure water were efficient for TPC and TS recovery through this extraction procedure. According to the regression model computed, the most significant solvent mixtures to obtain high TPC and TS recovery from dried garlic powder are, respectively, the binary mixture with 75% water and 25% acetone and pure water. These optimized extracts presented oxygen radical absorbance capacity. Pure water was better for total antioxidant capacity, and the binary mixture of water-acetone (75:25) was better for DPPH scavenging activity. These optimized extracts can be used for industrial and research applications.

Forced degradation tests are studies used to assess the stability of active pharmaceutical ingredients (APIs) and their formulations. These tests are performed submitting the API under extreme conditions in order to know the main degradation products in a short period of time. The results of these studies are used to assess the degradation susceptibility of APIs and to validate chromatographic analytical methods. However, most of degradation studies are performed using one-factor-at-the-time (OFAT) which does not consider the interactions between degradation variables. This work proposes the use of Design of Experiment (DoE) approach in forced degradation of albendazole (ABZ). It was used a central composite design (CCD) to evaluate the forced degradation in a multivariate way. Experiments were performed taking into account the variables pH, temperature, oxidizing agent (H 2 O 2 ) and UV radiation. It was verified the influence of the variables and their interactions on the ABZ degradation. The ABZ oxidation showed to be the main degradation route for ABZ, which is strongly influenced by the temperature. The hydrolysis was relevant at alkaline medium and high temperature. LC-IT-MS n was used to identify the degradation products. It was found three known degradation products (albendazole-2-amino, albendazole sulfoxide and albendazole sulfone) and a new derivate of albendazole molecule (albendazole sulfoxide with a chlorine). This last one was isolated and characterized by UPLC-QToF-MS and NMR analyses., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The fungus Beauveria bassiana is widely used for pest control; however, biostability and dispersion for broth pulverization are limiting factors for its application in the field. In this context, formulation techniques such as microencapsulation are viable alternatives. The aim of this work is to optimize B. bassiana formulations by spray dryer and evaluate its stability and biological activity against Spodoptera cosmioides compared to ionic gelatinization formulations. The fungus was biocompatible with all evaluated biopolymers (lignin, cellulose, starch, humic substances, and alginate). The encapsulation by spray drying was optimized by factorial design in an inlet and outlet air temperature of 120°C and 68°C, respectively; aspirator rate of 35 m 3 ·h -1 , feed flow rate of 12 mL·min -1 ; and drying gas flow at 35 L·h -1 . The ionic gelation capsules were obtained using a 0.5% quantity of conidia in a 1% sodium alginate solution dropped into a 0.5 mol·L -1 CaCl 2 solution using a peristaltic pump. Spray drying provided smaller microcapsules than those by ionic gelation. Both techniques produced more stable conidia when exposed to temperature and UV-radiation than non-formulated B. bassiana . The formulations prepared by spray drying showed gains at aqueous dispersion. Biological assays against Spodoptera cosmioides showed a mortality rate of up to 90%. These results demonstrate the suitability of encapsulating B. bassiana conidia stably in aqueous dispersion without loss of viability and virulence., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Felizatti, Manzano, Rodrigues, da Silva, Fernandes and Forim.)

Although many copper-based antimicrobial compounds have been developed to control pathogenic bacteria and fungi in plants and applied for crop protection, there is evidence that several plant pathogens have developed resistance to copper-based antimicrobial compounds, including some Xanthomonas species. Xylella is a bacterial genus belonging to the Xanthomonas family; and X. fastidiosa, which is responsible for citrus variegated chlorosis (CVC) in sweet orange, may develop resistance to one or more copper-based antimicrobials. Because of the time required for the development and approval of new antimicrobials for commercial use, the discovery of novel bactericidal compounds is essential before the development of resistance to the antimicrobials currently in use becomes widespread. Here, we explored the antimicrobial potential of two newly synthesized antimicrobials complexes and one natural compound against X. fastidiosa. Several nuclear magnetic resonance (NMR) assays with high resolution and sensitivity were developed to identify new diastereoisomers in the context of octahedral ruthenium - [Ru(narin)(phen) 2 ]PF 6- and magnesium naringenin 5-alkoxide - [Mg(narin)(phen) 2 ]OAc - complexes, obtained in the present work. The NMR assays proved to be powerful tools for the identification of isomers in metal complexes. Moreover, a protocol for the in-vivo determination of the effects of these complexes against X. fastidiosa was developed. The main trunks of X. fastidiosa infected plants were injected with the two complexes as well as with the limonoid azadirachtin using a syringe; the number of bacterial cells in the plants following treatment was estimated via real-time quantitative PCR (qPCR). Importantly, the administration of both complexes and of azadirachtin drastically reduced the number of X. fastidiosa cells in vivo., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Fruit processing waste, such as kernels (endocarp + seed) of avocado [Persea americana Mill. (Lauraceae)], could be used as raw material in the preparation of botanical insecticides. In light of this potential, this study assessed the insecticidal action of extracts and fractions from kernels of two avocado cultivars (Breda and Margarida) on Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) biotype B, an important pest species in tropical conditions. Ethanolic and aqueous extracts prepared from kernels of P. americana, regardless of the plant cultivar used, caused promising insecticidal activity to whitefly nymphs. Based on yield in crude extracts [10.32 and 9.85% (w/w), respectively, for cultivars Breda and Margarida], on the bioassay results with crude extracts and on the chemical profiles, the ethanolic extract of kernels of P. americana cv. Breda was chose for the continuation of the study. Thus, the ethanolic extract of kernels of cv. Breda (LC 50  = 197.84 ppm and LC 90  = 567.19 ppm) was selected and subjected to fractionation by the liquid-liquid partition technique. The hexane and dichloromethane fractions of this extract caused significant mortality of nymphs. The analysis using the ultraviolet (UV) and hydrogen nuclear magnetic resonance ( 1 H NMR) showed the presence of long-chain aliphatic compounds (alkanols or acetogenins of Lauraceae), alkylfurans (or avocadofurans), and unsaturated fatty acids in these fractions, which are possibly related to bioactivity observed in B. tabaci, besides saccharides. The results show that kernels of P. americana are promising sources of compounds with insecticidal action for the control of B. tabaci biotype B, a great opportunity to transform environmental problems into eco-friendly solutions to agriculture.

Rationale: Xylella fastidiosa causes citrus variegated chlorosis (CVC) in sweet orange trees. A diagnostic method for detecting CVC before the symptoms appear, which would inform citrus producers in advance about when the plant should be removed from the orchard, is essential for reducing pesticide application costs., Methods: Chemometrics was applied to high-performance liquid chromatography diode array detector (HPLC-DAD) data to evaluate the similarities and differences between the chromatographic profiles. A liquid chromatography/atmospheric pressure chemical ionization mass spectrometry selected reaction monitoring (LC/APCI-MS-SRM) method was developed to identify the major compounds and to determine their amounts in all samples., Results: We evaluated the effect of this bacterium on the variation in the chemical profile in citrus plants. The organs of C. sinensis grafted on C. limonia were analyzed. Chemometrics was applied to the obtained data, and two major groups were differentiated. Flavonoids were observed in one group (leaves) and coumarins in the second (roots), both at higher concentrations in the plants with CVC symptoms than in those without the symptoms and those in the negative control. The rootstocks also interfered in the metabolism of the scion., Conclusions: The developed LC/APCI-MS-SRM method for detecting CVC before the symptoms appear is simple and accurate. It is inexpensive, and many samples can be screened per hour using 1 mg of leaves. Knowledge of the influence of the rootstock on the chemical profile of the graft is limited. This study demonstrates the effect of the rootstock in synthesizing flavonoids and increasing its content in all parts of the graft., (© 2020 John Wiley & Sons, Ltd.)

Essential oils are volatile compounds commonly used by several industries, easily degradable, which restrains their applications. Therefore, we developed and validated a methodology for producing microcapsules loaded with orange essential oil, using a spray-drying process. The experimental design results showed that the combination between a low flow transfer rate (0.15 L h -1 ) of the colloidal suspension, a higher drying air flow rate (536 L h -1 ), and an inlet air temperature of 150 °C to the spray-dryer were the most important parameters for the atomization efficiency. The method optimization resulted in microcapsules with powder recovery between 7.6 and 79.9% (w w -1 ), oil content ranging from 8.9 to 90.4% (w w -1 ), encapsulation efficiency between 5.7 and 97.0% (w w -1 ), and particle sizes with a high frequency of distribution less than 4 μm. In these experiments, gelatin and lignin were evaluated as biopolymers of encapsulation. We also developed an analytical method using headspace gas chromatography. The matrix effects could be addressed by using matrix-matched calibration curves. The chromatographic analysis was linear and selective for D-limonene between 0.025 and 3.00 µg mL -1 , with correlation coefficients higher than 0.99. The analytical method had limits of detection and quantitation of 0.024 and 0.073 mg g -1 for gelatin and 0.039 and 0.119 mg g -1 for lignin, respectively.

Citrus sinensis and Citrus limonia were obtained by germination from seeds, and isotopic-labeling experiments using d-[1- 13 C]glucose were performed with the seedlings. After 60 days, the seedlings were analyzed by high-performance liquid chromatography-ultraviolet-solid-phase extraction-nuclear magnetic resonance, data and the 13 C enrichment patterns of xanthyletin and seselin indicated that the pyran ring was formed by the methylerythritol phosphate pathway and that the coumarin moiety was derived from the shikimate pathway in both compounds. This information regarding the biosynthetic pathway can be used to increase resistance against phytopathogens, because xanthyletin and seselin are reported to have antimicrobial activity on the growth of Xylella fastidiosa , which causes citrus variegated chlorosis in orange.

Essential oils (EO) from aromatic and medicinal plants generally perform a diverse range of biological activities because they have several active constituents that work in different mechanisms of action. EO from Citrus peel have an impressive range of food and medicinal uses, besides other applications. EO from Citrus reticulata, C. sinensis and C. deliciosa were extracted from fruit peel and analyzed by GC-MS. The major constituent of EO under evaluation was limonene, whose concentrations were 98.54%, 91.65% and 91.27% for C. sinensis, C. reticulata and C. deliciosa, respectively. The highest potential of inhibition of mycelial growth was observed when the oil dose was 300 μL. Citrus oils inhibited fungus growth in 82.91% (C. deliciosa), 65.82% (C. sinensis) and 63.46% (C. reticulata). Anti-Sclerotinia sclerotiorum activity of 90% pure limonene and at different doses (20, 50, 100, 200 and 300 μL) was also investigated. This monoterpene showed to be highly active by inhibiting 100% fungus growth even at 200 and 300 μL doses. This is the first report of the in vitro inhibitory effect of natural products from these three Citrus species and its results show that there is good prospect of using them experimentally to control S. sclerotiorum, in both greenhouse and field conditions.

In this study, the chemical composition and antibacterial and antiproliferative potential of the essential oil obtained from fresh leaves of Psidium myrtoides (PM-EO) against oral pathogens and human tumour cell lines were investigated for the first time. GC-FID and GC-MS analyses showed that trans -β-caryophyllene (30.9%), α-humulene (15.9%), α-copaene (7.8%), caryophyllene oxide (7.3%) and α-bisabolol (5.3%) are the major constituents of PM-EO. The antibacterial activity of PM-EO against a panel of oral pathogens was investigated in terms of their minimal inhibitory concentrations (MIC) using the broth microdilution method. PM-EO displayed moderate activity against Streptococcus mitis (MIC = 100 μg/mL), S. sanguinis (MIC = 100 μg/mL), S. sobrinus (MIC = 250 μg/mL), and S. salivarius (MIC = 250 μg/mL), and strong activity against S. mutans (MIC = 62.5 μg/mL). The antiproliferative activity in normal (GM07492A, lung fibroblasts) and tumour cell lines (MCF-7, HeLa, and M059 J) was performed using the XTT assay. PM-EO showed 50% inhibition of normal cell growth at 359.8 ± 6.3 μg/mL. Antiproliferative activity was observed against human tumour cell lines, with IC 50 values significantly lower than that obtained for the normal cell line, demonstrating IC 50 values for MCF-7 cells (254.5 ± 1.6 μg/mL), HeLa cells (324.2 ± 41.4 μg/mL) and M059 J cells (289.3 ± 10.9 μg/mL). Therefore, the cytotoxicity of PM-EO had little influence on the antibacterial effect, since it showed antibacterial activity at lower concentrations. Our results suggest that PM-EO is a promising source of new antibacterial and antitumour agents.

A high performance liquid chromatography (HPLC) method was developed for the simultaneous isolation, on a semi-preparative scale, of chavibetol and methyleugenol from the crude essential oil of P. pseudocaryophyllus leaves. The purity of the isolated compounds and their quantifications were developed using GC/FID. Chavibetol was isolated with high purity (98.7%) and mass recovery (94.6%). The mass recovery (86.4%) and purity (85.3%) of methyleugenol were lower than those of chavibetol. Both compounds were identified on the basis of spectral analysis. The results suggest that the method can provide chavibetol with high purity, mass recovery, and productivity from crude essential, which will be used in bioassays against stored insect pests.

The aim of our study was to produce and characterize poly-ε-caprolactone (PCL) nanospheres containing essential oils from Zanthoxylum riedelianum fruit and to evaluate their stability gains as well as their insecticidal and deterrent activities against whitefly ( Bemisia tabaci ). The PCL nanospheres exhibited a homogeneous spherical morphology, with particle diameters between 106.7 nm and 129.2 nm, pH of approximately 6, zeta potential (ZP) lower than -19.0 mV and encapsulation efficiency higher than 98%. Only 43% of the nanoencapsulated essential oil (NSEO) was degraded in response to ultraviolet light, whereas the essential oil (EO) degraded by 76% over the same period. In a free-choice test, the NSEO and EO reduced the number of whitefly eggs by approximately 70%. NSEO and EO at 1.5% killed 82.87% and 91.23% of 2nd-instar nymphs of whitefly, respectively. Although NSEO displayed lower insecticidal activity, it offers a greater advantage over the free EO, due to protection conferred by polymer against photodegradation. Therefore, its usage may optimize the maintenance of essential oils in the field through photoprotection and controlled release. Our results suggest that the EO of Z. riedelianum fruit can be used for B. tabaci management strategy; nevertheless, the benefits of NSEO require further evaluation at the field level.

Sclerotinia sclerotiorum is responsible for the white mold of soybeans, and the difficulty to control the disease in Brazil is causing million-dollar damages. Stachybotrys levispora has shown activity against S. sclerotiorum. In our present investigation, we analyzed the chemical basis of this inhibition. Eight compounds were isolated, and using spectroscopic methods, we identified their structures as the known substances 7-dechlorogriseofulvin, 7-dechlorodehydrogriseofulvin, griseofulvin, dehydrogriseofulvin, 3,13-dihydroxy-5,9,11-trimethoxy-1-methylbenzophenone, griseophenone A, 13-hydroxy-3,5,9,11-tetramethoxy-1-methylbenzophenone, and 12-chloro-13-hydroxy-3,5,9,11-tetramethoxy-1-methylbenzophenone. Griseofulvin inhibited the mycelial growth of S. sclerotiorum at 2 μg mL -1 . Thus, the antagonistic effect of S. levispora to S. sclerotiorum may well be due to the presence of griseofulvins. Our results stimulate new work on the biosynthesis of griseofulvins, to locate genes that encode key enzymes in these routes and use them to increase the production of these compounds and thus potentiate the fungicide effect of this fungus. S. levispora represents an agent for biocontrol, and griseofulvin represents a fungicide to S. sclerotiorum.

The aim of this work was to quantify B-complex vitamins in pharmaceutical samples by surface enhanced Raman spectroscopy technique using gold colloid substrate. Synthesis of gold nanoparticles was performed according to an adapted Turkevich method. Initial essays were able to suggest the orientation of molecules on gold nanoparticles surface. Central Composite design was performed to obtain the highest SERS signal for nicotinamide and riboflavin. The evaluated parameters in the experimental design were volume of AuNPs, concentration of vitamins and sodium chloride concentration. The best condition for nicotinamide was NaCl 2.3×10 -3 molL -1 and 700μL of AuNPs colloid and this same condition showed to be adequate to quantify thiamine. The experimental design for riboflavin shows the best condition at NaCl 1.15×10 -2 molL -1 and 2.8mL of AuNPs colloid. It was possible to quantify thiamine and nicotinamide in presence of others vitamins and excipients in two solid multivitamin formulations using the standard addition procedure. The standard addition curve presented a R 2 higher than 0.96 for both nicotinamide and thiamine, at orders of magnitude 10 -7 and 10 -8 molL -1 , respectively. The nicotinamide content in a cosmetic gel sample was also quantified by direct analysis presenting R 2 0.98. The t-student test presented no significant difference regarding HPLC method. Despite the experimental design performed for riboflavin, it was not possible its quantification in the commercial samples., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Diketopiperazines can be generated by non-enzymatic cyclization of linear dipeptides at extreme temperature or pH, and the complex medium used to culture bacteria and fungi including phytone peptone and trypticase peptone, can also produce cyclic peptides by heat sterilization. As a result, it is not always clear if many diketopiperazines reported in the literature are artifacts formed by the different complex media used in microorganism growth. An ideal method for analysis of these compounds should identify whether they are either synthesized de novo from the products of primary metabolism and deliver true diketopiperazines. A simple defined medium ( X. fastidiosa medium or XFM) containing a single carbon source and no preformed amino acids has emerged as a method with a particularly high potential for the grown of X. fastidiosa and to produce genuine natural products. In this work, we identified a range of diketopiperazines from X. fastidiosa 9a5c growth in XFM, using Ultra-Fast Liquid Chromatography coupled with mass spectrometry. Diketopiperazines are reported for the first time from X. fastidiosa , which is responsible for citrus variegated chlorosis. We also report here fatty acids from X. fastidiosa , which were not biologically active as diffusible signals, and the role of diketopiperazines in signal transduction still remains unknown., Competing Interests: The authors declare no conflict of interest.

Indiscriminate use of synthetic pesticides can be hazardous to both humans and the environment, but the use of natural products as a source of bio-based products, such as Melia azedarach extracts, is an interesting approach to overcome these hazards. Unfortunately, the limonoids found in M. azedarach with desired insecticidal properties (e.g. azadirachtin) may also be present with limonoids toxic to mammals. The goal of this report was to develop a fast and reliable MS-based experiment to characterize meliatoxins in crude extracts of M. azedarach, in order to provide unequivocal assessment of the safety for extracts for application in the field. MS and MS/MS experiments using MALDI ionization were evaluated as tools for the assignment of characteristic ions produced by each meliatoxin in crude extracts.The use of different experiments in combination, such as the analysis of fragment m/z 557 and [M + Na] + (adducts ions m/z 681 and m/ z 667), MALDI-MS can be used for detection of meliatoxins A 1 /B 1 or A 2 /B 2 in a crude extract and may be used to discriminate meliatoxins A from B, respectively. Subsequent MS/MS experiments can distinguish between the presence of group 1 and/or 2 in each class of meliatoxins classifying the proposed approach as a quick and efficient quality control method of meliatoxins in real M. azedarach samples., (Georg Thieme Verlag KG Stuttgart · New York.)

The chemical composition of volatile oils from 22 genotypes of Citrus and related genera was poorly differentiated, but chemometric techniques have clarified the relationships between the 22 genotypes, and allowed us to understand their resistance to D. citri. The most convincing similarities include the synthesis of (Z)-β-ocimene and (E)-caryophyllene for all 11 genotypes of group A. Genotypes of group B are not uniformly characterized by essential oil compounds. When stimulated with odor sources of 22 genotypes in a Y-tube olfactometer D. citri preferentially entered the arm containing the volatile oils of Murraya paniculata, confirming orange jasmine as its best host. C. reticulata × C. sinensis was the least preferred genotype, and is characterized by the presence of phytol, (Z)-β-ocimene, and β-elemene, which were not found in the most preferred genotype. We speculate that these three compounds may act as a repellent, making these oils less attractive to D. citri.

This study evaluated the effect of Euclea natalensis gel on the reduction of erosive wear with or without abrasion, in enamel and dentin. During two five-day experimental crossover phases, volunteers (n = 10) wore palatal devices containing human enamel and dentin blocks (E = 8 and D = 8). The gel was applied in a thin layer in the experimental group, and was not applied in the control group. In the intraoral phase, volunteers used the palatal appliance for 12 h before the gel treatment, and were instructed to start the erosive challenges 6 h after the gel application. Erosion was performed with Coca-Cola® (for 5 min) 4 times/day. The appliance was then put back into the mouth and was brushed after 30 minutes. After intraoral exposure, the appliances were removed and the specimens were analyzed using profilometry (mean ± SD, μm). The Euclea natalensis gel caused less wear in enamel in the experimental group (EROS = 12.86 ± 1.75 µm; EROS + ABRAS = 12.13 ± 2.12 µm) than in the control group (EROS = 14.12 ± 7.66 µm; EROS + ABRAS = 16.29 ± 10.72 µm); however, the groups did not differ from each other significantly. A statistically significant value was found for erosion and eros + abrasion in dentin (p = 0.001). Euclea natalensis may play a role in the prevention of dentin loss under mild erosive and abrasive conditions. A clinical trial is required to confirm these promising results in a clinical situation.

A high performance liquid chromatography-ultraviolet (HPLC-UV) method was developed for quantifying hesperidin and rutin levels in leaves and stems of Citrus limonia, with a good linearity over a range of 1.0-80.0 and 1.0-50.0 μg mL(-1) respectively, with r(2)>0.999 for all curves. The limits of detection (LOD) for both flavonoids were 0.6 and 0.5 μg mL(-1), respectively, with quantification (LOQ) being 2.0 and 1.0 μg mL(-1), respectively. The quantification method was applied to Citrus sinensis grafted onto C. limonia with and without CVC (citrus variegated chlorosis) symptoms after Xylella fastidiosa infection. The total content of rutin was low and practically constant in all analyses in comparison with hesperidin, which showed a significant increase in its amount in symptomatic leaves. Scanning electron microscopy studies on leaves with CVC symptoms showed vessel occlusion by biofilm, and a crystallized material was noted. Considering the difficulty in isolating these crystals for analysis, tissue sections were analyzed by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) to confirm the presence of hesperidin at the site of infection. The images constructed from MS/MS data with a specific diagnostic fragment ion (m/z 483) also showed higher ion intensities for it in infected plants than in healthy ones, mainly in the vessel regions. These data suggest that hesperidin plays a role in the plant-pathogen interaction, probably as a phytoanticipin. This method was also applied to C. sinensis and C. limonia seedlings, and comparison with the graft results showed that the rootstock had an increased hesperidin content ∼3.6 fold greater in the graft stem than in the stem of C. sinensis seedlings. Increase in hesperidin content by rootstock can be related to induced internal defense mechanisms., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

This study identified the threshold concentration of limonoids for the complete inhibition of oviposition of Ceratitis capitata (Wiedemann) in grapes 'Itália.' Choice and no-choice experiments with the insect were performed. The three no-choice bioassays were conducted following a completely randomized design with 18 treatments (three densities of insects [one, two, or three females]×five concentrations of limonoids and control) and 20 replicates. In a free choice bioassay, two fruits per cage (a treatment grape and a control) were provided for ovipositing. Three densities of insects (one, two, or three females) were used, with 15 replicates. Bioassays were conducted at 25±2°C, 60±10% relative humidity, and a photoperiod of 14:10 (L:D) h. The inhibition of oviposition of C. capitata was concentration dependent, with infestation occurring at lower concentrations of azadirachtin (+3-tigloylazadirachtol) and complete inhibition occurring at concentrations at or exceeding 100 ppm azadirachtin (+28.5 ppm of 3-tigloylazadirachtol), maintaining protective effects even at the most densely populated treatment (three females per fruit). When the pest had a free choice of host grapes (treatment vs. control), severe inhibition was observed at concentrations≥50 ppm azadirachtin (+14.3 ppm of 3-tigloylazadirachtol). We conclude that a threshold concentration of 100 ppm azadirachtin (+28.5 ppm of 3-tigloylazadirachtol) is capable of preventing grape infestation. This concentration is likely to provide a reliable level of protection, as the experimental population density of three females per fruit usually does not occur in the field and wild flies usually have more host options., (© The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

We evaluated oil and powder formulations of Melia azedarach for controlling larvae of Diabrotica speciosa (Germar) in corn and plant enhancement. Five concentrations of each formulation were evaluated and compared to fipronil (negative control) and distilled water (positive control). After treatment, the number of surviving insects (larvae, pupae, and adults), the adult body weight, the sex ratio, and the longevity were recorded, while the height, dry weight of aerial part and roots, and number of leaves of plants were measured. The oil formulation at 4.0 mL reduced the larvae population of D. speciosa similarly to the insecticide fipronil, which resulted in greater height, dry weight of the root system, and number of leaves. Powder formulation at concentrations of 40, 80, and 160 mg caused larval mortality above 80%; however, these concentrations did not prevent reduction of plant height and dry weight of aerial part. Further studies assessing the residual period of M. azedarach control against D. speciosa larvae and its phytotoxicity, which are common traits associated with azadirachtin application, are necessary to subsidize the next steps of this alternative control strategy.

Previous investigations of H. oreadica reported the presence of a wide spectrum of complex limonoids and dihydrocinnamic acids. Our interest in the Rutaceae motivated a reinvestigation of H. oreadica, H. brasiliana and H. superba searching for other secondary metabolites present in substantial amounts for taxonomic analysis. In a continuation of the investigation of the H. oreadica, three new limonoids have now been isolated 9α-hydroxyhortiolide A, 11β-hydroxyhortiolide C and 1(S*)-acetoxy-7(R*)-hydroxy-7-deoxoinchangin. All the isolated compounds from the Hortia species reinforce its position in the Rutaceae. With regard to limonoids the genus produces highly specialized compounds, whose structural variations do not occur in any other member of the Rutaceae, thus, it is evident from limonoid data that Hortia takes an isolated position within the family. In addition, H. superba afforded the unexpected coumarin 5-chloro-8-methoxy-psoralen, which may not be a genuine natural product. Solid-state cross-polarisation/magic-angle-spinning 13C nuclear magnetic resonance, X-Ray fluorescence and Field-emission gun scanning electron microscopy experiments show that the Sephadex LH-20 was modified after treatment with NaOCl, suggesting that when xanthotoxin (8-methoxy-psoralen) was extracted from cleaning of the gel column, chlorination of the aromatic system occurred.

Objective: To evaluate the efficacy of Neem (Azadirachta indica) experimental gel for the prevention of erosive wear on bovine dentin, in vitro., Material and Methods: One hundred dentin blocks were allocated into 5 experimental groups (20 samples each): C (control group, without gel); CG (control group, only base gel); F (fluoride gel, 1.23% NaF; pH 4.1, Dentsply; Brazil); N (Neem gel, 10% neem extract; pH 4.1, manipulation); NF (Neem+fluoride gel, 10% Neem extract and 1.23% NaF; pH 4.1, manipulation). The blocks were stored in artificial saliva for 24 hours. After this, they were submitted to six alternating re- and demineralization cycles. The blocks were analyzed for wear (profilometry). The results were submitted to statistical analysis by ANOVA and Tukey tests (P<0.05)., Results: The mean wear (±SD, µm) was shown as follows in groups: C (13.09±0.99), CG (10.60±1.99), F (10.90±1.44), N (12.68±1.13) and NF (10.84±1.65). All gels showed some preventive action when compared with control group. However, significant differences were found only between Neem+fluoride gel and fluoride gel., Conclusion: A single application of a neem-containing fluoride gel reduced dentin erosion, thus it is a possible alternative in reducing dental wear. Further research should investigate the action mechanism and the synergism between them.

Dioxins are a class of extremely hazardous molecules that might pose a threat to the environment. This work evaluated the microbial degradation of 1,2,3,4-tetrachlorodibenzo-p-dioxin (1,2,3,4-TCDD), in liquid broth using three brown-rot fungi and one white-rot fungi as control. A fast and reliable extraction method with recoveries of over 98% together with a validated GC-MS method was developed, and applied to quantify 1,2,3,4-TCDD in liquid broth, mycelia and reaction flask, with detection limits of 10 ppb. Among the four strains tested, brown-rot fungus Aspergillus aculeatus showed best results, removing up to 21% of dioxin after 30-day incubation. The results open both a path for biotechnological interest in bioremediation purposes and environmental behavior studies by using brown-rot fungus., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Biodegradable nanoparticles have been widely explored as carriers for controlled delivery of therapeutic molecules; however, studies describing the development of nanoparticles as carriers for biopesticide products are few. In this work, a new method to prepare nanoparticles loaded with neem (Azadirachta indica) extracts is presented. In this study, nanoparticles were formulated as colloidal suspension and (spray-dried) powder and characterized by evaluating pH, particle size, zeta potential, morphology, absolute recovery, and entrapment efficiency. A high-performance liquid chromatography method was used for nanoparticle characterization. The best formulations presented absolute recovery and entrapment efficiencies of approximately 100% and a release profile based on swelling and relaxation of the polymer or polymer erosion. The biological data of the formulated products against Plutella xylostella showed 100% larval mortality. The nanoparticle information improved the stability of neem products against ultraviolet radiation and increased their dispersion in the aqueous phase.

Xylella fastidiosa is a plant pathogen bacterium that causes diseases in many different crops. In citrus, it causes Citrus Variegated Chlorosis (CVC). The mechanism of pathogenicity of this bacterium is associated with its capacity to colonize and form a biofilm in the xylem vessels of host plants, and there is not yet any method to directly reduce populations of this pathogen in the field. In this study, we investigated the inhibitory effect of N-Acetylcysteine (NAC), a cysteine analogue used mainly to treat human diseases, on X. fastidiosa in different experimental conditions. Concentrations of NAC over 1 mg/mL reduced bacterial adhesion to glass surfaces, biofilm formation and the amount of exopolysaccharides (EPS). The minimal inhibitory concentration of NAC was 6 mg/mL. NAC was supplied to X. fastidiosa-infected plants in hydroponics, fertigation, and adsorbed to organic fertilizer (NAC-Fertilizer). HPLC analysis indicated that plants absorbed NAC at concentrations of 0.48 and 2.4 mg/mL but not at 6 mg/mL. Sweet orange plants with CVC symptoms treated with NAC (0.48 and 2.4 mg/mL) in hydroponics showed clear symptom remission and reduction in bacterial population, as analyzed by quantitative PCR and bacterial isolation. Experiments using fertigation and NAC-Fertilizer were done to simulate a condition closer to that normally is used in the field. For both, significant symptom remission and a reduced bacterial growth rate were observed. Using NAC-Fertilizer the lag for resurgence of symptoms on leaves after interruption of the treatment increased to around eight months. This is the first report of the anti-bacterial effect of NAC against a phytopathogenic bacterium. The results obtained in this work together with the characteristics of this molecule indicate that the use of NAC in agriculture might be a new and sustainable strategy for controlling plant pathogenic bacteria.

Neem derivatives have been widely touted as replacements for pesticides. A feasible replacement of synthetic insecticides in the management of fruit flies could be to use neem products in baits. This study evaluated the bioactivity of neem (Azadirachta indica A. Juss) derivatives in bait for adults of Anastrepha fraterculus (Wiedemann) and Ceratitis capitata (Wiedemann). The estimated LCs50 values for A. fraterculus and C. capitata were 7,522 ppm (18.40 ppm of azadirachtin) and 1,368 ppm (3.35 ppm of azadirachtin), respectively, using an aqueous extract of neem seeds in bait after 10 d of experimentation. No significant differences in the mortality of A. fraterculus and C. capitata adults exposed to baits made from different extracts and neem oil were observed after 3 h or 2 or 6 d; differences among the treatments were observed only on the 10th day of the evaluation. We conclude that neem derivatives applied as a bait spray over citrus plants did not demonstrate a toxic effect on A. fraterculus and C. capitata. The reasons for the low efficacy of the neem bait on Tephritid fruit flies are discussed.

A novel analytical method using HPLC-MS/MS operating in selected reaction monitoring (SRM) for evaluation of fungi efficacy to decolorize Remazol Brilliant Blue R (RBBR) dye solution was developed, validated and applied. The method shows high sensibility allowing the detection of 4.6 pM of RBBR. Four fungal strains were tested in liquid medium, three strains of Aspergillus (Aspergillus aculeatus, Aspergillus flavus and Aspergillus fumigatus) and Phanerochaete chrysosporium. All fungi were able to degrade the dye, with efficiencies ranging from 40% for P. chrysosporium up to 99% for A. flavus during a 30-day incubation period. During the experiment, increased accumulation of degradation products was observed in A. flavus cultures containing RBBR. Through the use of full scan HPLC-MS technique it was possible to propose the biogenesis of the microbial metabolic degradation pathway. Screening using microorganisms and RBBR may be hereafter used to investigate microbial biodegradation of high toxicity molecules such as dioxins., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Objective. To evaluate the effect of an experimental gel containing Euclea natalensis extract on dentin permeability. Methods. Thirty-six dentin discs, 1-mm-thick. The discs were prepared from the coronal dentin of extracted human third molars that were divided into 3 groups (n = 10). The dentin discs in each group were treated with the groups following experimental materials: (FG): 1.23% fluoride gel, pH 4.1; (EG): Euclea natalensis extract gel, pH 4.1; (CG): control gel, pH 4.1. The gels were applied to the occlusal slide of the dentin under the following conditions: after 37% phosphoric acid and before 6% citric acid. The hydraulic conductance (HC) of each condition was determined four times using a fluid flow apparatus (Flodec). The data were analyzed using Two-way ANOVA and Tukey's test (P < 0.05). Results. The greatest mean reduction in HC was produced in group EG dentin discs (61.2%; P < 0.05). Even after acid challenge with 6% citric acid the great reduction occurred in group EG (66.0%; P < 0.05) than other groups (CG-77.1%, FG-90.8%). Conclusion. E. natalensis gel not only reduced dentin permeability, but also resisted posttreatment citric acid challenge without changing its permeability. Further research has to confirm this promising result in the clinical situation.

The effect of four extracts from neem seeds (Azadirachta indica) containing 2000, 5000, 9000 and 10,000 ppm of azadirachtin A (AZA), quantified by high-performance liquid chromatography (HPLC) and diluted to 1.25%; 2.5%; 5.0%; 10.0% and 12.8% was verified by in vitro tests with engorged females and larvae of the cattle tick Rhipicephalus microplus. The results from the bioassays with the engorged females showed that the main toxic effect of the extracts was reduction of the reproductive parameters, with a sharp drop in the number of eggs laid and the hatching rate, mainly when the extracts were diluted to 10.0% and 12.8%. The product effectiveness (PE) calculations for all the solutions tested showed that the AZA solution at 10,000 ppm (N10) was the most effective. However, statistical analysis of the PE data obtained for the proportional AZA concentrations in the different diluted extracts showed significance (P<0.05) of the effects included in the model (extract dilution, principle effect (classificatory) of the assay (extract) and the interaction between the two), indicating significant variations due to the dilution, the test and the interaction between the two factors in the tests with engorged females. For solutions N2, N5, and N9, it was not possible to estimate LC(90) values in the dilution range tested. The lowest LC(50) was observed for extract N5, and although extract N10 was the only extract for which the LC(90) could be estimated within the range tested, the LC(50) was higher than for N5 and N9. These results suggest that substances other than AZA present in the extracts influenced the efficacy, especially up to a certain LC range. In the tests with larvae, no mortality was observed, indicating zero effectiveness of all the extracts tested. The results of the tests with engorged females showed that the neem extracts had acaricide activity, inhibiting egg laying and the larval hatching rate. Complementary studies are necessary to develop new methods to isolate and/or identify other substances besides AZA contained in this plant, to enable using products made from it as acaricides., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Objective: The aim of this study was to evaluate the capacity of potassium oxalate, fluoride gel and two kinds of propolis gel to reduce the hydraulic conductance of dentin, in vitro., Material and Methods: The methodology used for the measurement of hydraulic conductance of dentin in the present study was based on a model proposed in literature. Thirty-six 1-mm-thick dentin discs, obtained from extracted human third molars were divided into 4 groups (n=9). The groups corresponded to the following experimental materials: GI-10% propolis gel, pH 4.1; GII-30% propolis gel; GIII-3% potassium oxalate gel, pH 4,1; and GIV-1.23% fluoride gel, pH 4.1, applied to the dentin under the following surface conditions: after 37% phosphoric acid and before 6% citric acid application. The occluding capacity of the dentin tubules was evaluated using scanning electron microscopy (SEM) at ×500, ×1,000 and ×2,000 magnifications. Data were analyzed statistically by two-way ANOVA and Tukey's test at 5% significance level., Results: Groups I, II, III, IV did not differ significantly from the others in any conditions by reducing in hydraulic conductance. The active agents reduced dentin permeability; however they produced the smallest reduction in hydraulic conductance when compared to the presence of smear layer (P<0.05). The effectiveness in reducing dentin permeability did not differ significantly from 10% or 30% propolis gels. SEM micrographs revealed that dentin tubules were partially occluded after treatment with propolis., Conclusions: Under the conditions of this study, the application of 10% and 30% propolis gels did not seem to reduce the hydraulic conductance of dentin in vitro, but it showed capacity of partially obliterating the dentin tubules. Propolis is used in the treatment of different oral problems without causing significant great collateral effects, and can be a good option in the treatment of patients with dentin sensitivity.

Much attention has been given to the development of botanical insecticides to provide effective natural control of cattle ectoparasites without harming animals, consumers, and environment. This study evaluated the efficacy of a commercial neem cake in controlling Haematobia irritans infestation on cattle. The study was conducted at the Embrapa Southeast Cattle Research Center (CPPSE), in São Carlos, SP, Brazil, from April to July 2008. The neem cake mixed in mineral salt in a 2% concentration was provided to 20 Nelore cows during nine weeks and had its efficacy evaluated by comparison of the infestation level against a control group. Fly infestations were recorded weekly by digital photographs of each animal from both groups and the number of flies was later counted in a computer-assisted image analyzer. Quantification of neem cake components by high-performance liquid chromatography revealed the presence of azadirachtin (421 mg.kg(-1)) and 3-tigloyl-azadirachtol (151 mg.kg(-1)) in the tested neem cake. Addition of the 2% neem cake reduced mineral salt intake in about 22%. The 2% neem cake treatment failed to reduce horn fly infestations on cattle during the 9-week study period.

Introduction: Melia azedarach adapted to cool climates was selected as rootstocks for vegetative propagation of Azadirachta indica. Cleft grafting of A. indica on M. azedarach rootstock showed excellent survival. Little is known about the chemistry of grafting., Objective: The roots, stems, leaves and seeds of this graft were examined in order to verify if grafted A. indica would produce limonoids different from those found in non-grafted plants. Intact matured fruits were also studied to verify if they were free of meliatoxins., Methodology: After successive chromatographic separations the extracts afforded several limonoids. HPLC-MS/MS and MALDI-MS were used to develop sensitive methods for detecting azadirachtin on all aerial parts of this graft and meliatoxins in fruits, respectively., Results: The stem afforded the limonoid salannin, which was previously found in the oil seeds of A. indica. Salannin is also found in the root bark of M. azedarach. Thus, the finding of salannin in this study suggests that it could have been translocated from the M. azedarach rootstock to the A. indica graft. HPLC-MS/MS analyses showed that azadirachtin was present in all parts of the fruits, stem, flowers and root, but absent in the leaves. The results of MALDI-MS analyses confirmed the absence of meliatoxins in graft fruits., Conclusion: This study showed that A. indica grafted onto M. azedarach rootstock produces azadirachtin, and also that its fruits are free of meliatoxins from rootstocks, confirming that this graft forms an excellent basis for breeding vigorous Neem trees in cooler regions.

High-speed counter-current chromatography (HSCCC) with a two-phase solvent system (hexane-ethanol-acetonitrile-water 10:8:1:1, v/v) was applied to examine the leaves of Hortia oreadica, which afforded the known limonoid guyanin (1), the alkaloids rutaecarpin (2) and dictamnine (6), the dihydrocinnamic acid derivatives methyl 5,7-dimethoxy-2,2-dimethyl-2H-1-benzopyran-6-propanoate (3), 5,8-dimethoxy-2,2-dimethyl-2H-1-benzopyran-6-propanoic acid (4), together with the new E-3,4-dimethoxy-alpha(3-hydroxy-4-carbomethoxyphenyl)cinnamic acid (5). The recovery of compounds 1-6 was determined by comparison with LC-atmospheric pressure chemical ionization MS/MS data: 66.2%, 93.1%, 102.5%, 101.2%, 99.0% and 84.9%, respectively. Compound 3 showed IC(50) of 23.6microM against Plasmodium falciparum and 15.6microM against Trypanosoma brucei rhodesienses and was not toxic to KB cells (IC(50)>100microM).

The alkaloid ricinine, an insecticide for leaf-cutting ant (Atta sexdens rubropilosa), was obtained from Ricinus communis. A two-phase solvent system composed of CH(2)Cl(2)/EtOH/H(2)O (93:35:72, v/v/v) was used for high-speed counter-current chromatographic (HSCCC) isolation of ricinine in high yield and with over 96% purity, as determined by liquid and gas chromatography-mass spectrometry (LC-MS and GC-MS). Identification of ricinine was performed by comparison of (1)H NMR, (13)C NMR and LC-MS/MS data.