Identification of Citrus medica var. sarcodactylis Infected with Citrus Huanglongbing / 中国实验方剂学杂志

Objective::To analyze and compare different samples in many aspects to identify Citrus medica var. sarcodactylis infected with Huanglongbing(HLB) timely and accurately, in order to prevent and control the disease in time. Method::HLB was identified through character analysis, reverse transcription-polymerase chain reaction(RT-PCR), enzyme digestion reaction and Real-time PCR. Result::In terms of characters, there were typically variegated yellow leaves and relatively small fruit, even with deformity but without " red nose fruit" among C. medica var. sarcodactylis infected with HLB. All of these can be used as the basis for the preliminary identification of HLB in the fields. According to the RT-PCR test results and enzyme digestion reaction, when the primer was OI1/OI2c, there was specific band of 1 160 bp, which could be cut into 520 bp and 640 bp by Xba I enzyme. These results were consistent with the characters of other citrus plants infected with HLB. According to the Real-time PCR detection results, C. medica var. sarcodactylis infected with HLB had amplification curves and dissolved peaks, with the melting temperature was 82 ℃ and Ct between 24.6 to 28.2, while the normal plants were not amplified. Conclusion::Character analysis can be used to roughly distinguish HLB in the fields, but with a certain subjectivity. RT-PCR or Real-time PCR can be used to identify C. medica var.sarcodactylis infected with HLB in a timely and accurate manner, and qPCR detection is more sensitive and quantitative. Through the combination of character analysis and molecular identification, C. medica var.sarcodactylis infected with HLB can be determined more timely and accurately.

Survival analyses of Diaphorina citri immatures on young citrus orchard in São Paulo, Brazil / Análises de sobrevivência de imaturos de Diaphorina citri em pomar jovem de citros em São Paulo, Brasil

The survival of eggs and nymphs of Diaphorina citri Kuwayama (Hemiptera: Liviidae) was estimated on young plants (< 1 year) of Citrus sinensis (cultivar Valencia) grafted on Citrus limonia in 3 seasons (Winter and Spring 2013; and Autumn 2014) in Ribeirão Preto, São Paulo State, Brazil. Young shoots with eggs were selected from natural or artificial infestations. Eggs, small nymphs (1st to 3rd instar), and large nymphs (4th and 5th instar) were counted every 3 days. Shoots with large nymphs were caged until the emergence of the adults. The lowest eggs and nymph survival were observed during Winter (0.0%), and Autumn (0.4%); the highest survival rates were recorded during Spring (12.2%). The biotic factor of mortality identified was the parasitism of large nymphs by Tamarixia radiata Waterston (Hymenoptera: Eulophidae), observed in all seasons. Air temperatures (minimum and maximum), and air relative humidity (minimum) could not explain the immature survival differences between Spring (12.2%) and Autumn (0.4%). Rainfall during Spring (165 mm) was higher than during Autumn (48 mm) and may have favored the survival of D. citri in the Spring. Adverse abiotic factors seem to be more important on immature D. citri survival than the biotic factor (parasitism).(AU)

A sobrevivência de ovos e ninfas de Diaphorina citri Kuwayama (Hemiptera: Liviidae) foi estimada em plantas jovens (< 1 ano) de Citrus sinensis (cultivar Valencia) enxertadas em Citrus limonia em 3 estações (inverno e primavera de 2013, e outono de 2014) em Ribeirão Preto, estado de São Paulo, Brasil. Ramos novos com ovos foram selecionados a partir de infestações naturais ou artificiais. Ovos, ninfas pequenas (ínstar 1 ao 3) e ninfas grandes (ínstar 4 e 5) foram contados a cada 3 dias. Ramos com ninfas grandes foram colocados em gaiolas até a emergência dos adultos. As menores sobrevivências de ovos e ninfas foram observadas no inverno (0,0%) e no outono (0,4%), e a maior sobrevivência ocorreu na primavera (12,2%). O principal fator biótico de mortalidade identificado foi o parasitismo de ninfas grandes por Tamarixia radiata Waterston (Hymenoptera: Eulophidae), observado em todas as estações. As temperaturas (mínima e máxima) e umidade relativa (mínima) do ar não explicaram as diferenças nas sobrevivências de imaturos entre a primavera (12,2%) e o outono (0,4%). As chuvas na primavera (165 mm) foram maiores que no outono (48 mm) e podem ter favorecido a sobrevivência de D. citri na primavera. Fatores abióticos adversos parecem ser mais importantes na sobrevivência de imaturos de D. citri que o fator biótico (parasitismo).(AU)

Métodos para la detección presuntiva de Huanglongbing (HLB) en cítricos / Methods for presumtive detection of Huanglongbing (HLB) in Citrus

RESUMEN Huanglongbing (HLB) es considerada mundialmente como la enfermedad más amenazadora para la citricultura, y ha impactado principalmente en Asia, Sudáfrica y Brasil. A la fecha, no se cuenta con un tratamiento efectivo para la detección de esta enfermedad, que pueda ayudar a disminuir su propagación y la consecuente eliminación de árboles infectados. El método más confiable hasta ahora, para la identificación del HLB, es la prueba de reacción en cadena de la polimerasa (qrt-PCR), la cual es costosa y tardada. Esta es la primera revisión de los diferentes métodos existentes o en desarrollo para la detección e identificación del HLB; clasificados en métodos de análisis y reconocimiento de patrones en imágenes, espectrofotométricos, cromatográficos y moleculares a través del insecto vector. Algunos de estos métodos representan alternativas innovadoras con diferente nivel de eficiencia en tiempo, costo y confiabilidad en la detección de árboles enfermos de HLB o en el manejo de la enfermedad, con relación a los tradicionalmente utilizados.

ABSTRACT Huanglongbing (HLB) is considered worldwide as the most threatening disease for citrus and has impacted mainly in Asia, South Africa and Brazil. Until now, no effective treatment for the detection of this disease (is available, which can help diminish its spread and the consequent removal of infected trees). The most reliable identification HLB test method to date is the polyme-rase chain reaction (qrt-PCR), which is costly and time-consuming. This is the first review of the different existing or developing methods for HLB detection and identification, classified into analysis and patternrecognition in images, spectrophotometric, chromatographic and molecular through the insect vector. Some of these methods represent innovative alternatives with different levels of efficiency in time, cost and reliability in detecting diseased trees or in managing the disease, compared to those traditionally used.

Estudio de la estabilidad de emulsiones con estructuras líquido-cristalinas, y su aplicación farmacéutica mediante el agregado de un principio activo liposoluble: Econazol / Study of emulsions stability with liquid crystals structure and their pharmaceutical application by the addition of a liposoluble principe active: Econazole

La piel es un órgano apropiado para administrar principios activos con el fin de obtener efectos locales o sistémicos. Las formulaciones de uso tópico más comunes son: lociones, emulsiones, suspensiones, cremas, pomadas; las cuales deben reunir determinadas condiciones para ser aplicadas sobre la piel. El objetivo del presente trabajo es seleccionar una emulsión preparada con la técnica de formación de cristales líquidos compuesta de ácido esteárico, vaselina líquida, trietanolamina, propilparabeno, metilparabeno y agua, a la que se le incorpora un ingrediente farmacéutico activo liposoluble: econazol. El econazol, principio activo cuya vía de administración es la tópica y su acción es local, es una sustancia soluble en aceites, que se aloja en la fracción liposoluble de los cristales líquidos y en la fase interna de la emulsión sin que se modifique el perfil reológico ni la estabilidad de los sistemas. Se estudió además del HLB y de sus comportamientos reológicos, la presencia de cristales líquidos con luz polarizada, la existencia de gotas secundarias con luz común y la estabilidad de los sistemas por centrifugación y estrés térmico a temperaturas de 40 ºC. Los valores obtenidos en los estudios realizados, demostraron que la emulsión lograda presenta un perfil reológico y las condiciones de estabilidad adecuadas para ser utilizada como crema medicinal.

The skin is the appropiate organ to administrate active principles in orden to obtain local or systemic effects. Formulations for topical use most common are: lotions, emulsions, suspensions, creams, ointments, which must gather certain conditions to be applied to the skin. In this work, the objective is to select an emulsion prepared by a technique of liquid crystals formation composed by stearic acid, mineral oil, triethanolamine, propylparaben, methylparaben and water. To this formula we incorporated a pharmaceutical active liposoluble ingredient: econazole. Econazole, a principle active with topical administration and local action, is a substance soluble in oils, which stays in the liposoluble fraction of the liquid crystals and in their internal phase of the emulsion without modifying their rheological profile not even the stability of systems. Besides the HLB of the systems and their rheological behaviour we also study the presence of liquid crystals with polarized light, the existence of secondary drops with common light and systems stability by centrifugation, thermal stress and temperatures of 40 °C. The values obtained from the studies made, demonstrated that the emulsion achieved present a rheological profile and stability conditions suitable for medicinal use cream.

HLB value, an important parameter for the development of essential oil phytopharmaceuticals

Essential oils are used primarily as natural preservatives, flavourants and fragrances in cosmetic products. Several pharmacopeias possess monographs of plants which are good sources of essential oils, such as Brazilian Pharmacopeia, including Illicium verum Hook. f., Schisandraceae and Rosmarinus offi cinalis. Since determination of Hydrophile-Lipophile Balance (HLB) value of essential oils appears as a critical step for development of emulsions and other semi-solid formulations, evaluation of required HLB values for I. verum and R. offi cinalis essential oils is the aim of this study. They were obtained by hydrodistillation and several emulsions were prepared by changing emulsifiers. The couple sorbitan oleate/polysorbate 20 provided best emulsions and was used at different ratios, at a total blend concentration of 5% w/w. The lowest mean droplet diameters for R. offi cinalis and I. verum emulsions were obtained at HLB 16.5 (97.12 nm) and 16.7 (246.6 nm), respectively. Moreover, emulsions with R. offi cinalis were finer and presented some bluish reflection, characteristic of nanoemulsions. The lowest turbidity value for R. offi cinalis emulsion was also obtained at HLB 16.5 (0.33). Thus, the present study describes for the first time HLB values for R. offi cinalis (16.5) and I. verum (16.7) essential oils, contributing to their physicochemical characterization and technology development of phytopharmaceuticals.

Desenvolvimento de emulsões óleo de oliva/água: avaliação da estabilidade física / Development of olive oil-in-water emulsions: assessment of physical stability

Emulsões óleo de oliva/água, na presença de agentes emulsionantes não-iônicos, foram avaliadas quanto à estabilidade física. Assim, prepararam-se emulsões fazendo uso de diferentes emulsionantes, sendo um hidrofílico e o outro lipofílico, nas diferentes proporções. Às emulsões mais estáveis, adicionaram-se agentes auxiliares da emulsificação, visando otimizar a estabilidade; e estudos de estabilidade foram conduzidos, submetendo as amostras em condições e períodos diversos de armazenamento. Para caracterização da estabilidade, as amostras foram examinadas macroscopicamente e submetidas às análises de pH, centrifugação, viscosidade, potencial zeta e distribuição de tamanho de partícula. Os resultados demonstraram que as emulsões óleo de oliva/água não apresentaram alteração, ou seja mantiveram-se estáveis, quanto às propriedades organolépticas, bem como físicoquímicas, quando armazenadas à temperatura ambiente e protegidas da luz. Das emulsões obtidas, as que apresentaram maior estabilidade provêm da associação de agentes emulsionantes que resultaram em equilíbrio hidrofílico-lipofílico (EHL) equivalente a 12. As emulsões provenientes da associação de agentes emulsionantes que possuem cadeias de ácidos graxos insaturados similares ao óleo de oliva produziram estabilidade máxima, demonstrando que a similaridade estrutural entre os componentes da fase oleosa e os agentes emulsionantes é essencial para a estabilidade da emulsão.

Olive oil-in-water emulsions, developed with non-ionic emulsifiers, were assessed with regard to physical stability. Emulsions were prepared with two different emulsifiers, one of which was hydrophilic and the other lipophilic, in various proportions. To improve emulsion stability, auxiliary emulsifiers were added to the stablest emulsions and stability studies were carried out, in which the samples were stored for different periods and under various conditions. To test emulsion stability, the samples were examined macroscopically and various physicochemical properties, such as pH, centrifugation, viscosity, zeta potential and particle size distribution, were assessed. The results showed that olive oil-in-water emulsions are organoleptically and physicochemically stable, when stored at room temperature and protected from the light. Out of the emulsions developed, the most stable was based on an emulsifier blend that resulted in a hydrophilic-lipophilic balance (HLB) of 12. A blend of emulsifiers with unsaturated fatty acids of similar chain length to that of olive oil produced the stablest emulsions, showing that structural similarity between the hydrocarbon moieties of the oil phase and the surfactant is essential to successful emulsion stabilization.