Application of virus-induced gene silencing technology to investigate the phytochrome metabolism mechanism: a review / 生物工程学报

Color is an important indicator for evaluating the ornamental traits of horticultural plants, and plant pigments is a key factor affecting the color phenotype of plants. Plant pigments and their metabolites play important roles in color formation of ornamental organs, regulation of plant growth and development, and response to adversity stress. It has therefore became a hot topic in the field of plant research. Virus-induced gene silencing (VIGS) is a vital genomics tool that specifically reduces host endogenous gene expression utilizing plant homology-dependent defense mechanisms. In addition, VIGS enables characterization of gene function by rapidly inducing the gene-silencing phenotypes in plants. It provides an efficient and feasible alternative for verifying gene function in plant species lacking genetic transformation systems. This paper reviews the current status of the application of VIGS technology in the biosynthesis, degradation and regulatory mechanisms of plant pigments. Moreover, this review discusses the potential and future prospects of VIGS technology in exploring the regulatory mechanisms of plant pigments, with the aim to further our understandings of the metabolic processes and regulatory mechanisms of different plant pigments as well as improving plant color traits.

Evaluation of Natural Pigments Extracted from Tomato Wastes and the Possibility of their Encapsulation

This study aimed to evaluate the best methods from extracting natural pigments from tomato fruit wastes by four techniques used to extract lycopene and β-carotene each of them consisting of three solvents: ethanol, acetone and hexane in the following ratios (1:1:1), (2:1:1), (1:2:1) and (1:1:2) ml respectively. We studied too the possibility of encapsulation by freeze drying with a mixture of gelatin and gum Arabic as a carrier in nine microcapsules differing with respect to the total encapsulant (E) (2.5, 5.0, and 7.5%) and core (C) concentrations, the latter varying in relation to the total weight of encapsulant (25, 50, and 75%). The nine microcapsules were coded as follows: (1) E2.5/C25; (2) E2.5/C50; (3) E2.5/C75; (4) E5.0/C25; (5) E5.0/C50; (6) E5.0/C75; (7) E7.5/C25; (8) E7.5/C50; and (9) E7.5/C75. We found that the best solvent mixture for the extraction process was 1:1:2, and the most efficient microcapsules were E5.0/C75, E5.0/C50 and E2.5/C25. By studying the stability of the best three microcapsules when exposed to different values of heat, light, oxygen and pH, it was found that the most stable of them was E5.0/C75, followed by E5.0/C50 and then E2.5/C25. It was therefore recommended that further future studies are needed to evaluate the potential of this microcapsule as a natural additive in food, pharmaceuticals and cosmetics.

Structure of Gardenia Blue Pigment and Its Inhibitory Activity on Monoamine Oxidase B / 中国实验方剂学杂志

ObjectiveTo study the chemical structure of gardenia blue pigment and its inhibitory activity against monoamine oxidase B （MAO-B）， in order to seek a potential feasible way for rational utilization and value enhancement of iridoids in Gardeniae Fructus. MethodIridoid glycosides in Gardeniae Fructus were hydrolyzed by cellulase to obtain their aglycones and reacted with amino acids. Then， the products were purified by column chromatography packed with D101 macroporous resin and preparative liquid chromatography to obtain gardenia blue pigments， and the gardenia blue pigments were identified by nuclear magnetic resonance （NMR） and mass spectrometry （MS）. Benzylamine was used as the reaction substrate of MAO-B and in vitro incubated with gardenia blue pigment monomers， high performance liquid chromatography （HPLC） was employed to determine the production of benzaldehyde for evaluating the inhibitory effect of gardenia blue pigments on MAO-B， the mobile phase was methanol （A） -50 mmol·L-1 potassium phosphate buffer （B， pH 3.2） （2∶3）， and the detection wavelength was 245 nm. ResultEight compounds of gardenia blue pigment A-H were synthesized and identified. In MAO-B inhibition test， compared with geniposide， the inhibitory activity of gardenia blue pigment D and E was significantly enhanced （P<0.05）. Compared with the 6β-hydroxygeniposide， the inhibitory activity of gardenia blue pigment G and H was significantly enhanced （P<0.05， P<0.01）. All the four gardenia blue pigments showed better MAO-B inhibitory activity than the prototype compounds. ConclusionGardenia blue pigment is a simple compound formed by one molecule of amino acid and one molecule of iridoid. Some gardenia blue pigments have better MAO-B inhibitory activity than the prototype compounds. The activity of gardenia blue pigment produced by different substrates is different， and the high-value gardenia blue pigment can be prepared based on experimental optimization， which can expand the application range of gardenia blue pigment and enrich the comprehensive utilization of iridoids from Gardeniae Fructus.

Acute retinal pigment epitheliitis: a case presentation and literature review / Epitelite pigmentar retiniana aguda: apresentação de caso e revisão da literatura

ABSTRACT Acute retinal pigment epitheliitis (ARPE) is an idiopathic, self-limiting inflammatory retinal disorder that particularly affects healthy young individuals. The characteristic fundoscopic appearance of the acute retinal pigment epitheliitis includes a fine pigment stippling surrounded by a yellow-white hypopigmented halos in the macula. Although the exact pathogenesis of the disease remains unknown, some reports have suggested a relationship between a viral infection and acute retinal pigment epitheliitis. Acute retinal pigment epitheliitis is a rare disorder, and only single case reports or case series are found in the literature. The clinical and demographic characteristics of patients with this disease are not fully understood because of its rarity. In this study, we searched the literature to collect clinical and demographic features of the reported cases. We detail the characteristics of acute retinal pigment epitheliitis were pointed and discuss the pathogenesis of the disease.(AU)

RESUMO A epitelite pigmentar retiniana aguda (EPRA) é uma doença inflamatória idiopática e autolimitada da retina, que afeta especialmente indivíduos jovens e saudáveis. À fundoscopia, a aparência característica dessa entidade é de um pontilhado fino do pigmento, cercado de halos hiperpigmentados branco-amarelados na mácula. A patogênese exata da doença ainda é desconhecida, mas alguns relatos apontam uma relação entre epitelite pigmentar retiniana aguda e infecções virais. A epitelite pigmentar retiniana aguda é uma condição rara e na literatura há apenas relatos de casos individuais ou séries de casos. As características clínicas e demográficas da doença não são totalmente compreendidas, devido à sua raridade. Para este relato, foi feita uma busca na literatura para coletar os dados clínicos e demográficos dos casos relatados. Finalmente, são apontadas as características da epitelite pigmentar retiniana aguda e discute-se a patogênese da doença.(AU)

Qualidade da carne de frangos da linhagem Label Rouge suplementados com minerais biocomplexados e criados em sistema alternativo / [Meat quality of Label Rouge lineage chicken supplemented with biocomplexed minerals and reared in an alternative system]

Objetivou-se avaliar a influência da suplementação com minerais biocomplexados sobre a qualidade da carne de frangos da linhagem Label Rouge, de ambos os sexos, criados em sistema alternativo. O delineamento experimental foi inteiramente ao acaso (DIC), disposto em esquema fatorial (3x2), sendo três dietas (controle; 0,50ppm de selênio; 0,40ppm de cromo) e dois sexos. Para a estabilidade lipídica, foi utilizado o DIC, disposto em esquema fatorial (3x2x2), sendo três dietas, dois sexos e dois tempos de armazenamento (zero e 12 meses). Os parâmetros avaliados no peito e na coxa foram: pH final, cor (L* - luminosidade, a* - índice de vermelho, b* - índice de amarelo, C* - índice de saturação e h* - ângulo de tonalidade), perda de peso por cozimento (PPC), força de cisalhamento (FC) e estabilidade lipídica. Além disso, foram avaliadas a proporção das formas químicas da mioglobina da coxa e a quantificação do conteúdo de selênio no peito. Não houve efeito isolado da dieta sobre os parâmetros físicos e químicos do peito e da coxa e sobre a concentração de selênio no peito. As fêmeas apresentaram médias superiores de L* (57,57), b* (10,55) e C* (10,60) do peito; e os machos de L* (55,09) da coxa. Entre os machos, as aves alimentadas com a dieta controle e com cromo expressaram valores superiores de FC da coxa em relação às suplementadas com selênio; para o tratamento com cromo, os machos apresentaram média superior de FC da coxa e, em contrapartida, para o tratamento com selênio, as fêmeas manifestaram a maior média. As amostras submetidas ao período de 12 meses de armazenamento mostraram maiores valores de substâncias reativas ao ácido tiobarbitúrico (TBARS) em ambos os cortes. A suplementação com cromo e selênio biocomplexados não provocou alterações na qualidade da carne; as fêmeas demonstraram melhores atributos de qualidade da carne.(AU)

The objective of the present study was to evaluate the influence of the supplementation with biocomplexed minerals on the quality of the meat of label Rouge lineage chicken, of both genders, reared in an alternative system. The experimental design was completely randomized (CRD) in a factorial scheme (3x2), with three diets (control; 0.50ppm of selenium; 0.40ppm of chromium) and two genders. For lipid stability, the CRD arranged in a factorial scheme (3x2x2) was used, being three diets, two genders and two storage times (0 and 12 months). The parameters evaluated in the breast and in the thigh were: final pH, color (L* - luminosity, a* - red index, b* - yellow index, C* - saturation index and h* - tonality angle), weight loss per cooking (WLC), shear force (SF) and lipid stability. In addition, we evaluated the proportion of chemical forms of myoglobin of the thigh and the quantification of selenium content in the breast. There was no isolated effect of diet on the physical and chemical parameters of the breast and thigh and selenium concentration in the breast. Females had higher mean values of L* (57.57), b* (10.55) and C* (10.60) of the breast; and males of L* (55.09) of the thigh. Among males, poultry fed with the control diet and chromium showed higher SF values of the thigh than those supplemented with selenium; for the treatment with chromium, the males presented superior average of SF of the thigh and, in contrast, for the treatment with selenium, the females had the highest average. Samples submitted to the period of 12-month of storage showed higher values of substances reactive to thiobarbituric acid (TBARS) in both cuts. Supplementation with biocomplexed chromium and selenium did not cause changes in meat quality; females presented better attributes of meat quality.(AU)

Evaluation of Sample Preparation Procedures for Determination of Cr(VI) in Cr2O3 Pigments by Vis Spectrophotometry

Abstract Six sample preparation procedures were evaluated for selective extraction of Cr(VI) from commercial samples of chromium oxide green (Cr2O3) pigments prior to formation of its diphenylcarbazone complex [CrDPCO]- for determination by visible spectrophotometry: (I) water-soluble chromium; (II) EPA method 3060A without Mg2+; (III) EPA method 3060A with Mg2+; (IV) Na3PO4 based extraction; (V) method IRSA16 based on acidic extraction and; (VI) Na2CO3 based extraction. Evaluation of the influence of concomitant Cr(III) ions, time and stability of the [CrDPCO]- complex was investigated. Recoveries of soluble and insoluble Cr(VI) species were 86% and 80%, respectively, using procedure (VI). Direct calibration against aqueous standards prepared in the extraction medium was successful for Cr(VI) in the concentration range 0.05-1.50 μg L-1. Limits of detection and quantitation were 0.3 µg g-1 and 1.0 µg g-1, respectively, for 250 mg subsamples/25 mL. Procedure (VI) was applied to the analysis of four commercial samples of Cr2O3 pigments, three determined to have Cr(VI) within compliance limits below 1.0 µg g-1, but one at 16.6 ± 0.6 µg g-1, prohibiting use of this pigment in cosmetic formulations. This sample was conveniently employed to evaluate the accuracy of the method. The recommended procedure is simple and accurate and has been adopted by Tecpar's laboratory of Parana Institute of Technology (Curitiba, Brazil).

Recovery of Red Pigments from Monascus purpureus FTC 5357 by Extraction of Fermented Solids: Operational Conditions and Kinetics

Abstract Solvent extraction of red pigments from fermented solids is reported. The pigments were produced by solid-state fermentation of oil palm frond (OPF) biomass with the food-safe fungus Monascus purpureus FTC 5357. The effects of extraction solvent and other operational conditions (pH, temperature, agitation rate, contact time) on the recovery of pigments are discussed. The recovery was maximized using aqueous ethanol (60% ethanol by vol) as solvent at pH 6, 30 °C, with an extraction time of 16 h and an agitation rate of 180 rpm. A fermented solids dry mass of 1 g was used for each 160 mL of solvent during extraction. The kinetics of extraction were assessed by fitting the experimental data to different models. Peleg's model proved to be the best for describing solid-liquid extraction of the pigments under the above specified conditions. The highest extraction yield of red pigments under the above specified optimal conditions was 207(6.08 AU g(1 dry fermented solids.

Dynamics of change of chlorophyll and carotenoid pigments in leaves of Chrozophora tinctoria species spread in Nakhchivan Autonomous Republic (Azerbaijan Republic)

Background:The pigment composition and the dynamics of pigment accumulation were investigated to determine the degree of adaptation of Chrozophora tinctoria to the environmental conditions in the Nakhchivan Autonomous Republic (AR) of Azerbaijan Republic.Methods:The leaves were collected (transects size 5×5 m, in triplicate) and dried with 15-20 pieces, pigments were separated by solvent. The pigment density was determined by the absorption peak on a spectrophotometer, and then the pigment density was calculated.Results:The chlorophyll pigments in the dry weight of leaves varied between 2-6 mg and carotenoids0.8-1.2 mg/g were found. Also, chlorophyll (a-b) pigments increased in 1.2 times, and carotenoids decreased in 3.6 times depending on environmental conditions.Conclusions:Photosynthetic apparatus of the species actives in all cenosis with response adaptive to changing of environmental conditions depends from the time of leaf collection for the Chrozophora tinctoria species as shown results of researches

Melanin bleaching with potassium permanganate for melanocytic lesions diagnosis in veterinary medicine / Despigmentação de melanócitos com permanganato de potássio para diagnóstico de lesões melanocíticas na medicina veterinária

The aim of this study was to microscopically re-evaluate the melanocytic lesions diagnoses established by the Animal Pathology Laboratory of the Federal University of Uberlândia, in the state of Minas Gerais, Brazil, over a period of eleven years; in addition, to perform a comparative analysis between the conventional histopathological (CH) method and the use of the melanin bleaching (MB) technique with potassium permanganate, sulfuric acid, and oxalic acid solutions. The results of the MB method presented a disagreement in 24.32% of the diagnosis previously by CH, with low agreement (61.0%) and low Kappa coefficient (0.2267). Melanoma was the most frequent lesion, more frequent in elderly and non-breed female dogs. The most frequent melanoma location was in the cutaneous tissue. The presence or absence of a pagetoid spread in cutaneous samples, distribution of melanin, pattern of cell layout, cell morphology, degree of cellular atypia, and the number of mitoses verified after MB were the most important criteria to confirm the diagnosis of malignancy or benignity of the lesions. Evaluating pathologists considered MB to be essential for the majority of diagnoses and an efficient complementary method for the diagnosis of melanocytic lesions, even in cases with a moderate degree of pigmentation.(AU)

Este estudo objetivou reavaliar microscopicamente os diagnósticos de lesões melanocíticas estabelecidos pelo setor de Patologia Animal da Universidade Federal de Uberlândia, em um período de 11 anos, e, com base nesse levantamento, realizar uma análise comparativa entre o método histopatológico convencional (HC) e o método de despigmentação de melanócitos (DM) com permanganato de potássio, ácido sulfúrico e ácido oxálico. A DM revelou discordância em 24,32% dos diagnósticos previamente estabelecidos por HC, apresentando baixa concordância (61,0%) e baixo valor de coeficiente Kappa (0,2267). A alteração mais frequente foi o melanoma, com maior ocorrência em cadelas idosas sem raça definida (SRD). A localização mais frequente dos melanomas foi cutânea. A presença ou ausência de disseminação pagetoide nos casos cutâneos, a forma de distribuição da melanina, o padrão de disposição das células, a morfologia celular, o grau de atipia celular e a quantidade de mitoses verificada após a despigmentação foram critérios de elevada importância para firmar o diagnóstico quanto à malignidade ou benignidade da lesão. A despigmentação foi considerada pelos patologistas avaliadores como essencial para o diagnóstico na maioria dos casos, o que leva a concluir que ela constitui um método complementar eficiente no diagnóstico das lesões melanocíticas, mesmo em casos com grau moderado de pigmentação.(AU)

The Influence of Harmful Substances on the Pigments of Leaves of Decorative Trees

Republic of Uzbekistan the composition of the atmospheric air on the main streets, the level of traffic on these streets, the selection of resistant species of phenol trees for growing ornamental trees, and the laws of changing the amount of pigments in these trees depending on the streets and season. At the same time, the types of ornamental trees and their resistance to pollution from the pollution of carbon dioxide, nitrogen oxides and sulfur dioxide are mainly represented by atmospheric air pollution.

Antibacterial pigment production by Serratia marcescens using different casein types obtained from milk / Producción de un pigmento antibacteriano con Serratia marcescens usando diferentes tipos de caseína obtenida de leche

ABSTRACT Prodigiosin, a pigment produced by Serratia marcescens inhibits the growth of different microorganisms and the proliferation of some human cancer cell lines. Prodigiosin is usually produced by fermentations of substrates such as starch and proteins, and pigment yield depends on the concentration of the carbon source, stirring speed of cultures, temperature and time of incubation, nitrogen sources, and pH of medium. In the present study, pigment production was assessed using two types of casein as substrate; pigment yield was greater when casein precipitated with vinegar was used as substrate than when high purity casein was. The maximum prodigiosin production was achieved with 10 g/L casein at pH 8.0. Additionally, supplementation of culture media with glucose was found to considerably decrease prodigiosin production and growth inhibition of Staphylococcus aureus, which is directly related to pigment yield. Production in stirred-tank bioreactor at 0.75 vvm aeration was higher than that at 0.5 and 1.0 vvm. Substrate type, concentration and pH affected pigment production in Erlenmeyer flasks, whereas aeration rate influenced pigment production in a stirred-tank bioreactor.

RESUMEN La prodigiosina es un pigmento producido por Serratia marcescens que inhibe el crecimiento de diferentes microorganismos y la proliferación de algunas líneas celulares de cáncer humano. La prodigiosina generalmente se produce a través de fermentaciones con sustratos como almidón y proteínas y su producción depende de la concentración de la fuente de carbono, la velocidad de agitación en los cultivos, la temperatura y el tiempo de incubación, las fuentes de nitrógeno y el pH del medio. En este trabajo se evaluó la formación de pigmento usando dos tipos de caseína como sustrato; la producción de pigmento fue mayor cuando se utilizó caseína precipitada con vinagre que cuando se utilizó caseína de alta pureza. La máxima concentración de prodigiosina se logró con 10 g/L de caseína a un pH de 8.0. Se encontró además que suplementar los medios de cultivo con glucosa disminuye considerablemente la producción de prodigiosina y la inhibición de Staphylococcus aureus, la cual está relacionada directamente con la producción de pigmento. La producción en biorreactor de tanque agitado con una aireación de 0.75 vvm fue mayor que con 0.5 y 1.0 vvm. El tipo de sustrato, la concentración y el pH afectaron la producción de pigmento en Erlenmeyer mientras que la velocidad de aireación influyó sobre la producción de pigmento en biorreactor de tanque agitado.

An Implication Of Actinomycetes On Human Well-Being: A Review

This review conceptualizes about the actinomycetes and its contribution to human health by playing a key role as bioactive secondary metabolites, such as enzymes, antibiotics and pigments, leading to their diverse applications and use in various industries. These searches have been uncommonly successful, and around 66% of naturally happening antibiotics, including many medically important, have been isolated from actinomycetes. The speedy occurrence of antimicrobial resistance among pathogens has led to a renewed interest to search for novel antimicrobial agents, but these antibiotics are not enough for the treatment of all diseases because there is a berserk requirement for a novel actinomycetes to combat against the antibiotic-resistant strains of pathogenic microorganisms, which are quickly expanding bit by bit. Actinomycetes are the important providers to the pharmaceutical and other industries and are well known for their capacity to produce secondary metabolites many of which are active against pathogenic microorganisms.

Responses of antioxidant enzymes, photosynthetic pigments and carbohydrates in micropropagated Pitcairnia encholirioides L.B. Sm. (Bromeliaceae) under ex vitro water deficit and after rehydration / Respostas de enzimas antioxidantes, pigmentos fotossintéticos e carboidratos em Pitcairnia encholirioides L.B. Sm. (Bromeliaceae) micropropagadas, sob déficit hídrico ex vitro e após reidratação

Abstract In this study, the activities of antioxidant enzymes, photosynthetic pigments, proline and carbohydrate contents in Pitcairnia encholirioides under ex vitro conditions of water deficit were evaluated. Results show that plants under progressive water stress, previously in vitro cultured in media supplemented with 30 g L-1 sucrose and GA3, accumulated more proline and increased peroxidase (POD) activity and the contents of photosynthetic pigments and carbohydrates. For plants previously in vitro cultured with 15 g L-1 sucrose and NAA, no differences were found for proline content and there were reductions in activities of peroxidase (POD), catalase (CAT) and poliphenoloxidase (PPO), and in contents of carbohydrates, with progress of ex vitro water deficit. After rehydration, plants showed physiological recovery, with enzymatic activities and contents of metabolites similar to those found in the controls not submitted to dehydration, regardless of the previous in vitro culture conditions. These results show that micropropagated P. encholirioides has high tolerance to dehydration once in ex vitro conditions, which can ensure the survival of plants from tissue culture when transferred to its natural environment, emphasizing the importance of such biotechnology for the propagation of endangered species.

Resumo Neste estudo, foram avaliadas as atividades de enzimas antioxidantes, pigmentos fotossintéticos, conteúdo de prolina e carboidratos em Pitcairnia encholirioides sob déficit hídrico em condições ex vitro. Os resultados mostraram que as plantas sob estresse hídrico progressivo, previamente cultivadas in vitro em meio de cultura suplementado com 30 g L-1 de sacarose e GA3 acumularam mais prolina e aumentaram a atividade da peroxidase (POD) e os teores de pigmentos fotossintéticos e carboidratos. Para plantas previamente cultivadas in vitro com 15 g L-1 de sacarose e ANA, não foram encontradas diferenças nos conteúdos de prolina e houve reduções nas atividades da peroxidase (POD), catalase (CAT) e polifenoloxidase (PPO), e no conteúdo de carboidratos, com o progresso do déficit hídrico ex vitro. Após a reidratação, as plantas apresentaram recuperação fisiológica, com atividades enzimáticas e conteúdo de metabólitos semelhantes aos encontrados nos controles não sujeitos à desidratação, independentemente das condições de cultivo in vitro. Estes resultados mostram que P. encholirioides micropropagada tem alta tolerância à desidratação uma vez em condições ex vitro, o que pode garantir a sobrevivência de plantas provenientes da cultura de tecidos quando transferidas para seu ambiente natural, enfatizando a importância desta biotecnologia para a propagação de espécies ameaçadas.

Responses of antioxidant enzymes, photosynthetic pigments and carbohydrates in micropropagated Pitcairnia encholirioides L.B. Sm. (Bromeliaceae) under ex vitro water deficit and after rehydration / Respostas de enzimas antioxidantes, pigmentos fotossintéticos e carboidratos em Pitcairnia encholirioides L.B. Sm. (Bromeliaceae) micropropagadas, sob déficit hídrico ex vitro e após reidratação

In this study, the activities of antioxidant enzymes, photosynthetic pigments, proline and carbohydrate contents in Pitcairnia encholirioides under ex vitro conditions of water deficit were evaluated. Results show that plants under progressive water stress, previously in vitro cultured in media supplemented with 30 g L-1 sucrose and GA3, accumulated more proline and increased peroxidase (POD) activity and the contents of photosynthetic pigments and carbohydrates. For plants previously in vitro cultured with 15 g L-1 sucrose and NAA, no differences were found for proline content and there were reductions in activities of peroxidase (POD), catalase (CAT) and poliphenoloxidase (PPO), and in contents of carbohydrates, with progress of ex vitro water deficit. After rehydration, plants showed physiological recovery, with enzymatic activities and contents of metabolites similar to those found in the controls not submitted to dehydration, regardless of the previous in vitro culture conditions. These results show that micropropagated P. encholirioides has high tolerance to dehydration once in ex vitro conditions, which can ensure the survival of plants from tissue culture when transferred to its natural environment, emphasizing the importance of such biotechnology for the propagation of endangered species.

Neste estudo, foram avaliadas as atividades de enzimas antioxidantes, pigmentos fotossintéticos, conteúdo de prolina e carboidratos em Pitcairnia encholirioides sob déficit hídrico em condições ex vitro. Os resultados mostraram que as plantas sob estresse hídrico progressivo, previamente cultivadas in vitro em meio de cultura suplementado com 30 g L-1 de sacarose e GA3 acumularam mais prolina e aumentaram a atividade da peroxidase (POD) e os teores de pigmentos fotossintéticos e carboidratos. Para plantas previamente cultivadas in vitro com 15 g L-1 de sacarose e ANA, não foram encontradas diferenças nos conteúdos de prolina e houve reduções nas atividades da peroxidase (POD), catalase (CAT) e polifenoloxidase (PPO), e no conteúdo de carboidratos, com o progresso do déficit hídrico ex vitro. Após a reidratação, as plantas apresentaram recuperação fisiológica, com atividades enzimáticas e conteúdo de metabólitos semelhantes aos encontrados nos controles não sujeitos à desidratação, independentemente das condições de cultivo in vitro. Estes resultados mostram que P. encholirioides micropropagada tem alta tolerância à desidratação uma vez em condições ex vitro, o que pode garantir a sobrevivência de plantas provenientes da cultura de tecidos quando transferidas para seu ambiente natural, enfatizando a importância desta biotecnologia para a propagação de espécies ameaçadas.

Influence of High Light Intensity and Nitrate Deprivation on Growth and Biochemical Composition of the Marine Microalgae Isochrysis galbana

Abstract Isochrysis galbana is a brown microalgae widely used as a feed for marine organism in aquaculture. The aim of present study is to investigate the growth, biochemical composition, fatty acid profile, photosynthetic parameters and antioxidant activity (radical scavenging activity) of Isochrysis galbana cells cultivated under different levels (sub-optimum; 50 ± 1.5,optimum; 125 ± 2.5 and supra-optimum; 325 ± 3.5 µmol photons m-2 s -1) of photosynthetic active radiation (PAR), and subsequently treated with different concentration of nitrate deprivation (8mM, 2mM and 0.5mM). The experiment was carried out under controlled conditions utilizing a factorial design 3x3 (light intensity and nitrate concentration). The result depicts that PAR positively influences the growth of Isochrysis galbana which is maximum under supra-optimum PAR. Nitrate deprivation (2mM & 0.5mM) induced decline in growth in terms of dry weight is observed as 60.1% & 61.9% in suboptimum and 26.5% and 34.9% in supra-optimum respectively over the values recorded in their respective controls. Supra-optimum PAR decreased primary photosynthetic pigment Chl a and Chl c by 15.7% and 8.5%, whereas carotenoid content increased by 45.9% in supra-optimum PAR which displays potential interest as antioxidant agent in addition to total phenolic content and radical scavenging activity. The results suggest that combined stress of high light and nitrogen deprivation shifts the metabolic physiology from protein synthesis to energy reserve (carbohydrate and lipid) and accumulation of saturated fatty acid on expense of unsaturated fatty acid except docosahexaenoic acid. These valuable compounds exhibit potential applications in mariculture, nutraceutical and biofuel industry.

L-DOPA extradiol dioxigenase de amanita muscaria: revisão da sequência codificadora, expressão heteróloga, caracterização funcional e contextualização filogenética / Amanita muscaria L-DOPA extradiol dioxygenase: coding sequence review, heterologous expression, functional characterization, and phylogenetic context

Extradiol dioxigenases são enzimas que catalisam a clivagem oxidativa de ligações C-C entre grupos hidroxila fenólicos adjacentes utilizando catecóis como substratos. Esta classe de enzimas é bem caracterizada em bactérias, onde catalisam a degradação de compostos aromáticos. Na maioria das plantas Caryophyllales, como a beterraba, primavera e a maravilha, L-3,4-diidroxifenilalanina (L-DOPA) extradiol dioxigenases (DODAs) catalisam a clivagem oxidativa de L-DOPA na posição 4,5 gerando o ácido betalâmico, aldeído precursor das betalaínas, uma classe de pigmentos naturais que substitui as antocianinas na pigmentação dessas espécies. Alguns fungos basidiomicetos também produzem betalaínas, como o agário-das-moscas (Amanita muscaria). Nesse organismo, DODA é capaz de catalisar uma clivagem adicional na posição 2,3 da L-DOPA, formando muscaflavina, um isômero do ácido betalâmico que dá origem a uma outra classe de pigmentos naturais: as higroaurinas. Desde a caracterização do gene dodA, o qual codifica para a DODA de A. muscaria (AMAMU), não existem relatos na literatura que explorem a promiscuidade catalítica desta enzima, sua relação com outras linhagens de DODAs e a síntese quimioenzimática de betalaínas a partir desta enzima. Dessa forma, buscamos contextualizar as relações filogenéticas e funcionais entre AMAMU e diferentes linhagens de DODAs, bem como estabelecer um método que viabilize a clonagem, expressão heteróloga e caracterização funcional destaenzima. As análises filogenéticas revelaram que AMAMU possui uma evolução convergente com DODAs de plantas e bactérias e que, apesar de AMAMU ser funcionalmente homóloga à DODA da bactéria Escherichia coli, esta última apresenta homologia com DODAs de plantas. Logo, não há uma relação direta entre a sequência primária de DODAs e sua função. Nós também demonstramos que não há uma relação entre a expressão de transcritos de BvDODA1, e de seu parálogo BvDODA2, e a diferença de pigmentação entre variedades de beterrabas amarelas e vermelhas. A clonagem da sequência codificadora (CDS) publicada para o gene dodA de A. muscaria resultou na retenção do primeiro íntron, o que impedia a sua expressão. Então, uma nova CDS de 558 nucleotídeos foi proposta para este gene, a qual inclui um códon de início da tradução que se mantém na fase de leitura e codifica para uma proteína de 185 resíduos, 43 a menos que AMAMU. A expressão desta CDS resultou na proteína recombinante AmDODA, capaz de catalisar a síntese de ácido betalâmico e muscaflavina a partir de L-DOPA e D-DOPA. AmDODA possui um tamanho aproximado de 22 kDa, com um pH ótimo de atividade de 8,5 e uma constante de Michaelis (KM) de 3,7 ± 0,9 mmol L-1 e de velocidade máxima (Vmax) de 3,3 ± 0,4 µ mol min-1 mg-1. Sua utilização foi demonstrada na síntese quimioenzimática de betalaínas-modelo com potencial aplicação como sondas para microscopia confocal de fluorescência de dois fótons. Neste contexto, esta Tese explora os aspectos moleculares, bioquímicos e biológicos da DODA do fungo A. muscaria e traz importantes contribuições acerca da pigmentação por betalaínas na natureza

Extradiol dioxigenases are enzymes that catalyze the oxidative cleavage of C-C bonds between adjacent phenolic hydroxyl groups using catechols as substrates. This class of enzymes is well characterized in bacteria, where they catalyze the degradation of aromatic compounds. In most plants of the Order Caryophyllales, such as beet, paperflower and four o'clock flower, L-3,4-dihydroxyphenylalanine (L-DOPA) extradiol dioxygenases (DODAs) catalyze the oxidative 4,5-cleavage of L-DOPA generating the betalamic acid, an aldehyde precursor of the betalains, a class of natural pigments that replaces anthocyanins in the pigmentation of these species. Some basidiomycete fungi also produce betalains, such as the fly agaric (Amanita muscaria). In this organism, DODA is able to catalyze an additional 2,3-cleavage of L-DOPA, yielding muscaflavine, an isomer of betalamic acid that gives rise to another class of natural pigments: the hygroaurins. Since the characterization of the dodA gene, which encodes the A. muscaria DODA (AMAMU), there are no reports in the literature that explore the catalytic promiscuity of this enzyme, its relation to other DODAs and the chemoenzymatic synthesis of betalains from this enzyme. Thus, we seek to contextualize the phylogenetic and functional relationships between AMAMU and different DODA lineages, as well as to establish a method that enable the cloning, heterologous expression and functional characterization of this enzyme. Phylogenetic analysis revealed that AMAMU has a convergent evolutionwith plant and bacterial DODAs and that although AMAMU is functionally homologous to the DODA of the Escherichia coli bacteria, this latter is homologous to the plant DODAs. Therefore, there is no direct relationship between the primary sequence of DODAs and their function. We have also shown that there is no relationship between the expression of BvDODA1 transcripts, and its BvDODA2 paralogue, and the pigment difference between yellow and red beet varieties. Cloning of the published coding sequence (CDS) for the dodA gene of A. muscaria resulted in the retention of the first intron, which prevented its expression. Then, a new CDS of 558 nucleotides was proposed for this gene, which includes a translation start codon that remains in the open reading frame and encodes for a protein 185 residues long, 43 less than AMAMU. Expression of this CDS resulted in the recombinant AmDODA protein, able to catalyze the synthesis of betalamic acid and muscaflavine from L-DOPA and D-DOPA. AmDODA has an approximate size of 22 kDa, with an optimum activity pH of 8.5 and a Michaelis constant (KM) of 3.7 ± 0.9 mmol L-1 and a maximum velocity (Vmax) of 3.3 ± 0.4 µmol min-1 mg-1. Its use was demonstrated in the chemoenzymatic synthesis of betalains-model with potential application as probes for confocal microscopy of two-photon fluorescence. In this context, this thesis explores the molecular, biochemical and biological aspects of the DODA of the fungus A. muscaria and brings important contributions about the pigmentation by betalains in nature

Carotenoid production in sugarcane juice and synthetic media supplemented with nutrients by Rhodotorula rubra l02

ABSTRACT In order for the use of biological carotenoids to become feasible, it is necessary to have adequate low cost sources and improved methods of cultivation. The aim of this study was to evaluate the effect of supplementation with nitrogen, phosphorus, zinc, and magnesium, on the biomass and carotenoid volumetric production by yeast Rhodotorula rubra L02 using a complex medium (sugarcane juice) and synthetic media (sucrose and maltose) as substrates. The experimental design used for each substrate was randomized in blocks with 16 treatments and 3 repetitions. The treatments were compound for 15 different combinations of nutrients (N; Mg; Zn; P, N + Mg; N + Zn; N + P; Mg + Zn; Mg + P; Zn + P; N + P + Zn; N + P + Mg; N + Zn + Mg; P + Zn + Mg; N + Zn + Mg + P) alone and combined, and a control. The results were submitted to analysis of variance and Tukey test at 5% significance level. Among the treatments evaluated, the highest production of dry biomass, with both maltose and sucrose, was observed for Mg (1.60 g/L and 1.94 g/L, respectively). Additionally, another treatment that stood out in terms of biomass production was the control treatment with maltose (1.54 g/L). After the incubation time, killer activity was not observed since there was no formation of inhibition halo around the L02 yeast.

Structural, histochemical and photosynthetic profiles of galls induced by Eugeniamyia dispar (Diptera: Cecidomyiidae) on the leaves of Eugenia uniflora (Myrtaceae) / Perfiles estructurales, histoquímicos y fotosintéticos de las agallas inducidas por Eugeniamyia dispar (Diptera: Cecidomyiidae) en las hojas de Eugenia uniflora (Myrtaceae)

Abstract Gall-inducing insects manipulate the structural, histochemical and physiological profiles of host-plant tissues to develop galls. We evaluated galls induced by Eugeniamyia dispar on the leaves of Eugenia uniflora in an attempt to answer the following questions: (i) How does this gall-inducing insect change the structural and histochemical profiles of the host-plant organ? (ii) Despite structural changes, can gall tissues maintain photosynthetic activity? Starch, proteins, reducing sugars and reactive oxygen species were detected mainly in the nutritive tissue surrounding the larval chamber. Despite structural changes, the galls induced by E. dispar on E. uniflora retain chlorophyllous tissue, although its amount and photosynthetic activity are less than that of non-galled leaves. This reduced photosynthetic activity, in association with the presence of large intercellular spaces, could improve gas diffusion and, consequently, avoid hypoxia and hypercarbia in gall tissue.(AU)

Resumen Los insectos que inducen las agallas manipulan los perfiles estructurales, histoquímicos y fisiológicos de los tejidos de la planta hospedera para su desarrollo. Nosotros evaluamos las agallas inducidas por Eugeniamyia dispar en las hojas de Eugenia uniflora en un intento de responder las siguientes preguntas: (i) ¿Cómo este insecto inductor de agallas cambia los perfiles estructurales e histoquímicos en el órgano de la planta hospedera? (ii) A pesar de las modificaciones estructurales, ¿pueden los tejidos de la agalla mantener la actividad fotosintética? El almidón, las proteínas, los azúcares reductores y las especies reactivas de oxígeno se detectaron principalmente en la capa de tejido nutritivo que rodea a la cavidad larval. A pesar de las modificaciones estructurales, las agallas inducidas por E. dispar en E. uniflora retienen su tejido clorofílico, aunque su cantidad y actividad fotosintética son menores que en las hojas no agalladas. Esta actividad fotosintética reducida, asociado a la presencia de grandes espacios intercelulares, pueden mejorar la difusión de gases y, en consecuencia, evitar la hipoxia y la hipercapnia en los tejidos de las agallas.(AU)

Function analysis of anthocyanidin synthase from Morus alba L. by expression in bacteria and tobacco

Background: Flavonoids are a kind of important secondary metabolite and are commonly considered to provide protection to plants against stress and UV-B for a long time. Anthocyanidin synthase (ANS), which encodes a dioxygenase in the flavonoid pathway, catalyzes the conversion of leucoanthocyanidins to anthocyanidins, but there is no direct evidence indicating that it provides tolerance to stress in plants. Results: To investigate whether ANS can increase tolerance to abiotic stress, MaANS was isolated from mulberry fruits and transformed into tobacco. Our results suggested that the bacterially expressed MaANS protein can convert dihydroquercetin to quercetin. Overexpression of MaANS remarkably increased the accumulation of total flavonoids in transgenic lines and anthocyanins in corollas of flowers. Transgenic lines showed higher tolerance to NaCl and mannitol stress. Conclusions: These results indicated that MaANS participates in various dioxygenase activities, and it can protect plants against abiotic stress by improving the ROS-scavenging ability. Thus, this alternative approach in crop breeding can be considered in the improvement of stress tolerance by enriching flavonoid production in plants

A Study on Acid-Base Indicator Property of Flowers of Gomphrena globosa

Indicators are used to determine the equivalence point in acid � base titrations (neutralization titrations). They exhibit sharp color change with respect to change in pH. Popularly used indicators for neutralization titrations are synthetic chemicals. They are found to have hazardous effects in human body. The highly colored pigments obtained from plants are found to exhibit color changes with variation in pH. A study was done to check the indicator action of aqueous extract of flower pigments and compared with that of already existing synthetic indicators. Extraction was done using hot water and a definite volume of extract was added which gave accurate and reliable results for different types of neutralization titrations - strong base against strong acid, strong acid against a weak base, weak base against strong acid and weak acid against weak base. The work proved to be acceptable in introducing flower pigments as a suitable substitute to the synthetic indicators.