Simultaneous allergen inactivation and detoxification of castor bean cake by treatment with calcium compounds

Ricinus communis L. is of great economic importance due to the oil extracted from its seeds. Castor oil has been used for pharmaceutical and industrial applications, as a lubricant or coating agent, as a component of plastic products, as a fungicide or in the synthesis of biodiesel fuels. After oil extraction, a castor cake with a large amount of protein is obtained. However, this by-product cannot be used as animal feed due to the presence of toxic (ricin) and allergenic (2S albumin) proteins. Here, we propose two processes for detoxification and allergen inactivation of the castor cake. In addition, we establish a biological test to detect ricin and validate these detoxification processes. In this test, Vero cells were treated with ricin, and cell death was assessed by cell counting and measurement of lactate dehydrogenase activity. The limit of detection of the Vero cell assay was 10 ng/mL using a concentration of 1.6 x 10(5) cells/well. Solid-state fermentation (SSF) and treatment with calcium compounds were used as cake detoxification processes. For SSF, Aspergillus niger was grown using a castor cake as a substrate, and this cake was analyzed after 24, 48, 72, and 96 h of SSF. Ricin was eliminated after 24 h of SSF treatment. The cake was treated with 4 or 8% Ca(OH)2 or CaO, and both the toxicity and the allergenic properties were entirely abolished. A by-product free of toxicity and allergens was obtained.

Effect of eccentric training on mitochondrial function and oxidative stress in the skeletal muscle of rats

The objective of the present study was to investigate the effects of eccentric training on the activity of mitochondrial respiratory chain enzymes, oxidative stress, muscle damage, and inflammation of skeletal muscle. Eighteen male mice (CF1) weighing 30-35 g were randomly divided into 3 groups (N = 6): untrained, trained eccentric running (16°; TER), and trained running (0°) (TR), and were submitted to an 8-week training program. TER increased muscle oxidative capacity (succinate dehydrogenase and complexes I and II) in a manner similar to TR, and TER did not decrease oxidative damage (xylenol and creatine phosphate) but increased antioxidant enzyme activity (superoxide dismutase and catalase) similar to TR. Muscle damage (creatine kinase) and inflammation (myeloperoxidase) were not reduced by TER. In conclusion, we suggest that TER improves mitochondrial function but does not reduce oxidative stress, muscle damage, or inflammation induced by eccentric contractions.

Quercetin postconditioning attenuates myocardial ischemia/reperfusion injury in rats through the PI3K/Akt pathway

Quercetin (Que), a plant-derived flavonoid, has multiple benefical actions on the cardiovascular system. The current study investigated whether Que postconditioning has any protective effects on myocardial ischemia/reperfusion (I/R) injury in vivo and its potential cardioprotective mechanisms. Male Sprague-Dawley rats were randomly allocated to 5 groups (20 animals/group): sham, I/R, Que postconditioning, Que+LY294002 [a phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway inhibitor], and LY294002+I/R. I/R was produced by 30-min coronary occlusion followed by 2-h reperfusion. At the end of reperfusion, myocardial infarct size and biochemical changes were compared. Apoptosis was evaluated by both TUNEL staining and measurement of activated caspase-3 immunoreactivity. The phosphorylation of Akt and protein expression of Bcl-2 and Bax were determined by Western blotting. Que postconditioning significantly reduced infarct size and serum levels of creatine kinase and lactate dehydrogenase compared with the I/R group (all P<0.05). Apoptotic cardiomyocytes and caspase-3 immunoreactivity were also suppressed in the Que postconditioning group compared with the I/R group (both P<0.05). Akt phosphorylation and Bcl-2 expression increased after Que postconditioning, but Bax expression decreased. These effects were inhibited by LY294002. The data indicate that Que postconditioning can induce cardioprotection by activating the PI3K/Akt signaling pathway and modulating the expression of Bcl-2 and Bax proteins.

Liver cancer stem cells are selectively enriched by low-dose cisplatin

Accumulating evidence has indicated the importance of cancer stem cells in carcinogenesis. The goal of the present study was to determine the effect of low-dose cisplatin on enriched liver cancer stem cells (LCSCs). Human hepatoblastoma HepG2 cells were treated with concentrations of cisplatin ranging from 1 to 5 μg/mL. Cell survival and proliferation were evaluated using a tetrazolium dye (MTT) assay. LCSCs were identified using specific markers, namely aldehyde dehydrogenase-1 (ALDH1) and CD133. The percentage of ALDH1+ or CD133+ cells was examined by flow cytometric analysis. The expression of ALDH1 and/or CD133 in HepG2 cells was determined by immunocytochemical analysis. Low-dose cisplatin treatment significantly decreased cell survival in HepG2 cells after 24 or 72 h. However, the percentage of LCSCs in the surviving cells was greatly increased. The percentage of ALDH1+ or CD133+ cells was increased in a time- and dose-dependent manner after treatment with 1-4 μg/mL cisplatin, whereas 5 μg/mL cisplatin exposure slightly reduced the number of positive cells. These findings indicate that low-dose cisplatin treatment may efficiently enrich the LCSC population in HepG2 cells.

Successful domino liver transplantation in maple syrup urine disease using a related living donor

Maple syrup urine disease (MSUD) is an autosomal recessive disease associated with high levels of branched-chain amino acids. Children with MSUD can present severe neurological damage, but liver transplantation (LT) allows the patient to resume a normal diet and avoid further neurological damage. The use of living related donors has been controversial because parents are obligatory heterozygotes. We report a case of a 2-year-old child with MSUD who underwent a living donor LT. The donor was the patient's mother, and his liver was then used as a domino graft. The postoperative course was uneventful in all three subjects. DNA analysis performed after the transplantation (sequencing of the coding regions of BCKDHA, BCKDHB, andDBT genes) showed that the MSUD patient was heterozygous for a pathogenic mutation in the BCKDHB gene. This mutation was not found in his mother, who is an obligatory carrier for MSUD according to the family history and, as expected, presented both normal clinical phenotype and levels of branched-chain amino acids. In conclusion, our data suggest that the use of a related donor in LT for MSUD was effective, and the liver of the MSUD patient was successfully used in domino transplantation. Routine donor genotyping may not be feasible, because the test is not widely available, and, most importantly, the disease is associated with both the presence of allelic and locus heterogeneity. Further studies with this population of patients are required to expand the use of related donors in MSUD.

Putative role of ischemic postconditioning in a rat model of limb ischemia and reperfusion: involvement of hypoxia-inducible factor-1α expression

Hypoxia-inducible factor-1α (HIF-1α) is one of the most potent angiogenic growth factors. It improves angiogenesis and tissue perfusion in ischemic skeletal muscle. In the present study, we tested the hypothesis that ischemic postconditioning is effective for salvaging ischemic skeletal muscle resulting from limb ischemia-reperfusion injury, and that the mechanism involves expression of HIF-1α. Wistar rats were randomly divided into three groups (n=36 each): sham-operated (group S), hindlimb ischemia-reperfusion (group IR), and ischemic postconditioning (group IPO). Each group was divided into subgroups (n=6) according to reperfusion time: immediate (0 h, T0), 1 h (T1), 3 h (T3), 6 h (T6), 12 h (T12), and 24 h (T24). In the IPO group, three cycles of 30-s reperfusion and 30-s femoral aortic reocclusion were carried out before reperfusion. At all reperfusion times (T0-T24), serum creatine kinase (CK) and lactate dehydrogenase (LDH) activities, as well as interleukin (IL)-6, IL-10, and tumor necrosis factor-α (TNF-α) concentrations, were measured in rats after they were killed. Histological and immunohistochemical methods were used to assess the skeletal muscle damage and HIF-1α expression in skeletal muscle ischemia. In groups IR and IPO, serum LDH and CK activities and TNF-α, IL-6, and IL-10 concentrations were all significantly increased compared to group S, and HIF-1α expression was up-regulated (P<0.05 or P<0.01). In group IPO, serum LDH and CK activities and TNF-α and IL-6 concentrations were significantly decreased, IL-10 concentration was increased, HlF-1α expression was down-regulated (P<0.05 or P<0.01), and the pathological changes were reduced compared to group IR. The present study suggests that ischemic postconditioning can reduce skeletal muscle damage caused by limb ischemia-reperfusion and that its mechanisms may be related to the involvement of HlF-1α in the limb ischemia-reperfusion injury-triggered inflammatory response.

Aplicação de revestimentos comestíveis em pêssego (Prunus persica)

O objetivo do trabalho foi avaliar a influência de revestimentos comestíveis sobre a vida-de-prateleira, perda de massa, atividade metabólica e contaminação microbiológicas em pêssegos in natura. Foram utilizados pêssegos brancos cv. Perola de Mairinque (IAC 769-8) revestidos com soluções de 1% de goma gelana (p/v) e 1% goma gelana + 1% sorbitol (p/v), embalados em caixas de papelão e armazenados a 10ºC / 80-85% UR por 16 dias. Frutos sem revestimento serviram de controle. Os revestimentos à base de goma gelana e gelana+sorbitol não alteraram a atividade fisiológica dos pêssegos, medida através da taxa de respiração, teor de sólidos solúveis e acidez titulável ao longo da armazenagem. Não houve diferença entre as taxas de perda de massa, que ficaram em torno de 1,4g/100g.dia. Os revestimentos não alteraram a aceitação dos pêssegos em termos de sabor e em termos de aparência, o controle obteve notas superiores aos revestidos devida a opacidade que o revestimento conferiu aos frutos. Na contagem total de microrganismos aeróbios mesófilos e bolores e leveduras não houve diferença entre os tratamentos ao longo da armazenagem, mas em termos de microrganismos psicrotróficos os frutos revestidos apresentaram uma menor contagem em relação ao controle. Revestimentos à base de goma gelana e gelana+sorbitol não aumentaram a vida-de-prateleira de pêssegos armazenados sob refrigeração devido à composição do revestimento, que não reduziu a perda de massa nem a atividade fisiológica dos frutos.

Determinação simultânea de açúcares e polióis por cromatografia líquida de alta eficiência (CLAE-IR) em sorvetes de baixas calorias ("diet"/ "light")

Um método simples e rápido para a preparação da amostra e quantificação simultânea de açúcares e polióis em sorvetes "diet"/"light" por CLAE-IR foi investigado. Os açúcares frutose, glicose, lactose, maltose, sacarose, juntamente com glicerol e sorbitol foram separados. Diferentes variáveis foram testadas na preparação da amostra e nas condições cromatográficas para a separação dos componentes. A melhor condição para a separação dos analitos da matriz foi obtida através de duas extrações consecutivas com água : etanol (1:8 v/v, seguida de 1:4 v/v). Para a separação cromatográfica em uma coluna CLC-NH2 , a fase móvel foi composta de acetonitrila: água (77,5:22,5 v/v) na vazão de 1mL/min, a 30 ºC. O tempo de corrida foi menor do que 20 min. Amostras de sorvetes "diet"/"light" de 3 diferentes marcas, com sabores de morango, chocolate, flocos e baunilha, recolhidas em Campinas-SP, apresentaram valores de açúcares totais entre 9-15%. Somente os sorvetes com sabor morango apresentaram frutose (0,1-0,5%), e somente aqueles sorvetes com sabor de flocos mostraram a presença de sacarose em maiores quantidades. Os maiores valores de açúcares foram encontrados para lactose, independente da marca ou sabor testados (3,5-10%). As quantidades de sorbitol variaram de 3-4%. A mesma análise foi realizada com sorvete normal (controle) que apresentou valor de açúcares totais de aproximadamente 29%, sendo que destes 16,8%, foi sacarose. Não foram detectados polióis. Na amostra de sorvete controle foi realizado teste de recuperação de sorbitol (99,3%). Os valores encontrados foram comparados com os valores permitidos pela legislação.

Influence of carbon source and the fermentation process on levan production by Zymomonas mobilis analyzed by the surface response method

The aim of this study is to assess sugar cane juice and sucrose as substrates, the batch and fed batch processes and their interaction in the levan production using a complete factorial design. Zymomonas mobilis was cultivated in different sugar cane juice and sucrose concentrations in two fermentation processes at 25 °C for 20 h. A complete factorial design (2³) was used to analyze the effects of the type and concentration of the substrate, as well as the batch and fed batch processes. A complete second factorial design (2²) was used to observe the importance of sugar cane juice. The results indicated that the batch process improved the levan production reaching 40.14 g/L. The addition of sugar cane juice was not statistically significant for levan formation, however sugar cane juice stimulated biomass, sorbitol and ethanol production. The best medium for levan production was 150 g/L sucrose in batch.

Qualidade pós-colheita de nêsperas submetidas ao armazenamento sob baixa temperatura e atmosfera modificada

A curta vida-de-prateleira da nêspera (Eryobotria japonica Lindl.) é um dos fatores que encarecem o preço final deste fruto constituindo um obstáculo para o aumento da sua produção e popularização de seu consumo. Neste contexto, o investimento em estratégias de armazenamento representa uma contribuição para a cadeia de produção da nêspera. Assim, o objetivo deste trabalho é apresentar uma alternativa de baixo custo para a extensão do tempo de armazenamento deste fruto. Nêsperas da cultivar Precoce de Itaquera, maduras, foram acondicionadas em bandejas de poliestireno teraftalato e embaladas com filme de polivinilcloreto de 14 µm de espessura. Os frutos foram armazenados em diferentes temperaturas (18 e 6 °C) na presença e ausência de sachês de permanganato de potássio para remoção do etileno. Durante o armazenamento, os frutos foram periodicamente analisados quanto à produção de etileno e à respiração. Os frutos amostrados para análises posteriores foram descascados e a polpa congelada. Os frutos foram também monitorados quanto à perda de massa fresca e analisados os teores de glicose, frutose, sacarose e sorbitol por HPAEC-PAD. Os teores de etileno nos grupos armazenados na presença dos sachês foram significativamente menores em comparação aos demais grupos, independente da temperatura de armazenamento. A respiração mostrou dependência da temperatura, sendo menor nos frutos armazenados a 6 °C, independente da presença dos sachês. Os teores de açúcares solúveis apresentaram flutuações durante o armazenamento a baixa temperatura, mas sem alterações significativas em relação aos teores iniciais, com exceção do sorbitol que aumentou. Os efeitos do armazenamento à baixa temperatura, combinados com a presença de sachês absorvedores de etileno, sobre a respiração e os teores de etileno podem ser apontados como relevantes para o aumento na vida-de-prateleira das nêsperas, sem alterações nos níveis de açúcares solúveis, fundamentais para o desenvolvimento do sabor do fruto. A combinação da embalagem com baixas temperaturas e a presença de sachês absorvedores de etileno pode constituir uma estratégia viável para o armazenamento da nêspera, estendendo significativamente a pós-colheita.

Influência dos teores de aveia e de gordura nas características tecnológicas e funcionais de bolos

O presente trabalho objetivou estudar o emprego de aveia e de gordura nas características tecnológicas e funcionais de bolos. No estudo foram utilizados os ingredientes flocos e farinha de aveia (Avena sativa L.) do cultivar IAC 7 (30% de flocos grossos tostados e 70% de farinha), previamente selecionados com base na composição química, farinha de trigo, açúcar, sal, fermento químico, emulsificante, leite em pó, ovo inteiro, sorbitol e aroma de baunilha. O experimento foi realizado em delineamento composto central rotacional aplicável à metodologia de superfície de resposta (MSR), sendo os teores de aveia (flocos, farinha) e gordura estabelecidos como variáveis independentes. Os resultados de densidade da massa, viscosidade da massa, composição química, energia metabolizável, volume específico, atividade de água, características internas, porosidade e cor foram tratados por análise de regressão múltipla, obtendo-se modelo matemático de segunda ordem com os termos lineares, quadráticos e de interação significativos. A aveia pode ser utilizada na elaboração de bolos de valor calórico reduzido e como fonte de fibras alimentares atendendo às características de alimento funcional. A densidade da massa variou com a quantidade de gordura da formulação, enquanto que a viscosidade, com a concentração de aveia. A porosidade do miolo foi influenciada pelas concentrações de aveia e de gordura utilizadas nas formulações. Com a elevação do teor de aveia e a diminuição de gordura nas formulações ocorreu um aumento da área, diâmetro e perímetro médios dos alvéolos.

Study of bacteria Gluconobacter sp.: isolation, purification, phenotypic and molecular identification

The importance of the study of acetic bacteria, on species of the Gluconobacter genus is based on its industrial application, as these possess the capacity of bioconversion of sorbitol to sorbose, enabling the process of vitamin C production. The study involved samples collected in industries of soft drinks, flowers, fruits and honey, followed by purification, phenotypic identification, molecular identification with the use of primer defined from Nucleotide Sequence Database consultation. Strains preserved were identified as members of the Acetobacteraceae family, Gluconobacter genus. 110 strains had been isolated of substrate: Pyrostegia venusta (ker-gawler), honey, Vitis vinifera (grape), Pyrus communis (pear), Malus sp. (apple) and in two samples of soft drinks. Of this total 57 strains had been recovered in manitol medium (manitol, yeast extract, peptone), 12 in YMG medium (glucose, manitol, yeast extract, ethanol, acetic acid), 41 in enrichment medium (De Ley and Swings) and later in the GYC medium (glucose, yeast extract and calcium carbonate). 68 strains were identified as Gram negative bacilli rods. Of these, 31 were characterized biochemically as belonging to the Acetobacteriaceae family as they were catalase positive, oxidase negative and producers of acid from glucose. The characterization of these strains was complemented with the biochemistry tests: gelatin liquefaction, nitrate reduction, indole and H2S production, oxidation of ethanol to acetic acid and molecular tests for genus identification. Only eight strains were characterized as pertaining to the Gluconobacter genus. The strains are maintained in collection cultures at the Microbiology Laboratory of the Biology Department at the São Paulo State University (UNESP) in Assis, stored in malt extract at -196 ºC.

Microorganisms screening for limonene oxidation

Limonene is a monoterpene obtained in large amounts from essential oils and is used as a raw material for the synthesis of flavors and fine chemicals. Several pathways or routes for the microbial degradation of limonene making use of the cytochrome P450-dependent monooxygenases have been described. In this study, we present a fermentative screening of microorganisms in order to verify their ability to perform the desirable conversion. In parallel, the PCR technique was used to select the microorganisms that contain the limC gene, which is responsible for the conversion of carveol to carvone. The microorganisms selected by PCR were not able to bioconvert limonene. From this result, we can suppose that these strains do not have the gene that codifies the enzyme responsible for the transformation of limonene into carveol. The results obtained in the fermentative screening showed that 4 microorganisms were able to bioconvert limonene into carveol. In addition, the amplification results showed the presence of fragments of 800 pb, expected for the limC gene. Therefore, the results obtained in the bioconversion and evaluation of the limC gene did not allow a correlation showing that these strains do not contain all the enzymes responsible for the conversion of limonene to carvone.

Development and characterization of orally-disintegrating films for propolis delivery

This study aimed at evaluating the effect of different concentrations of hydrolyzed collagen (HC) on the properties of an orally disintegrating film containing propolis ethanol extract (PEE) as an active component. The films were evaluated in terms of total phenols, mechanical properties, solubility, contact angle, disintegration time, and microstructure. The films were prepared by casting with 2 g of protein mass (gelatin and HC), 30 g of sorbitol/100 g of protein mass, and 100 g of PEE/100 g of protein mass. HC was incorporated at concentrations of 0, 10, 20, and 30 g/100 g of protein mass. It was found that increased concentrations of HC reduced tensile strength and increased elongation; however, all films showed plastic behavior. An increase in solubility at 25 ºC, a reduction in the contact angle, and disintegration time were also observed. Thus, higher concentrations of collagen led to more hydrophilic and more soluble polymeric matrices that showed shorter dissolution time, favoring the use of these materials as carriers for active compounds to be delivered in the oral cavity.

Development and optimization of a HPLC-RI method for the determination of major sugars in apple juice and evaluation of the effect of the ripening stage

The sugars in apple juice prove its authenticity and its sensory and nutritional properties. The aim of this study was to develop and validate a simple analytical method using high performance liquid chromatography with refractive index detection (HPLC-RI) to determinate and quantify the sugars sucrose, D-glucose, D-fructose, and D-sorbitol polyol in apple juices, as well as to analyze the juices from the Fuji suprema and Lis Gala cultivars at three ripening stages. The analytical performance parameters evaluated indicated that the method was specific for the compounds analyzed, and the linearity of the calibration curves of sugars showed high correlation coefficients (close to 1.0). The limits of detection and quantification are consistent with recommendations available in the literature for this type of matrix. Sample preparation is simple and generates small amount of residues. Over 70% of the sugars were determined in the juices of apples at the pre-ripe stage, with an increase during senescence. This method is applicable for the determination of sugars in juices and evaluation of apple ripening.