Proteinograma sérico de bovinos da raça Curraleir

Estudou-se o perfil eletroforético das proteínas séricas de bovinos sadios da raça Curraleiro por meio da técnica de eletroforese em gel de acrilamida contendo dodecil sulfato de sódio. Utilizaram-se amostras de soro sanguíneo de 228 bovinos da raça Curraleiro, 51 machos e 177 fêmeas, com idades entre sete meses e 12 anos, pertencentes a dois rebanhos localizados nos Estados de Goiás e Tocantins. Foram quantificadas proteína total e concentração plasmática de fibrinogênio. Verificaram-se variações nas concentrações das diferentes frações proteicas. Foram detectadas 26 proteínas e identificadas 10 delas. A ceruloplasmina esteve ausente em 78,1% dos indivíduos, e a α-antitripsina não foi detectada em nenhum animal. Proteína total, globulina, IgA, IgG e fibrinogênio aumentaram com a idade e houve correlação positiva entre os níveis séricos de haptoglobina e α1-glicoproteína ácida.

Perfil eletroforético do proteinograma sérico de eqüinos com obstrução experimental do cólon menor

Avaliaram-se as alterações do proteinograma sérico de eqüinos submetidos à isquemia e reperfusão do cólon menor por distensão intraluminal. Foram utilizados 10 animais submetidos à laparotomia pelo flanco, em posição quadrupedal, para a indução de obstrução no cólon menor durante um período de quatro horas. Cinco animais foram instrumentados, mas sem distensão (grupo controle - G1). em cinco outros animais, foi realizada isquemia mural por distensão do cólon menor via manguito inflado com 40mmHg (grupo distendido - G2). Foram colhidas amostras de sangue antes da intervenção cirúrgica (M1), com 4 horas da colocação do manguito (M2) e com 3 horas (M3) e 12 horas (M4) de reperfusão. Após centrifugação e fracionamento das amostras, as proteínas de fase aguda foram separadas por eletroforese em gel de poliacrilamida contendo SDS-PAGE, e suas concentrações determinadas por densitometria computadorizada. Foram encontradas 19 proteínas no fracionamento eletroforético, com peso molecular variando de 185.000 a 14.000 Daltons (Da). Os pesos moleculares encontrados, correspondentes às proteínas mais conhecidas, foram ceruloplasmina, 130.000 Da; proteína C-reativa, 122.000 Da; transferrina, 85.000 Da; α1-antitripsina, 61.000 Da; haptoglobina, 47.000 Da; e glicoproteína ácida, 40.000 Da. Os resultados mostram que proteínas de fase aguda se alteram após o trauma cirúrgico.

Proteinograma sérico de bezerros recém-nascidos da raça Holandesa obtido por eletroforese em gel de poliacrilamida

The serum protein concentration of newborn Holstein calves determined by means of sodium dodecyl sulphate-polyacrylamide (SDS-PAGE) was studied. Blood samples from 40 healthy newborn calves were obtained 48 hours after birth. Calves had been given 3 liters of colostrum within 2 hours after birth, following by dose corresponding by 15% of animal weight each 24 hours. The results showed three different proteinograms: 19 calves had 14 proteins with molecular weights (MW) ranging from 28,000 D to 170,000D (proteinogram 1); 11 calves had 14 proteins with MW ranging from 18,000 to 170,000 D (proteinogram 1); and 10 calves had 12 proteins with MW ranging from 28,000 D to 170,000 D (proteinogram 3). The three groups presented similar IgG levels. The highest serum concentration of ceruloplasmin were verified in proteinogram 3, which had the lowest serum level of protein with MW 58,000D. It was verified a1-antitrypsin only in proteinogram 2, which had no proteins with MW of 42,000 D and 37,000D. The highest serum concentrations of IgA and protein with MW 58,000 D, and the lowest serum levels of transferrin, haptoglobin, and acid glycoprotein were verified in proteinogram 3. Measurement of serum protein concentrations by SDS-PAGE may be useful in monitoring the occurrence of hypogammaglobulinemia and the neonatal disease in calves.

Passive immunity transfer and serum constituents of crossbred calves

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Proteinograma e teores de cobre, ferro e zinco no soro sanguíneo de ovelhas da raça Santa Inês com mastite experimental por Staphylococcus aureus

Costa N.A., Simao L.C.V., Santos R.A., Afonso J.A.B., Fagliari J.J., Cardoso E.C., Soares P.C. & Mendonca C.L. 2010. [Proteinogram and serum concentrations of copper, iron and zinc in Santa Ines ewes with Staphylococcus aureus experimentally induced mastitis.] Proteinograma e teores de cobre, ferro e zinco no soro sanguineo de ovelhas da raca Santa Ines com mastite experimental por Staphylococcus aureus. Pesquisa Veterinaria Brasileira 30(5):435-442. Clinica de Bovinos, Campus Garanhuns, Universidade Federal Rural de Pernambuco (UFRPE), Avenida Bom Pastor s/n, Boa Vista, Caixa Postal 152, Garanhuns, PE 55292-901, Brazil. E-mail: carlalopes.mendonca@gmail.comThe aim of the present study was to evaluate the effect of Staphylococcus aureus experimentally induced mastitis on proteinogram and serum concentrations of cupper, iron and zinc levels of Santa Ines primiparous ewes. The right mammary gland of ten healthy ewes was inoculated with 1,0x10(4) UFC/mL of S. aureus. Clinical examination and determination of serum concentrations of proteins by electrophoresis in polyacrylamide gel (SDS-PAGE), cupper, iron and zinc, as well plasma level of fibrinogen were measured before the inoculation (control) and 12h, 24h, 36h, 48h, 60h, 72h, 84h, 96h, 108h, 120h, 132h, 168h, 180h, 288h and 336h after bacteria inoculation. All animals experimentally infected presented clinical mastitis and subsequent loss of mammary gland function. The electrophoretogram allowed the identification of 23 proteins with molecular weights (MW) ranged from 26.000 to 185.000 daltons (Da) including acute-phase proteins, IgG and IgA. A significant increase (P<0,05) in haptoglobin, ceruloplasmin, IgG and IgA concentrations was observed. Antitrypsin and acid glicoprotein concentrations did not alter. The levels of iron and zinc decreased and the cupper concentration increased. A positive correlation between plasma fibrinogen and serum ceruloplasmin (r=0.74), haptoglobin (r=0.62) and IgA (r=0.62) was also identified. Results showed the importance of ceruloplasmin and haptoglobin as acute-phase proteins in ewes with intramammary infections and confirms fibrinogen as an inflammatory marker because its high correlation with specific proteins. The alterations in the serum levels of Cu, Fe and Zn suggest the action of inflammatory mediators triggered by S. aureus.

Serum protein concentrations, including acute phase proteins, in calves experimentally infected with Salmonella Dublin

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Influência do puerpério sobre o proteinograma sérico de caprinos da raça Saanen obtido por eletroforese em gel de poliacrilamida

Aiming to evaluate the puerperal influence on the proteinogram of Saanen goats, 108 samples of blood serum from 12 goats were collected, and the results were presented at nine times: just after parturition, 1, 3, 5, 7, 10, 15, 21 and 30 days after parturition. Total amount of serum proteins were determined by the biuret technique, and the sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was used to the protein fractionation. In this last method, 17 protein bands were observed, from which molecular weights varied between 25 KDa and 275 KDa. In addition, it was possible to identify the following protein fractions: immunoglobulin A (180 KDa), ceruloplasmin (115 KDa), transferrin (79 KDa), albumin (65 KDa), heavy-chain immunoglobulin G (58 KDa), haptoglobin (45 KDa), acid glycoprotein (37 KDa) and light-chain immunoglobulin G (28 KDa). Another 9 nonidentified protein fractions presented, each molecular weights equal to 275 KDa, 140 KDa, 125 KDa, 103 KDa, 95 KDa, 41 KDa, 35 KDa, 30 Kda and 25 KDa. The results allow us to conclude that by the first week of puerperium, an improvement of acid glycoprotein occurs, whereas those others protein fractions do not suffer any puerperal influence.

Proteínas ligantes do insulin-like growth factor (IGFBPs) e dominância folicular em vacas Bos taurus indicus puras e cruzadas

O objetivo deste estudo foi identificar um marcador bioquímico de folículos bovinos com diâmetro maior do que 10 mm, o qual poderia ser utilizado para identificar folículos dominantes nesta espécie. Líquido folicular era aspirado de folículos com diâmetros menores do que 5 mm, entre 5 e 10 mm e maiores do que 10 mm, provenientes de ovários de 37 fêmeas bovinas abatidas. Após aspiração, o líquido folicular (LF) era acondicionado em ependorfs etiquetados e congelados. Os padrões eletroforéticos em SDS-PAGE das proteínas do LF foram determinados e verificou-se que os folículos com diâmetro maior do que 10 mm apresentaram um polipeptídeo com PM entre 39 e 43 KDa identificado como a proteína ligante de IGF-3 (IGFBP-3), o qual não foi observado em folículos com diâmetro menor do que 5 mm e entre 5 e 10 mm. Este estudo sugere que este polipeptídeo possa ser utilizado como marcador bioquímico de folículos maiores do que 10 mm.

Culture of osteogenic cells from human alveolar bone: A useful source of alkaline phosphatase

The aim of this study was to obtain membrane-bound alkaline phosphatase from osteoblastic-like cells of human alveolar bone. Cells were obtained by enzymatic digestion and maintained in primary culture in osteogenic medium until subconfluence. First passage cells were cultured in the same medium and at 7, 14, and 21 days, total protein content, collagen content, and alkaline phosphatase activity were evaluated. Bone-like nodule formation was evaluated at 21 days. Cells in primary culture at day 14 were washed with Tris-HCl buffer, and used to extract the membrane-bound alkaline phosphatase. Cells expressed osteoblastic phenotype. The apparent optimum pH for PNPP hydrolysis by the enzyme was pH 10.0. This enzyme also hydrolyzes ATP, ADP, fructose-1-phosphate, fructose-6-phosphate, pyrophosphate and beta-glycerophosphate. PNPPase activity was reduced by typical inhibitors of alkaline phosphatase. SDS-PAGE of membrane fraction showed a single band with activity of similar to 120 kDa that could be solubilized by phospholipase C or Polidocanol. (c) 2007 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.

Blood serum components and serum protein test of Hybro-PG broilers of different ages

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Recuperação e purificação de quitosanases usando adsorção em leito expandido com streamline DEAE com modelagem e simulação usando redes neurais

Expanded Bed Adsorption (EBA) is an integrative process that combines concepts of chromatography and fluidization of solids. The many parameters involved and their synergistic effects complicate the optimization of the process. Fortunately, some mathematical tools have been developed in order to guide the investigation of the EBA system. In this work the application of experimental design, phenomenological modeling and artificial neural networks (ANN) in understanding chitosanases adsorption on ion exchange resin Streamline® DEAE have been investigated. The strain Paenibacillus ehimensis NRRL B-23118 was used for chitosanase production. EBA experiments were carried out using a column of 2.6 cm inner diameter with 30.0 cm in height that was coupled to a peristaltic pump. At the bottom of the column there was a distributor of glass beads having a height of 3.0 cm. Assays for residence time distribution (RTD) revelead a high degree of mixing, however, the Richardson-Zaki coefficients showed that the column was on the threshold of stability. Isotherm models fitted the adsorption equilibrium data in the presence of lyotropic salts. The results of experiment design indicated that the ionic strength and superficial velocity are important to the recovery and purity of chitosanases. The molecular mass of the two chitosanases were approximately 23 kDa and 52 kDa as estimated by SDS-PAGE. The phenomenological modeling was aimed to describe the operations in batch and column chromatography. The simulations were performed in Microsoft Visual Studio. The kinetic rate constant model set to kinetic curves efficiently under conditions of initial enzyme activity 0.232, 0.142 e 0.079 UA/mL. The simulated breakthrough curves showed some differences with experimental data, especially regarding the slope. Sensitivity tests of the model on the surface velocity, axial dispersion and initial concentration showed agreement with the literature. The neural network was constructed in MATLAB and Neural Network Toolbox. The cross-validation was used to improve the ability of generalization. The parameters of ANN were improved to obtain the settings 6-6 (enzyme activity) and 9-6 (total protein), as well as tansig transfer function and Levenberg-Marquardt training algorithm. The neural Carlos Eduardo de Araújo Padilha dezembro/2013 9 networks simulations, including all the steps of cycle, showed good agreement with experimental data, with a correlation coefficient of approximately 0.974. The effects of input variables on profiles of the stages of loading, washing and elution were consistent with the literature

A lectin from the lichenized Basidiomycete Dictyonema glabratum

A lectin with hemagglutinating activity has been isolated from an aqueous extract of the symbiotic phenotype of Dictyonema glabratum and its cyanobacterial photobiont Scytonema sp. The purified lectin had a pi of 6.8 and its molecular mass was investigated by electrospray ionization mass spectrometry, gel filtration and SDS-PAGE, which indicated its native conformation as a dimer formed by two identical subunits of 16540 Da. The lectin is a glycoprotein with a low degree of glycosylation, containing galactose, xylose, glucose and mannose as neutral monosaccharides, in addition to glucosamine, which could indicate both N- and O-linkages. Amino acid analysis showed the predominance of nonpolar residues such as phenylalanine. Agglutination of human erythrocytes required divalent cations, which is affected by addition of EDTA. The lectin was more stable at 30 degreesC or less for at least 1 h and between pH 5.0 and 7.0. Among the various compounds tested for hemagglutination inhibition, N-acetylgalactosamine was the most active. The potential role of this lectin in recognition of the compatible cyanobacterial photobiont is discussed.

Thermal studies on protein isolates of white lupin seeds (Lupinus albus)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Protein Hydrolysis by Immobilized and Stabilized Trypsin

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Isolation, solubility and in vitro hydrolysis of chickpea vicilin-like protein

The chickpea vicilin-like globulin was isolated and chromatographed on Sepharose CL-6B and Sephacryl S-300. The native globulin with a molecular weight of 140 kDa was resolved in Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in seven polypeptide bands in the range of 12.4-67 kDa. The solubility profile of the protein in water and NaCl solutions was typical of a legume globulin. The purified vicilin-like globulin, native and heated, was hydrolyzed by pepsin, trypsin and chymotrypsin. The hydrolysis patterns indicated that the native vicilin-like protein was only partially degraded by the enzymes in comparison with casein. Heating increased its susceptibility to hydrolysis relative to the native form, for all the enzymes. However, the results obtained by the pH-drop method revealed that the in vitro digestibility of the vicilin-like protein was not altered by heating, while 11 S-like and total globulins suffered a small increase, indicating that the structural characteristics of storage globulins may be important factors limiting the protein digestion. (c) 2007 Swiss Society of Food Science and Technology. Published by Elsevier Ltd. All rights reserved.