Effects Of High Pressure Homogenization On The Activity, Stability, Kinetics And Three-dimensional Conformation Of A Glucose Oxidase Produced By Aspergillus Niger.

High pressure homogenization (HPH) is a non-thermal method, which has been employed to change the activity and stability of biotechnologically relevant enzymes. This work investigated how HPH affects the structural and functional characteristics of a glucose oxidase (GO) from Aspergillus niger. The enzyme was homogenized at 75 and 150 MPa and the effects were evaluated with respect to the enzyme activity, stability, kinetic parameters and molecular structure. The enzyme showed a pH-dependent response to the HPH treatment, with reduction or maintenance of activity at pH 4.5-6.0 and a remarkable activity increase (30-300%) at pH 6.5 in all tested temperatures (15, 50 and 75°C). The enzyme thermal tolerance was reduced due to HPH treatment and the storage for 24 h at high temperatures (50 and 75°C) also caused a reduction of activity. Interestingly, at lower temperatures (15°C) the activity levels were slightly higher than that observed for native enzyme or at least maintained. These effects of HPH treatment on function and stability of GO were further investigated by spectroscopic methods. Both fluorescence and circular dichroism revealed conformational changes in the molecular structure of the enzyme that might be associated with the distinct functional and stability behavior of GO.

Structural Connectivity Of The Default Mode Network And Cognition In Alzheimer׳s Disease.

Disconnectivity between the Default Mode Network (DMN) nodes can cause clinical symptoms and cognitive deficits in Alzheimer׳s disease (AD). We aimed to examine the structural connectivity between DMN nodes, to verify the extent in which white matter disconnection affects cognitive performance. MRI data of 76 subjects (25 mild AD, 21 amnestic Mild Cognitive Impairment subjects and 30 controls) were acquired on a 3.0T scanner. ExploreDTI software (fractional Anisotropy threshold=0.25 and the angular threshold=60°) calculated axial, radial, and mean diffusivities, fractional anisotropy and streamline count. AD patients showed lower fractional anisotropy (P=0.01) and streamline count (P=0.029), and higher radial diffusivity (P=0.014) than controls in the cingulum. After correction for white matter atrophy, only fractional anisotropy and radial diffusivity remained significantly lower in AD compared to controls (P=0.003 and P=0.05). In the parahippocampal bundle, AD patients had lower mean and radial diffusivities (P=0.048 and P=0.013) compared to controls, from which only radial diffusivity survived for white matter adjustment (P=0.05). Regression models revealed that cognitive performance is also accounted for by white matter microstructural values. Structural connectivity within the DMN is important to the execution of high-complexity tasks, probably due to its relevant role in the integration of the network.

Changes In Liver Cell Dna Methylation Status In Diabetic Mice Affect Its Ft-ir Characteristics.

Lower levels of cytosine methylation have been found in the liver cell DNA from non-obese diabetic (NOD) mice under hyperglycemic conditions. Because the Fourier transform-infrared (FT-IR) profiles of dry DNA samples are differently affected by DNA base composition, single-stranded form and histone binding, it is expected that the methylation status in the DNA could also affect its FT-IR profile. The DNA FT-IR signatures obtained from the liver cell nuclei of hyperglycemic and normoglycemic NOD mice of the same age were compared. Dried DNA samples were examined in an IR microspectroscope equipped with an all-reflecting objective (ARO) and adequate software. Changes in DNA cytosine methylation levels induced by hyperglycemia in mouse liver cells produced changes in the respective DNA FT-IR profiles, revealing modifications to the vibrational intensities and frequencies of several chemical markers, including νas -CH3 stretching vibrations in the 5-methylcytosine methyl group. A smaller band area reflecting lower energy absorbed in the DNA was found in the hyperglycemic mice and assumed to be related to the lower levels of -CH3 groups. Other spectral differences were found at 1700-1500 cm(-1) and in the fingerprint region, and a slight change in the DNA conformation at the lower DNA methylation levels was suggested for the hyperglycemic mice. The changes that affect cytosine methylation levels certainly affect the DNA-protein interactions and, consequently, gene expression in liver cells from the hyperglycemic NOD mice.

Development And Shelf-life Determination Of Pasteurized, Microfiltered, Lactose Hydrolyzed Skim Milk.

The segment of the world population showing permanent or temporary lactose intolerance is quite significant. Because milk is a widely consumed food with an high nutritional value, technological alternatives have been sought to overcome this dilemma. Microfiltration combined with pasteurization can not only extend the shelf life of milk but can also maintain the sensory, functional, and nutritional properties of the product. This studied developed a pasteurized, microfiltered, lactose hydrolyzed (delactosed) skim milk (PMLHSM). Hydrolysis was performed using β-galactosidase at a concentration of 0.4mL/L and incubation for approximately 21h at 10±1°C. During these procedures, the degree of hydrolysis obtained (>90%) was accompanied by evaluation of freezing point depression, and the remaining quantity of lactose was confirmed by HPLC. Milk was processed using a microfiltration pilot unit equipped with uniform transmembrane pressure (UTP) ceramic membranes with a mean pore size of 1.4 μm and UTP of 60 kPa. The product was submitted to physicochemical, microbiological, and sensory evaluations, and its shelf life was estimated. Microfiltration reduced the aerobic mesophilic count by more than 4 log cycles. We were able to produce high-quality PMLHSM with a shelf life of 21 to 27d when stored at 5±1°C in terms of sensory analysis and proteolysis index and a shelf life of 50d in regard to total aerobic mesophile count and titratable acidity.

Bay 41-2272, A Soluble Guanylate Cyclase Stimulator, Relaxes Isolated Human Ureter In A Standardized In Vitro Model.

To characterize the relaxation induced by BAY 41-2272 in human ureteral segments. Ureter specimens (n = 17) from multiple organ human deceased donors (mean age 40 ± 3.2 years, male/female ratio 2:1) were used to characterize the relaxing response of BAY 41-2272. Immunohistochemical analysis for endothelial and neuronal nitric oxide synthase, guanylate cyclase stimulator (sGC) and type 5 phosphodiesterase was also performed. The potency values were determined as the negative log of the molar to produce 50% of the maximal relaxation in potassium chloride-precontracted specimens. The unpaired Student t test was used for the comparisons. Immunohistochemistry revealed the presence of endothelial nitric oxide synthase in vessel endothelia and neuronal nitric oxide synthase in urothelium and nerve structures. sGC was expressed in the smooth muscle and urothelium layer, and type 5 phosphodiesterase was present in the smooth muscle only. BAY 41-2272 (0.001-100 μM) relaxed the isolated ureter in a concentration dependent manner, with a potency and maximal relaxation value of 5.82 ± 0.14 and 84% ± 5%, respectively. The addition of nitric oxide synthase and sGC inhibitors reduced the maximal relaxation values by 21% and 45%, respectively. However, the presence of sildenafil (100 nM) significantly potentiated (6.47 ± 0.10, P <.05) this response. Neither glibenclamide or tetraethylammonium nor ureteral urothelium removal influenced the relaxation response by BAY 41-2272. BAY 41-2272 relaxes the human isolated ureter in a concentration-dependent manner, mainly by activating the sGC enzyme in smooth muscle cells rather than in the urothelium, although a cyclic guanosine monophosphate-independent mechanism might have a role. The potassium channels do not seem to be involved.

Censored Linear Regression Models For Irregularly Observed Longitudinal Data Using The Multivariate-t Distribution.

In acquired immunodeficiency syndrome (AIDS) studies it is quite common to observe viral load measurements collected irregularly over time. Moreover, these measurements can be subjected to some upper and/or lower detection limits depending on the quantification assays. A complication arises when these continuous repeated measures have a heavy-tailed behavior. For such data structures, we propose a robust structure for a censored linear model based on the multivariate Student's t-distribution. To compensate for the autocorrelation existing among irregularly observed measures, a damped exponential correlation structure is employed. An efficient expectation maximization type algorithm is developed for computing the maximum likelihood estimates, obtaining as a by-product the standard errors of the fixed effects and the log-likelihood function. The proposed algorithm uses closed-form expressions at the E-step that rely on formulas for the mean and variance of a truncated multivariate Student's t-distribution. The methodology is illustrated through an application to an Human Immunodeficiency Virus-AIDS (HIV-AIDS) study and several simulation studies.

Crystal Structure Of (3e)-3-[(4-nitro-phen-oxy)-meth-yl]-4-phenyl-but-3-en-2-one.

In the title compound, C17H15NO4, the conformation about the C=C double bond [1.348 (2) Å] is E with the ketone group almost co-planar [C-C-C-C torsion angle = 7.2 (2)°] but the phenyl group twisted away [C-C-C-C = 160.93 (17)°]. The terminal aromatic rings are almost perpendicular to each other [dihedral angle = 81.61 (9)°] giving the mol-ecule an overall U-shape. The crystal packing feature benzene-C-H⋯O(ketone) contacts that lead to supra-molecular helical chains along the b axis. These are connected by π-π inter-actions between benzene and phenyl rings [inter-centroid distance = 3.6648 (14) Å], resulting in the formation of a supra-molecular layer in the bc plane.

Crystal Structure Of (3e)-3-(2,4-di-nitro-phen-oxy-meth-yl)-4-phenyl-but-3-en-2-one.

In the title compound, C17H14N2O6, the conformation about the C=C double bond [1.345 (2) Å] is E, with the ketone moiety almost coplanar [C-C-C-C torsion angle = 9.5 (2)°] along with the phenyl ring [C-C-C-C = 5.9 (2)°]. The aromatic rings are almost perpendicular to each other [dihedral angle = 86.66 (7)°]. The 4-nitro moiety is approximately coplanar with the benzene ring to which it is attached [O-N-C-C = 4.2 (2)°], whereas the one in the ortho position is twisted [O-N-C-C = 138.28 (13)°]. The mol-ecules associate via C-H⋯O inter-actions, involving both O atoms from the 2-nitro group, to form a helical supra-molecular chain along [010]. Nitro-nitro N⋯O inter-actions [2.8461 (19) Å] connect the chains into layers that stack along [001].

A New Platinum Complex With Tryptophan: Synthesis, Structural Characterization, Dft Studies And Biological Assays In Vitro Over Human Tumorigenic Cells.

A new platinum(II) complex with the amino acid L-tryptophan (trp), named Pt-trp, was synthesized and characterized. Elemental, thermogravimetric and ESI-QTOF mass spectrometric analyses led to the composition [Pt(C11H11N2O2)2]⋅6H2O. Infrared spectroscopic data indicate the coordination of trp to Pt(II) through the oxygen of the carboxylate group and also through the nitrogen atom of the amino group. The (13)C CP/MAS NMR spectroscopic data confirm coordination through the oxygen atom of the carboxylate group, while the (15)N CP/MAS NMR data confirm coordination of the nitrogen of the NH2 group to the metal. Density functional theory (DFT) studies were applied to evaluate the cis and trans coordination modes of trp to platinum(II). The trans isomer was shown to be energetically more stable than the cis one. The Pt-trp complex was evaluated as a cytotoxic agent against SK-Mel 103 (human melanoma) and Panc-1 (human pancreatic carcinoma) cell lines. The complex was shown to be cytotoxic over the considered cells.

Behavior Of Listeria Monocytogenes In A Multi-species Biofilm With Enterococcus Faecalis And Enterococcus Faecium And Control Through Sanitation Procedures.

The formation of mono-species biofilm (Listeria monocytogenes) and multi-species biofilms (Enterococcus faecium, Enterococcus faecalis, and L. monocytogenes) was evaluated. In addition, the effectiveness of sanitation procedures for the control of the multi-species biofilm also was evaluated. The biofilms were grown on stainless steel coupons at various incubation temperatures (7, 25 and 39°C) and contact times (0, 1, 2, 4, 6 and 8days). In all tests, at 7°C, the microbial counts were below 0.4 log CFU/cm(2) and not characteristic of biofilms. In mono-species biofilm, the counts of L. monocytogenes after 8days of contact were 4.1 and 2.8 log CFU/cm(2) at 25 and 39°C, respectively. In the multi-species biofilms, Enterococcus spp. were present at counts of 8 log CFU/cm(2) at 25 and 39°C after 8days of contact. However, the L. monocytogenes in multi-species biofilms was significantly affected by the presence of Enterococcus spp. and by temperature. At 25°C, the growth of L. monocytogenes biofilms was favored in multi-species cultures, with counts above 6 log CFU/cm(2) after 8days of contact. In contrast, at 39°C, a negative effect was observed for L. monocytogenes biofilm growth in mixed cultures, with a significant reduction in counts over time and values below 0.4 log CFU/cm(2) starting at day 4. Anionic tensioactive cleaning complemented with another procedure (acid cleaning, disinfection or acid cleaning+disinfection) eliminated the multi-species biofilms under all conditions tested (counts of all micro-organisms<0.4 log CFU/cm(2)). Peracetic acid was the most effective disinfectant, eliminating the multi-species biofilms under all tested conditions (counts of the all microorganisms <0.4 log CFU/cm(2)). In contrast, biguanide was the least effective disinfectant, failing to eliminate biofilms under all the test conditions.

Multimodal Mri-based Study In Patients With Spg4 Mutations.

Mutations in the SPG4 gene (SPG4-HSP) are the most frequent cause of hereditary spastic paraplegia, but the extent of the neurodegeneration related to the disease is not yet known. Therefore, our objective is to identify regions of the central nervous system damaged in patients with SPG4-HSP using a multi-modal neuroimaging approach. In addition, we aimed to identify possible clinical correlates of such damage. Eleven patients (mean age 46.0 ± 15.0 years, 8 men) with molecular confirmation of hereditary spastic paraplegia, and 23 matched healthy controls (mean age 51.4 ± 14.1years, 17 men) underwent MRI scans in a 3T scanner. We used 3D T1 images to perform volumetric measurements of the brain and spinal cord. We then performed tract-based spatial statistics and tractography analyses of diffusion tensor images to assess microstructural integrity of white matter tracts. Disease severity was quantified with the Spastic Paraplegia Rating Scale. Correlations were then carried out between MRI metrics and clinical data. Volumetric analyses did not identify macroscopic abnormalities in the brain of hereditary spastic paraplegia patients. In contrast, we found extensive fractional anisotropy reduction in the corticospinal tracts, cingulate gyri and splenium of the corpus callosum. Spinal cord morphometry identified atrophy without flattening in the group of patients with hereditary spastic paraplegia. Fractional anisotropy of the corpus callosum and pyramidal tracts did correlate with disease severity. Hereditary spastic paraplegia is characterized by relative sparing of the cortical mantle and remarkable damage to the distal portions of the corticospinal tracts, extending into the spinal cord.

The Conformational Analysis Of 2-halocyclooctanones.

The establishment of the most stable structures of eight membered rings is a challenging task to the field of conformational analysis. In this work, a series of 2-halocyclooctanones were synthesized (including fluorine, chlorine, bromine and iodine derivatives) and submitted to conformational studies using a combination of theoretical calculation and infrared spectroscopy. For each compound, four conformations were identified as the most important ones. These conformations are derived from the chair-boat conformation of cyclooctanone. The pseudo-equatorial (with respect to the halogen) conformer is preferred in vacuum and in low polarity solvents for chlorine, bromine and iodine derivatives. For 2-fluorocyclooctanone, the preferred conformation in vacuum is pseudo-axial. In acetonitrile, the pseudo-axial conformer becomes the most stable for the chlorine derivative. According to NBO calculations, the conformational preference is not dictated by electron delocalization, but by classical electrostatic repulsions.

The C-terminal Region Of The Human P23 Chaperone Modulates Its Structure And Function.

The p23 protein is a chaperone widely involved in protein homeostasis, well known as an Hsp90 co-chaperone since it also controls the Hsp90 chaperone cycle. Human p23 includes a β-sheet domain, responsible for interacting with Hsp90; and a charged C-terminal region whose function is not clear, but seems to be natively unfolded. p23 can undergo caspase-dependent proteolytic cleavage to form p19 (p231-142), which is involved in apoptosis, while p23 has anti-apoptotic activity. To better elucidate the function of the human p23 C-terminal region, we studied comparatively the full-length human p23 and three C-terminal truncation mutants: p23₁₋₁₁₇; p23₁₋₁₃₁ and p23₁₋₁₄₂. Our data indicate that p23 and p19 have distinct characteristics, whereas the other two truncations behave similarly, with some differences to p23 and p19. We found that part of the C-terminal region can fold in an α-helix conformation and slightly contributes to p23 thermal-stability, suggesting that the C-terminal interacts with the β-sheet domain. As a whole, our results suggest that the C-terminal region of p23 is critical for its structure-function relationship. A mechanism where the human p23 C-terminal region behaves as an activation/inhibition module for different p23 activities is proposed.

Modeling The Effects Of Temperature And Ph On The Resistance Of Alicyclobacillus Acidoterrestris In Conventional Heat-treated Fruit Beverages Through A Meta-analysis Approach.

In this work, all publicly-accessible published findings on Alicyclobacillus acidoterrestris heat resistance in fruit beverages as affected by temperature and pH were compiled. Then, study characteristics (protocols, fruit and variety, °Brix, pH, temperature, heating medium, culture medium, inactivation method, strains, etc.) were extracted from the primary studies, and some of them incorporated to a meta-analysis mixed-effects linear model based on the basic Bigelow equation describing the heat resistance parameters of this bacterium. The model estimated mean D* values (time needed for one log reduction at a temperature of 95 °C and a pH of 3.5) of Alicyclobacillus in beverages of different fruits, two different concentration types, with and without bacteriocins, and with and without clarification. The zT (temperature change needed to cause one log reduction in D-values) estimated by the meta-analysis model were compared to those ('observed' zT values) reported in the primary studies, and in all cases they were within the confidence intervals of the model. The model was capable of predicting the heat resistance parameters of Alicyclobacillus in fruit beverages beyond the types available in the meta-analytical data. It is expected that the compilation of the thermal resistance of Alicyclobacillus in fruit beverages, carried out in this study, will be of utility to food quality managers in the determination or validation of the lethality of their current heat treatment processes.

Aplicação do escore MELD em pacientes submetidos a transplante de fígado: análise retrospectiva da sobrevida e dos fatores preditivos a curto e longo prazo

BACKGROUND: The model for end-stage liver disease (MELD) was developed to predict short-term mortality in patients with cirrhosis. There are few reports studying the correlation between MELD and long-term posttransplantation survival. AIM: To assess the value of pretransplant MELD in the prediction of posttransplant survival. METHODS: The adult patients (age >18 years) who underwent liver transplantation were examined in a retrospective longitudinal cohort of patients, through the prospective data base. We excluded acute liver failure, retransplantation and reduced or split-livers. The liver donors were evaluated according to: age, sex, weight, creatinine, bilirubin, sodium, aspartate aminotransferase, personal antecedents, brain death cause, steatosis, expanded criteria donor number and index donor risk. The recipients' data were: sex, age, weight, chronic hepatic disease, Child-Turcotte-Pugh points, pretransplant and initial MELD score, pretransplant creatinine clearance, sodium, cold and warm ischemia times, hospital length of stay, blood requirements, and alanine aminotransferase (ALT >1,000 UI/L = liver dysfunction). The Kaplan-Meier method with the log-rank test was used for the univariable analyses of posttransplant patient survival. For the multivariable analyses the Cox proportional hazard regression method with the stepwise procedure was used with stratifying sodium and MELD as variables. ROC curve was used to define area under the curve for MELD and Child-Turcotte-Pugh. RESULTS: A total of 232 patients with 10 years follow up were available. The MELD cutoff was 20 and Child-Turcotte-Pugh cutoff was 11.5. For MELD score > 20, the risk factors for death were: red cell requirements, liver dysfunction and donor's sodium. For the patients with hyponatremia the risk factors were: negative delta-MELD score, red cell requirements, liver dysfunction and donor's sodium. The regression univariated analyses came up with the following risk factors for death: score MELD > 25, blood requirements, recipient creatinine clearance pretransplant and age donor >50. After stepwise analyses, only red cell requirement was predictive. Patients with MELD score < 25 had a 68.86%, 50,44% and 41,50% chance for 1, 5 and 10-year survival and > 25 were 39.13%, 29.81% and 22.36% respectively. Patients without hyponatremia were 65.16%, 50.28% and 41,98% and with hyponatremia 44.44%, 34.28% and 28.57% respectively. Patients with IDR > 1.7 showed 53.7%, 27.71% and 13.85% and index donor risk <1.7 was 63.62%, 51.4% and 44.08%, respectively. Age donor > 50 years showed 38.4%, 26.21% and 13.1% and age donor <50 years showed 65.58%, 26.21% and 13.1%. Association with delta-MELD score did not show any significant difference. Expanded criteria donors were associated with primary non-function and severe liver dysfunction. Predictive factors for death were blood requirements, hyponatremia, liver dysfunction and donor's sodium. CONCLUSION: In conclusion MELD over 25, recipient's hyponatremia, blood requirements, donor's sodium were associated with poor survival.