A rapid method to evaluate potential vasodilatory activity of organic nitrates

The kinetics of interaction between organic nitrates (3,3-bis(nitroxymethyl)oxetane) and cysteine were evaluated by the rate of nitrite ion formation at various concentrations of reagents and pH. The activities of natural reducing agents, including cysteine, glutathione, and NADH, in generating the nitrite ion from organic nitrates (3,3-bis(nitroxymethyl)oxetane) were compared. Cysteine was shown to be the most potent reducing agent. Studying the effectiveness of nitrates (trinitroglycerol, 3,3-bis(nitroxymethyl)oxetane, and nicorandil) at a concentration of 3 mM showed that the rate of nitrite ion accumulation in the reaction with 10 mM cysteine is 1.66, 0.37, and 0.02 microM/min, respectively.     

A rapid method to evaluate potential vasodilatory activity of organic nitrates

The kinetics of interaction between organic nitrates (3,3-bis(nitroxymethyl)oxetane) and cysteine were evaluated by the rate of nitrite ion formation at various concentrations of reagents and pH. The activities of natural reducing agents, including cysteine, glutathione, and NADH, in generating the nitrite ion from organic nitrates (3,3-bis(nitroxymethyl)oxetane) were compared. Cysteine was shown to be the most potent reducing agent. Studying the effectiveness of nitrates (trinitroglycerol, 3,3-bis(nitroxymethyl)oxetane, and nicorandil) at a concentration of 3 mM showed that the rate of nitrite ion accumulation in the reaction with 10 mM cysteine is 1.66, 0.37, and 0.02 μM/min, respectively. The reaction of organic nitrate with cysteine (Cys) is used as a test system for analyzing the effectiveness of nitrates in nitrite ion formation, which correlates with vasodilatory activity of these compounds (dilation of blood vessels).     

A new method to evaluate the thermal stability of lyophilized enzymes

Summary The thermal inactivation of lyophilized chymotrypsin was studied at controlled water activities. At 60 °C the enzyme showed good stability except at aw 0.97, whereas at 75 °C considerable inactivation occured at most water activities. Increasing the amount of buffer on the preparation decreased the stability significantly. The optimal temperature of enzymatic activity was increased 14 °C, when the water activity was decreased from 1 to 0.5.     

Effect of nutrient periodicity on microbial community dynamics

When microbes are subjected to temporal changes in nutrient availability, growth rate and substrate affinity can contribute to competitive fitness and thereby affect microbial community structure. This hypothesis was tested using planktonic bacterial communities exposed to nutrient additions at 1-, 3-, 7-, or 14-day intervals. Growth rates after nutrient addition were inversely proportional to the pulse interval and declined from 0.5 h(-1) to 0.15 h(-1) as the pulse interval increased from 1 to 14 days. The dynamics of community structure were monitored by 16S rRNA gene PCR-denaturing gradient gel electrophoresis. At pulse intervals of more than 1 day, the community composition continued to change over 130 days. Although replicate systems exposed to the same pulse interval were physiologically similar, their community compositions could exhibit as much dissimilarity (Dice similarity coefficients of <0.5) as did systems operated at different intervals. Bacteria were cultivated from the systems to determine if the physiological characteristics of individual members were consistent with the measured performance of the systems. The isolates fell into three bacterial divisions, Bacteroidetes, Proteobacteria, and Actinobacteria. In agreement with community results, bacteria isolated from systems pulsed every day with nutrients had higher growth rates and ectoaminopeptidase specific activities than isolates from systems pulsed every 14 days. However, the latter isolates did not survive starvation longer than those provided with nutrients every day. The present study demonstrates the dynamic nature of microbial communities exposed to even simple and regular environmental discontinuities when a substantial pool of species that can catabolize the limiting substrate is present.     

A quantitative method to evaluate neutralizer toxicity against Acanthamoeba castellanii.

A standard methodology for quantitatively evaluating neutralizer toxicity against Acanthamoeba castellanii does not exist. The objective of this study was to provide a quantitative method for evaluating neutralizer toxicity against A. castellanii. Two methods were evaluated. A quantitative microtiter method for enumerating A. castellanii was evaluated by a 50% lethal dose endpoint method. The microtiter method was compared with the hemacytometer count method. A method for determining the toxicity of neutralizers for antimicrobial agents to A. castellanii was also evaluated. The toxicity to A. castellanii of Dey-Engley neutralizing broth was compared with Page's saline. The microtiter viable cell counts were lower than predicted by the hemacytometer counts. However, the microtiter method gives more reliable counts of viable cells. Dey-Engley neutralizing medium was not toxic to A. castellanii. The method presented gives consistent, reliable results and is simple compared with previous methods.     

A method to evaluate survival of genetically engineered bacteria in soil extracts

A technique of potential use to the biotechnology industry was developed for studying the survival of bacteria in aqueous extracts of soil. The aqueous extracts of soil were placed into test tubes, amended as desired, inoculated with bacteria containing recombinant DNA, and incubated. Most bacteria introduced into filter-sterilized soil extracts were capable of multiplying and maintained populations of 10 E6 to 10 E8 cfu/ml over 13 days. However, bacteria introduced into nonsterile soil extracts at 10 E5 cfu/ml were found to decrease by 2–3 logs over a 13-day period. The soil extract method revealed that recombinant DNA plasmids had no significant effect on survival of thePseudomonas spp. andEscherichia coli strains examined. Extracts from soil provide a convenient and homogeneous milieu for estimating relative competitiveness and documenting survival characteristics of genetically engineered microorganisms. The use of aqueous extracts of soil offer convenience, a means of obtaining homogeneous cell suspensions, and ease of experimental replication over the inoculation of bacteria uniformly into soil.     

A method to evaluate and compare two different intraoral radiographs of the same patient

Objective of this study was to determine the accuracy of the method of the clinical intraoral densitometry, to compare differences in the calculation with or without subtraction of the background adjacent soft-tissues from the stepwedge (SW) and to verify which regression model best fitted the experimental data in order to express the measured values in equivalents of SW thickness. Two intraoral radiographs, one after another, were made for each of 6 patients. A copper SW (6 steps, thickness 0.05-0.3 mm) was attached to each radiograph, trying to avoid the superimposition of the bony structures. Films were processed and digitized. Grey levels were measured on each step of the SW, on the background of the SW and on the same 3 randomly chosen regions of interest (ROIs) on each digitized image. The measurement with and without the subtraction of optical densities of the background around the SW from the optical densities of the SW was performed. For the calculation of the SW thickness equivalents, the regression analysis was performed by using different regression models. The best fitting regression model was the 3rd degree polynomial. The results were more precise when using the subtraction of the background overlapping the SW.     

A method to evaluate the antioxidant system for radicals in erythrocyte membranes

The erythrocyte defense system against cellular oxidants is complex and efficient. Free radicals generated in cell membranes, however, are relatively sequestered from the cell's antioxidant mechanisms. When an oxidant challenge exceeds the capacity of the erythrocyte's antioxidant system, membrane damage may occur, causing red cell destruction and hemolytic anemia. In this study, we present a method for monitoring radical reduction in erythrocyte membranes, using fatty acid spin labels with nitroxide radicals on the hydrocarbon chains. About 50 microL of packed (about 5-6 x 10(8)), carbon monoxide (CO)-gassed red blood cells are used. The electron paramagnetic resonance signals of the 5-doxylstearic acid spin labels in the intact cells are obtained as a function of time, at 37 degrees C over a period of 2 h. The pseudo first-order rate constant for reduction of the spin label in normal adult intact cells under our experimental conditions is 4.3 +/- 1.8 x 10(-3)/min. The reproducibility and variability of the measurements are discussed. Since the measurements we describe reflect the extent of radical reductions occurring in cell membranes, we suggest that this method can be used to measure the ability to defend oxidants in membranes of erythrocytes with defective antioxidant systems. This method is particularly useful for measuring the modification of the antioxidant system toward radicals in membranes by drugs, chemicals, or environmental toxins.     

A simple method to evaluate biofilm formation in exopolysaccharide-producing strains of Sinorhizobium meliloti

ABSTRACT

Sinorhizobium meliloti

is a bacterium of great agroeconomic importance because of its ability to fix atmospheric nitrogen in symbiotic association with alfalfa (Medicago sativa) roots, and it is often used in model studies. We investigated the effects of exopolysaccharides (EPSs) in cell-to-cell and cell-to-surface interactions in S. meliloti. The microtiter plate assay, a quantitative spectrophotometric method, is used to study bio?lm formation by bacterial adherence to an abiotic surface. It consists in staining biofilms grown in microtiter plates. Here, we describe two microbiology laboratory classes designed for undergraduate students of Experimental Biological Chemistry, in which they learn about biofilm forming capacity by observing the behavior of both wild-type and mutant strains of S. meliloti.     

A quantitative colorimetric method to evaluate the functional state of human polymorphonuclear leukocytes

The colorimetric assay previously described by Mosmann for the measurement of cell viability and proliferation has been modified for the assessment of the functional state of human polymorphnuclear cells (PMNs). The ability of PMNs to reduce the tetrazolium salt MTT to formazan reflects directly the degree of stimulation induced by various agents. The underlying mechanism of MTT-reduction to formazan seems to be similar to that of nitroblue tetrazolium (NBT)-reduction. In contrast to the NBT-reduction assay, the formazan produced from MTT can easily be measured by an ELISA reader. Parallel experiments revealed a qualitative correlation between the concentration of formazan produced from MTT and the concentration of cytochrome C reduced by PMNs. Although oxidative burst may not be the actual lytic mechanism in cellular cytotoxicity of PMN, we also observed an association between MTT-reduction capacity and the cytotoxic activity of PMNs from normal donors in antibody dependent cellular cytotoxicity. Our results indicate that the MTT-reduction assay can be employed to estimate the functional state of polymorphnuclear granulocytes.     

A simple method to evaluate the concentration of pentachlorophenol degraders in contaminated soils

A new most probable number (MPN) method for the determination of pentachlorophenol (PCP) degraders in soil using the change in pH due to PCP degradation is compared with a well documented MPN method using radiolabeled PCP. The results of all MPN counts were similar within a 95% confidence limit. The results obtained in MPN per gram of dry soil using pH measurements were 1.8 (+3.1, -1.03) x10 (4) compared to 0.64 (+1.34, -0.42) x 10(4) when using production of [(14)C]CO(2).     

A simple and rapid one-time method to evaluate the non-acidic gas content from bioprocesses

This paper presents a rapid less than 2 min and low-cost method involving the use of alkali solution to capture the acidic gasses from a biogas, thereby providing an estimate of the percentage of non-acidic gasses. Such a method was mentioned in the literature but never fully described or optimized. After sampling an aliquot of gas from bioprocess, gas was injected in a sealed flask with a 3 M NaOH solution, and after equilibrium was obtained, the non-acidic gas volume was measured. The method was first calibrated with certified gasses with an accuracy observed between 98 and 105 %. Regarding the validation step, certified standard gas mixtures and nine biogas-laboratory batch reactors were used, the overall accuracy reported was 103 + 3 %. This rapid and low-cost method may either be used in laboratory conditions as a quick and low cost alternative to standard analysis equipment or in addition as a routine field control method used on full-scale plants.     

An image method to evaluate bagasse fiber dimensions

In the last 25 years the amount of textile nonwovens used for industrial and commercial applications increased more than 10 times. Bagasse fiber, a by-product from sugar cane industry, provides a natural resource for nonwoven industries. Even though underrated as a potential fiber, bagasse comes more and more into attention because of the increasing concern for disposal of agricultural residuals and the need for enhancing the sugar cane industry’s profitability. However, there is a lack of an instrumental method to evaluate bagasse fiber length and fineness. This paper presents a study on measuring the bagasse fineness using image analysis method. Cross-sections images of bagasse fibers were visualized using Scanning Electronic Microscopy (SEM). The computing software Scion Image was used to measure bagasse fiber length and cross-sectional area. Relationship between fiber fineness and cross-sectional area was analyzed using the statistical method of regression.     

A method to evaluate capacity and efficiency of water soluble antioxidants as peroxyl radical scavengers

In this paper, we report on a method to evaluate the activity of water soluble and H-atom donor antioxidants as peroxyl radical scavengers in a micelle system reproducing the conditions occurring in the upper small intestine in humans, during digestion and absorption of lipids. This method, which overcomes some of the problems of the total radical trapping antioxidant parameter (TRAP) assays, measures the peroxyl radical trapping capacity (n) and the peroxyl radical trapping efficiency IC50(-1) of antioxidants, that is the number "n" of peroxyl radicals trapped by one molecule of the studied antioxidant and the reciprocal of the antioxidant concentration that halves the steady-state concentration of peroxyl radicals, respectively. These two fundamental parameters characterizing the radical chain breaking of many water soluble antioxidants, among which dietary polyphenols, can be obtained with relatively good precision from a single experiment, on the basis of a rigorous treatment of the kinetic data.