Purification of gliadin subclasses has been difficult since they share many biochemical and physicochemical properties. In this report, the optimization of a preparative electrophoretic method to fractionate gliadins is described. Separation was performed in preparative 7% polyacrylamide gels at pH 3.1. The separation performance was tested using analytical electrophoresis at pH 3.1 and capillary electrophoresis. Preparative gels of different lengths were employed. Using 5-cm preparative gels, several fractions of alpha-, beta-, and gamma-gliadins and fast-mobility and slow-mobility omega-gliadins were collected in 40 h of separation. Resolution was maintained at a protein load of up to 30 mg in each run. The highest efficiency of recovery was achieved using aluminum lactate as the collecting buffer. Fractionation with 10 cm in length gels improved resolution but increased operation times. Gels of 2 cm in length did not separate alpha/beta- and gamma-gliadins efficiently but were useful to separate the two main fractions of omega-gliadins in shorter times. In conclusion, preparative electrophoresis at low pH allowed the separation of alpha-, beta-, gamma-, and omega-gliadin fractions from crude material under nondenaturing conditions.
A novel nanostructured material was successfully developed by combining a chitin matrix with graphene oxide nanosheets (Chi:nGO) and then used for the continuous flow adsorption of ciprofloxacin. The spectroscopic characterization indicated that none covalent interaction between both components would be occurring and the introduction of nGO did not interfere in chitin nanostructure rearrangement during gelling and later drying. SEM images and Mercury Intrusion Porosimetry results showed a wide pore size distribution ranging from nano to micrometers. The continuous flow adsorption was observed to be dependent on the pH which affects the electrostatic interaction. The flow rate, Na concentration and water hardness were evaluated to describe the adsorption process. The resistance to alkali allowed to regenerate and reuse the column for subsequent adsorption cycles. Finally, ciprofloxacin spiked real water samples were assessed and the results confirmed that the medium pH was the main parameter that defines the adsorption behavior.
A reproducible, rapid procedure for the simultaneous quantitative separation of S-adenosylmethionine and S-adenosylhomocysteine by capillary zone electrophoresis has been developed. Separations were performed by using an uncoated capillary of 60 cm effective length and 50 microm ID, 40 mM sodium phosphate buffer, pH 2.50, as background electrolyte solution, and 30 kV. On-line detection was carried out at 254 nm. Under the conditions selected we resolved a standard solution containing S-adenosylmethionine and S-adenosylhomocysteine in a run time shorter than 8 min. A mass detection limit in the range of 10 fmol was achieved. Adenosyl-L-methionine, S[methyl-3H] has also been assayed under the same experimental conditions. Other related compounds did not show interference, including those derived from the hydrolysis of S-adenosylmethionine. The present method allows simultaneous determination of these compounds, which play an important role in many microbiological and enzymatic research studies.
A reproducible, rapid procedure for the determination of glutathione in human blood by micellar electrokinetic chromatography has been developed. Whole blood samples were deproteinized with 5% w/v sulfosalicylic acid (final concentration). After centrifugation, the supernatant was directly injected for analysis, without further derivatization. Separations were performed by using an uncoated capillary of 30 cm effective length and 50 microns internal diameter (ID), 50 mM Tris-HCl, 30 mM sodium dodecyl sulfate (SDS), pH 7.00, as running buffer, and 10-20 kV. On-line detection was carried out at 214 nm and a detection limit in the range of femtomoles was achieved. Under the same experimental conditions, we resolved a mixed standard solution containing glutathione in its oxidize and reduced forms, lipoamide and alpha-lipoic acid. The corresponding migration times were reproducible. The present method allows rapid determination of these compounds, which play a critical role in oxidative stress, in cellular defense against injurious agents and whose levels are related to the toxicology and metabolism of several toxins and drugs, such as antineoplastic agents.
: Bioremediation is capable of reducing the hydrocarbon concentration of contaminated soil by 75È95% depending on the soil type, the kind of hydrocarbons and the history of the contamination. The impact of di †erent number of petrochemical sludge applications to soil on the degree of PAH elimination was assessed. A simple and reliable extraction and gas chromatographic method was used to facilitate more rapid determination of hydrocarbon contamination in soils and sludge wastes. Its application in a model laboratory bioremediation experiment and a pilot Ðeld study were used to illustrate its practical beneÐts. Post-remediation persistence of sludge constituents was evaluated after a single dose sludge application in the laboratory and after seven sludge applications in the Ðeld. A relative increase in the concentration of some PAHs was detected at the end of the experiments, but their individual concentrations were reduced to suggested values for industrial soils. The remaining concentration of total hydrocarbons in soil was found to be similar in both experiments, pointing to soil organic matter adsorption capacity as the factor determining hydrocarbon elimination limits in soil bioremediation.
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