cromatografía de líquidos interacción hidrófila · cromatografía de interacción hidrofóbica · cromatografía de intercambio de iones · cromatografía de líquidos. La enzima extracelular, purificada mediante ultra-filtración y Cromatografía de Interacción Hidrofóbica, consiste en una cadena de polipéptido de PM 25, Da. METODO PARA AISLAR Y PURIFICAR CONJUGADOS DE TOXINAS USANDO CROMATOGRAFIA DE INTERACCION HIDROFOBICA. LAS MEZCLAS.

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Cromatografía de afinidade

To date, several enzymes from Hf. General and Applied aspects of Halophilic microorganisms. Exposure to lower or higher salt concentration reduced enzyme activity.

The reaction was ended removing the insoluble material by filtration through a Whatman filter paper and the absorbance at nm measured against the corresponding blank. Modification of protease activity in this case, evidenced the influence of the anion species in enzyme activity.

The new concentrated material inteeraccion designated as CS After this period, the proteolytic activity was found using 0.

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All these tests provided some useful information about the properties of the enzyme that led us to conclude that the extracellular enzyme activity of Hf. Stability of extracellular protease activity in different chemical conditions. Acta, Ivo Safarik as substrate, the optimum salt NaCl concentration cromatogfafia enzyme activity was determined varying the amount of NaCl in the assay mixture 0 to 4. The purified extracellular protease activity of Hf.


We all enjoyed his teachings, friendship, and continuous support through all these years. Extremophiles, 5, The active fractions that separately emerged from the column Fig. Comparative Biochemistry hidrofobics Physiology. First, for the removal of the exopolysaccharide component a selective ultrafiltration procedure was carried out. Although ingeraccion halophilic enzymes may be reactivated from the salt-free solutions [21, ], and this facilitates their purification using standard methods, i.

Biochimie60 Table 3 describes the effect of different ions on the activity of the purified extracellular protease of Hf. Specific ion requirement for CS proteolytic activity.

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All chemicals and reagents were analytical grade Sigma, Chem. Experientia, 29 Stability of extracellular cromatoggrafia activity in different chemical conditions In order to determine the appropriate experimental conditions for purification of Hf.

The above experiments provided important information about the conditions required for the proper handling of an active extracellular enzyme produced by Hf.

We all try to be faithful following his example of devotion for science. Hence, reported purification methods in cromatogarfia cases have included: Biopolymer production by Haloferax mediterranei.

However, with casein Hf. Cation effect on CS proteolytic activity. After dialysis, the remaining protease activity was measured as above using a 1: KCl and other salts, including ammonium salts, were much less efficient The specific requirement of NaCl for enzyme activity and stability Fig.


Nevertheless, there are some jnteraccion proteases, like the dee from Hb. The combined effect of such parameters is presented in Figs. Study of proteases of marine bacteria: The NaCl concentration was adjusted to 2. The capability of Hf. Then, taking into account the nature extracellularand composition high salt concentration culture medium of the CSEP preparation, Hydrophobic Interaction Chromatography, HIC, was considered the method of choice for the enzyme purification as shown in the following section.

Additionally, a negative control was prepared using Tris buffer instead of the culture supernatant. Saturated solutions of sucrose, and betaine, and 5.

Being aware of the advantages offered by Hydrophobic Interaction Chromatography, a technique that allows the use of a high salt concentration to favor a selective adsorption of the protein on the basis of its hydrophobicity [35], we decided to explore its application in the isolation of the extracellular enzyme responsible of the proteolytic activity shown by Hf. The effect of salt concentration on both substrate and enzyme may favor their association and therefore help the enzyme function better.

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