Calculation of some equilibrium parameters and thermodynamic characteristics of sorption in the process of papain immobilization on fibrous carriers
Abstract
One of the current and promising areas in modern pharmaceutical practice is the development and implementation of immobilized enzyme preparations. Pharmaceutical forms containing the proteolytic enzyme papain (EC 3.4.22.2) have got anti-inflammatory, antioxidant, immunostimulating and regenerating effects and they are effective for treatment of various skin lesions and purulent complications. Immobilization of enzymes is aimed at obtaining prolonged-action drugs due to their increased stability and increased half-life of the enzyme, facilitating the diffusion of the substance in the human body, preventing autolysis, and increasing the resistance of the protein macromolecule to denaturing factors. This paper discusses the adsorption immobilization of papain using fibrous ion exchangers based on a polypropylene-styrene-divinylbenzene matrix as carriers: a weakly acidic cation exchanger K-4 and a highly basic anion exchanger A-1. Adsorption immobilization was carried out at temperatures of 293±2K, 313±2K using the variable concentration method in the range of 0.2-5.0∙10-2 mmol/dm3, pH 6.5. It was revealed that protein sorption on fibers at the temperature of 313±2K is described by the Langmuir model. Analysis of the obtained papain sorption isotherms the 293±2K showed that with an increase of the papain concentration in the external solution, the adsorption process is mainly due to protein-protein interactions. The formation of polymolecular layers of the enzyme is described by the Brunauer-Emmett-Teller equation. Physical sorption makes the most significant contribution to the total sorption capacity of fibrous carriers. An assumption has been made that in this case, supramolecular structures in the form of protein associates are formed. The catalytic activity of heterogeneous biocatalysts at pH 6.5 is 47-87% of the enzymatic activity of soluble papain.
In order to predict the conditions of enzyme binding to the carriers, the equilibrium parameters of protein sorption by the carriers under consideration (constants characterizing the interactions papain-fibrous carrier, papain-papain) and some thermodynamic characteristics of the immobilization process under study (sorption process energy, enthalpy, entropy) were calculated.
The maximum values of the sorption parameter and the constants of monomolecular sorption KS, calculated using the BET equation for K-4 and A-1 at 293 K, are characterized by higher values compared to the values obtained on hand of the Langmuir equation. This fact indicates the predomination of protein-protein interactions in the system during immobilization with an increase in the concentration of the solution. The calculated filling constant of the KL polylayers for K-4 takes on greater values (KL=3.97 dm3/mmol) compared to (KL=2.89 dm3/mmol), which indicates a greater affinity for the studied enzyme of this carrier. The results of the study can be considered as one of the stages in the technology of developing drugs production based on immobilized forms of the proteolytic enzyme papain.
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