Immobilisation of soybean peroxidase on different supports

MAGRI, MARÍA L; MIRANDA, MARÍA V; OSVALDO CASCONE

Río de Janeiro

Congreso; Enpromer 2005; 2005

  Enzymes have a great appeal in chemical processes as “green chemistry” reagents that will allow future sustainable developments. In this sense, enzyme technology has become a multidisciplinary field with applications in diverse processes. Peroxidases in particular have been extensively studied and show many attractive properties for biocatalysis such as wide specificity, high stability in solution and easy accessibility from plant materials. Soybean seed coat peroxidase (SBP) has arisen great interest due to a different activity and stability profile as compared to its widespread counterpart, horseradish peroxidase. Besides, SBP is obtained from soybean seed hulls, a low-cost agricultural by-product. In this work we study the immobilization of SBP on its natural support (soybean seed coats) and on polyaniline, a conducting polymer. Immobilization prevents inactivation in solution and extends the half-life of the enzyme. Experiments were designed to evaluate several variables, such as immobilization method, concentration of immobilization reagent, hydrogen peroxide to substrate molar ratio, optimum activity pH and immobilized enzyme stability. Since recent research has shown that SBP can be used for phenol removal in wastewater, the immobilized enzyme on both supports was assayed against a phenol solution. For seed coats, periodate and glutaraldehyde chemistries were assayed. Periodate failed to immobilize any amount of SBP, whereas glutaraldehyde was effective in immobilizing the enzyme. The optimum concentration of glutaraldehyde was determined to be 1%. Immobilization did not change the free enzyme optimum pH, which was 6.0. Treated seed coat stability was evaluated over a four-week period and the immobilized enzyme maintained its activity, while untreated seed coats lost their activity in 24 h under the same conditions. A reusability assay showed that seed coats could be reused once. In the case of polyaniline, SBP was optimally bound to a 2.5% glutaraldehyde-activated polyaniline at pH 6.0. A 6-h reaction was needed for completion and immobilization efficiency was 82%. The immobilized enzyme could be reused for 2 months (10 cycles assayed) without any loss of activity. Both immobilized species were used in phenol removal assays and performed well. A comparison of characteristics and functionality is also presented.