טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentLipkovsky Esther
SubjectPeroxidase Catalayzed Oxidation and Polymerization of
Catecholic Derivatives
DepartmentDepartment of Biotechnology
Supervisor Professor Emeritus Carlos Dosoretz


Abstract

Phenols and catechols are oxidized by oxidases and peroxidases to form phenoxy radicals which couple to form oligomeric and polymeric products that can be used for “green” polyphenol and polycatechol biosynthesis.

The present research focused on the characterization of the oxidation and polymerization reaction of catechols by horseradish peroxidase as function of the substituting groups affecting the electron density of the ring which indirectly affects the oxidation potential or easy of oxidation: catechol (no substitution group), 4-methylcatechol (electron-donating group) and 4-chlorocatechol (electron-withdrawing group). The research hypothesis is that catechols as well as their polymerization products are highly hydrophilic, and therefore the expected degree of polymerization in water is high (compared to corresponding phenols). Since polycatechols exhibit high water solubility, this should eliminate the need for water miscible solvents to obtain the desired polymeric products, and reduce the amount of enzyme needed due to enzyme denaturation. 

The work included characterization of the kinetics of oxidation and calculation of the kinetic constants of the model substrates; characterization of the polymerization reaction as function of substrates and peroxide concentrations; mass balance for carbon and nitrogen upon fractionation of reaction mixture at acidic pH; chemical characterization of reaction products by molecular weight determination using gel filtration chromatography and mass spectroscopy (MS-MS) and FTIR analysis. The findings show that hydrophobic substitution groups of opposed electronegativity have an opposed effect not only on the reaction kinetics but also on the degree of polymerization, the structure of the polymerization products and on their interaction with the enzyme.