Enzymatic degradation of natural polymers at physiological conditions

Abstract

U današnje vrijeme, prirodni polimeri se sve više istražuju zbog biorazgradivosti koja se temelji na hidrolitičkom ili enzimskom djelovanju na kemijskim vezama u polimerima. Materijali koji imaju svojstvo biokompatibilnosti koriste se za medicinsku primjenu kao nosači lijekova, u inženjerstvu tkiva i kože, pri izradi srčanih zalistaka, proteza krvnih žila itd. Do njihove razgradnje dolazi u kontaktu s tjelesnim fluidima i tkivima, a produkti ne smiju biti toksični. Najčešće korišteni biomaterijali su celuloza, kolagen, poli(ε-kaprolakton) i hidrogelovi. U ovom radu praćena je enzimska razgradnja poroznih struktura kitozana uz katalizator lizozim tijekom četiri tjedna inkubacije pri temperaturi od 37 °C. Enzimska razgradnja pratila se u fosfatom puferiranoj otopini soli (PBS, pH = 7,38) i u vodenom mediju. Porozne strukture karakterizirane su prije i nakon razgradnje instrumentalnim metodama rendgenske difrakcijske analize i infracrvene spektroskopije s Fourierovim transformacijama. Morfologija uzoraka istražena je pomoću pretražnog elektronskog mikroskopa. Dobiveni rezultati ukazuju na enzimsku biorazgradivost kitozana visokog stupnja deacetilacije (95 – 98 %) pri fiziološkim uvjetima, dok je aktivnost enzima u vodenom mediju izostala. Identifikacijska analiza potvrdila je zaostatak adsorbiranog lizozima koji utječe na konačni pad mase uzoraka tijekom razgradnje. Mikroskopska analiza ukazala je na promjenu mikrostrukture kitozanskih spužvi tijekom enzimske razgradnje, ali i na potencijalnu nehomogenost pripravljenih uzoraka.Nowadays, natural polymers are being more studied due to their biodegradability, which is based on hydrolytic or enzymatic actions on chemical bonds in polymers. Biocompatible materials are used in medical applications, such as drug delivery, tissue and skin engineering, manufacturing of heart valves, vascular prostheses etc. They degrade in contact with body fluids and tissue, and degradation products must not be toxic. The mostly used biomaterials are cellulose, collagen, poly(ε-caprolactone) and hydrogels. In this paper, enzymatic degradation of porous chitosan structures was monitored with lysozyme as a catalyst during four weeks of incubation at 37 °C. The enzymatic degradation was monitored in phosphate buffered saline (PBS, pH = 7.38), and water media. Scaffolds were analyzed before and after enzymatic degradation using X-ray diffraction analysis and infrared spectroscopy with Fourier transformations, while morphology of scaffolds was investigated by scanning electron microscopy. The results indicate enzymatic biodegradability of highly deacetylated chitosan (95 – 98%) at physiological conditions, while lysozyme showed the absence of its activity in aqueous medium. The scaffolds’ identification has confirmed the residues of absorbed lysozyme, which affects the final weight loss of the sample. Microscopic analysis has indicated changes in the microstructure of chitosan sponges during the enzymatic degradation, but also potential lack of homogeneity of prepared samples