RESUMO
The use of polymers as supports for enzyme immobilization is a strategy that enables to remove the enzymes from a chemical reaction and improve their efficiency in catalytic processes. In this work, cellulose triacetate (CTA) was used for physical adsorption of phospholipase Lecitase ultra (LU). CTA is more hydrophobic than cellulose, shows good performance in the lipases immobilization being a good candidate for immobilization of phospholipases. We investigated the immobilization of LU in CTA, the stability of the immobilized enzyme (CTA-LU) and the performance of CTA-LU using soybean oil as a substrate. LU was efficiently immobilized in CTA reaching 97.1% in 60 min of contact with an enzymatic activity of 975.8 U·g-1. The CTA-LU system presents good thermal stability, being superior of the free enzyme and increase of the catalytic activity in the whole range of pH values. The difference observed for immobilized enzyme compared to free one occurs because of the interaction between the enzyme and the polymer, which stabilizes the enzyme. The CTA-LU system was used in the transesterification of soybean oil with methanol, with the production of fatty acid methyl esters. The results showed that CTA-LU is a promising system for enzymatic reactions.
Assuntos
Celulose/análogos & derivados , Enzimas Imobilizadas/química , Fosfolipases/química , Adsorção , Catálise , Celulose/química , Estabilidade Enzimática , Esterificação , Interações Hidrofóbicas e Hidrofílicas , Metanol/metabolismo , Óleo de Soja/químicaRESUMO
Lecitase Ultra has been immobilized on cyanogen bromide agarose (via covalent attachment) and on octyl agarose (via physical adsorption on the hydrophobic support by interfacial activation). Both immobilized preparations have been incubated in dextran sulfate (DS) or polyethylenimine (PEI) solutions to coat the enzyme surface. Then, the activity versus different substrates and under different experimental conditions was evaluated. The PEI coating generally produced a significant increase in enzyme activity, in some cases even by more than a 30-fold factor (using the octyl-Lecitase at pH 5 in the hydrolysis of methyl phenyl acetate). In opposition, the DS coating usually produced some negative effects on the enzyme activity. The rate of irreversible inhibition of the covalent preparation using diethyl p-nitrophenylphosphate did not increase after PEI coating suggesting that the increase in Lecitase activity is not a consequence of the stabilization of the open form of Lecitase. Moreover, the coating greatly increased the stability of the immobilized Lecitase, for example using DS and the covalent preparation, the half-life was increased by a 30-fold factor in 30% acetonitrile. The stabilizing effect was not found in all cases, in certain cases even a certain destabilization is found (e.g., octyl-Lecitase-DS at pH 7). Thus, the effects of the ionic polymer coating strongly depend on the substrate, experimental conditions and immobilization technique employed.