Studies on the extraction and back-extraction of a recombinant cutinase in a reversed micellar extraction process

This work deals with the extraction and back-extraction of a recombinant cutinase using AOT reversed micelles in isooctane. The effect of pH, ionic strength, AOT concentration and temperature on the extraction and back-extraction of the cutinase was investigated. High extraction (97%) of the cutinase was achieved at pH 7.0 with a 50 mM Tris-HCl buffer solution containing 100 mM KCl, but a low activity was detected in the reversed micellar phase. At pH 9.0, cutinase was extracted (75%) to the reversed micelles with higher activity. Cutinase was recovered (50%) from a reversed micellar phase (100 mM AOT/isooctane) into a 50 mM Tris-HCl buffered solution at pH 9.0 with 100 mM KCl, and 20°C. Protein and cutinase activity global yields of 38 and 45%, respectively, were obtained for the global process, extraction and back-extraction steps, using low ionic strength, pH 9.0, 100 mM AOT and 20°C.Maria das Graças Carneiro da Cunha acknowledges a Ph.D. fellowship from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and Centro de Pesquisas Aggeu Magalhães, Brasil. This work was partly financed by the BRIDGE Programme (Contract BIOT-CT91-0274(DTEE)).