Aqueous two-phase systems (ATPS) of PEG/phosphate were used to recover collagenase from Penicillium aurantiogriseum from fermented broth. Experiments were carried out according to a 24-full factorial design using the PEG molar mass (MPEG), PEG concentration (CPEG), phosphate concentration (CPHOS) and pH as the independent variables, and the purification factor (PF), partition coefficient (K) and activity yield (Y) as the responses. All the responses increased in the top phase with increasing CPEG and decreasing MPEG, but the maximum value of PF (5.23) was obtained at the highest CPHOS (15% w/w) and the lowest pH (6.0), that of Y (167.01) at opposite levels of CPHOS (10% w/w) and pH (8.0), and that of K at the highest levels of both. The electrophoretic profile revealed that some protein contaminants were removed by the ATPS, and the extracted collagenase exhibited optimum activity at pH 8.0 and 45 ºC. The proposed ATPS appears to be a promising alternative to conventional first-step operations for direct recovery of collagenase from fermented broths, yielding a concentrate enzyme solution able to effectively hydrolyze collagen.
Two-phase partitioning and partial characterization of a collagenase from Penicillium aurantiogriseum URM4622: Application to collagen hydrolysis
CONVERTI, ATTILIO;
2013-01-01
Abstract
Aqueous two-phase systems (ATPS) of PEG/phosphate were used to recover collagenase from Penicillium aurantiogriseum from fermented broth. Experiments were carried out according to a 24-full factorial design using the PEG molar mass (MPEG), PEG concentration (CPEG), phosphate concentration (CPHOS) and pH as the independent variables, and the purification factor (PF), partition coefficient (K) and activity yield (Y) as the responses. All the responses increased in the top phase with increasing CPEG and decreasing MPEG, but the maximum value of PF (5.23) was obtained at the highest CPHOS (15% w/w) and the lowest pH (6.0), that of Y (167.01) at opposite levels of CPHOS (10% w/w) and pH (8.0), and that of K at the highest levels of both. The electrophoretic profile revealed that some protein contaminants were removed by the ATPS, and the extracted collagenase exhibited optimum activity at pH 8.0 and 45 ºC. The proposed ATPS appears to be a promising alternative to conventional first-step operations for direct recovery of collagenase from fermented broths, yielding a concentrate enzyme solution able to effectively hydrolyze collagen.
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