Thermostability of free and immobilized Aspergillus niger chlorogenate hydrolase

BUTIUK, A.P.; ADACHI, O.; MIGNONE, C.; HOURS, R.A.

Mar del Plata

Encuentro; VIII Encuentro Latinoamericano y del Caribe de Biotecnología; 2013

RED BIO Argentina 20

Chlorogenate hydrolase (EC 3.1.1.42, CHase) was formed by Aspergillus niger AKU 3302 grown in Czapek medium containing 0.125% (w/v) of concentrated yerba mate extract as inducer. The cell-free extract was prepared by grinding mycelia in a mortar in the presence of Na-phosphate buffer and centrifuging the resultant suspension. Free CHase solution and immobilized CHase (mycelia CHase) were put in tubes dipped in a water bath and kept at different combinations of time (20-60 min) and temperature (45-60°C) according to a Doehlert experimental design. The residual CHase activity was measured and expressed as % relative to a control (without heat treatment). Thermostability of free and immobilized CHase activity was compared. Immobilized CHase remained stable up to 55°C and 60 min retaining about 90% CHase activity whereas free CHase only retained about 40% of its initial activity under the same conditions. At temperatures higher than 55°C, the activity of the free enzyme declined rapidly. Immobilized CHase was more stable than free CHase and at 65°C and 60 min still retained 50% of residual activity. Once immobilized CHase was heated at 55°C for 60 min, vegetative mycelial growth was negative thus allowing to be packed in a column as stable immobilized catalyst.