Amylase for Apple Juice Processing: Effects of pH, Heat, and Ca2+ Ions

Liliana N. Ceci and Jorge E. Lozano*

PLAPIQUI (UNS-CONICET) Camino La Carrindanga Km.7, C.C. 717  8000 Bahía Blanca – Argentina

Article history:

Received July 4, 2001
Accepted November 8, 2001

Key words:

α-amylase, apple juice, starch hydrolysis


The aim of this paper was to evaluate the effects of pH, heat, and Ca2+ ions on the α-amylase activities in a commercial amylolytic enzyme (Tyazyme L300), used for apple juice processing. Kinetics of thermal inactivation was studied in acetate and citrate/phosphate buffers at different temperatures (55–70 °C) and enzyme concentrations (0.276 and 0.552 mL/100 mL). Maximum -amylase activity was observed at pH=3.4 in both buffers. Effects of the addition of calcium chloride during and after thermal treatments were also investigated. α-amylase activities were measured by an iodometric method and thermal inactivation constants and D values (time for reducing 90 % of the enzymatic activity) were estimated. The enzyme was more sensible to pH changes and heat when citrate ions were present in the reaction medium. If Ca2+ in the enzyme structure is bound to citrate then the resistance of the enzyme to pH changes and heat is lowered. Kinetics obtained according to Arrhenius equation and two enzymatic fractions (thermo-labile and thermoresistant) were observed too. In citrate buffer the following relation was observed for thermo- labile fraction: log (D value) = – 0.144 t/°C + 12.992. The level of thermal inactivation also depended on the enzyme concentration. Higher thermal inactivation rates were obtained by increasing the enzyme concentration in the case when citrate was present. It was also found that the addition of calcium chloride (1 g/L) after thermal treatment in media containing citrate reactivated the enzyme treated at 60 and 65 °C. The possible implications of these findings in apple juice processing were discussed.

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