UDC 557.152.321:663.316 ISSN 1330–9862 original scientific paper (FTB-1100) Amylase for Apple Juice Processing: Effects of pH, Heat, and Ca 2+ Ions Liliana N. Ceci and Jorge E. Lozano* PLAPIQUI (UNS-CONICET) Camino La Carrindanga Km.7, C.C. 717 8000 Bahía Blanca – Argentina Received: July 4, 2001 Accepted: November 8, 2001 Summary The aim of this paper was to evaluate the effects of pH, heat, and Ca 2+ ions on the a-amylase activities in a commercial amylolytic enzyme (Tyazyme L300), used for apple juice processing. Kinetics of thermal inactivation was studied in acetate and citrate/pho- sphate buffers at different temperatures (55–70 °C) and enzyme concentrations (0.276 and 0.552 mL/100 mL). Maximum a-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. a-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 Ca 2+ 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 thermo- resistant) were observed too. In citrate buffer the following relation was observed for ther- mo-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 obtai- ned 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 implica- tions of these findings in apple juice processing were discussed. Key words: a-amylase, apple juice, starch hydrolysis Introduction Apple juice may have a high content of starch, when processed from some apple varieties or unripe fruits at the beginning of the season (1). High insoluble starch concentrations (7.68 g/L) were detected in pressed »Granny Smith« apple juice from unripe fruits har- vested in »Río Negro« Valley region (Argentine), two weeks before the usual harvest. Soluble starch was also present in these cloudy juices but in significantly lower concentrations (0.51 g/L). The starch contents of pressed cloudy juices were gradually lower when unripe fruits were stored at room temperature and soluble and insol- uble starch were not detected three weeks after the har- vest. Insoluble starch was gelatinized when cloudy jui- ces were pasteurized. Polymeric carbohydrates, as starch, may cause slow filtration, membrane fouling, jellying after concentration or post-process cloudiness. However, starch may be eas- ily detected and treated by adding starch-degrading en- 33 L. N. CECI and J. E. LOZANO: Amylase for Apple Juice Processing, Food Technol. Biotechnol. 40 (1) 33–38 (2002) * Corresponding author; Phone: +54 291 486 1700; Fax: +54 291 486 1600; E-mail: jlozano@plapiqui.edu.ar