Anais do 15° SLACAN - Simpósio Latino Americano de Ciência de Alimentos e Nutrição

Biochemical characterization is fundamental to determining the conditions under which enzymes can be applied in various industrial processes. Immobilization is a powerful technique to overcome the disadvantages of soluble proteins due to their confinement in solid supports, insolubilizing the catalysts and allowing for their reuse and easy separation from the reaction medium. This work aimed to determine the temperature and pH for maximum activity of amylases immobilized in sodium alginate, as well as their stability at different temperatures and pH levels. Amylases were produced by Rhizopus arrhizus I1.2.1 in a submerged culture medium containing starch and urea as carbon and nitrogen sources (respectively), and magnesium sulfate and potassium phosphate as salt solutions at 30°C, under stationary conditions, for six days. The amylases were immobilized by mixing 100mL of the crude enzyme extract with 4.70g of sodium alginate. After mixing, the solution was dripped into a 6.26% (w/v) calcium chloride solution in distilled water. The amylolytic activity was quantified using the saccharifying method to obtain reducing sugars, using 4 beads and 1000µL of 1% (w/v) starch substrate in 100mM sodium citrate buffer (pH 6). The effect of temperature and pH on the enzymatic activity of the immobilized derivative was evaluated using a DCCR experimental design, varying the pH (4.0-8.0) and temperature (40-80°C). Stability at different temperatures (50-70±5°C) and pH (3-10±0.5) was evaluated for up to 60min. It was observed that the apparent optimal temperature and pH were 64°C and 6.02, respectively, with an R2=92%. Stability analysis revealed that the immobilized derivative exhibited stability with residual activity exceeding 50% at temperatures of 50 and 55°C, and at around pH 5. From the above, it was noted that the amylases under study have the potential for application in industrial processes, given their obtained activity and the formation of reducing sugars through starch hydrolysis.