COMPARISON OF DIFFERENT CONCENTRATIONS OF BETA-GALACTOSIDASE ENZYME IN THE PERCENTAGE OF LACTOSE HYDROLYSIS

The enzymatic hydrolysis of lactose by the enzyme beta-galactosidase plays an important role in the processing of dairy products, such as the production of low-lactose products for the consumption of intolerant individuals. In this work, the enzyme beta-galactosidase was produced by culturing the microorganism Saccharomyces fragilis IZ 275 in culture environment based on fresh cheese whey. The objective of this study was to compare the different concentrations of beta-galactosidase enzyme in the percentage of lactose hydrolysis. The fermentations were conducted at 150 RPM, at 35°C for 24 hours. After the fermentation the extraction was carried out by ultrasound using the Box-Benhken statistical design with 9 runs. The rupture of the yeast cells is done by an ultrasonic processor (UNIQUE, Model: DES500) with frequency of 20KHz and power of 500 Watts (RMS) using a titanium macro (13 mm diameter), maintaining suspension during the time delimited by the statistical design. After the extraction, the enzymatic activity analysis was performed by the ONPG of the design, the maximum enzymatic activity was observed in the run 9, under the conditions of extraction of 9 minutes and power of 372 W/cm². Having presented the best result, the run 9was chosen for the experiment. Extractions of 1 and 10% (v/v) of the supernatant containing the enzyme extracted were added in 10 mL of 5% lactose solution, incubated in an oven at 37°C, for 10 hours, withdrawing aliquots every 2 hours, inactivating them at 95°C for 10 minutes. The determination of glucose was performed by the glucose oxidase method, using the Glucose Kit (Laborclin®). To carry out the analysis, 1 mL of glucose was added in test tubes, to which 10 μL of the lactose solution were added by incubating them in a water bath for 10 minutes at 37°C, at the end of the reaction, the samples were incubated in a bath of ice and measured in a spectrophotometer at 505 nm, considering as white the glucose oxidase solution. It can be observed that after 8 hours of hydrolysis, the solution containing 10% of extracted enzyme obtained a higher result when compared to hydrolysis with 1% of enzyme, being 15.7% and 69.8%, respectively. On completion of 10 hours of hydrolysis, the 1% solution kept the hydrolysis percentage increasing with 27.6%, however the 10% enzyme solution showed a decrease in the percentage of hydrolysis, due to the excess glucose released in the solution, thus causing inhibition of the beta-galactosidase enzyme.