Determinação da inativação e estudo da cinética de reativação da fosfatase alcalina em leite pasteurizado sob diferentes condições de tratamento térmico

Abstract

Milk is one of the most relevant commodities in the world. It is estimated that about 10% of the world's population depends directly on dairy farming. Brazil is currently the third largest producer of milk in the world, adding more than 34 billion liters/year, behind only the United States and India. Dairy products contribute significantly to the needs of proteins, carbohydrates, lipids and micronutrients, being the main responsible for the supply of calcium to our organism; in addition to contributing to the intake of magnesium, selenium, riboflavin and vitamin B12. In Brazil, fluid milk to be consumed in natura must undergo a pasteurization process provided for in current legislation. To attest that the thermal processing was carried out effectively, immediately after pasteurization, the milk must show a negative test for alkaline phosphatase and a positive test for peroxidase. Thus, the objective of this work was to determine the activity of alkaline phosphatase in milk subjected to different heat treatment conditions, in addition to investigating a possible reactivation of the enzyme over a 72-hour milk shelf-life. Qualitative methods (official adopted by the Ministry of Agriculture, Livestock and Food Supply) and quantitative methods (Scharer colorimetric described by the American Public Health Association) were applied. With the results of this work, it was possible to define a parameter in which the official qualitative method detected the presence of alkaline phosphatase, which was defined as 4.13 µg of phenol mL-1 of milk. Furthermore, it was shown that there is no direct relationship between the addition of raw milk and the generated phenol concentration. Thus, the use of milk doping to evaluate the official qualitative method is not recommended, since the same dosages of raw milk may result in different concentrations of phenol, due to the composition of each milk. The data showed that the composition of the milk used in the heat treatments influenced the inactivation of the enzyme and the fat, apparently, was one of the components responsible for this result. It was shown that the official method was not adequate to verify the pasteurization of milk, since it did not detect the presence of the enzyme in a subpasteurization sample (70ºC/20s). Furthermore, the limit expressed by the Scharer method (1 µg phenol mL-1) to verify the pasteurization of the milk did not prove to be appropriate, since the properly pasteurized samples (72 ºC/20s, 75 ºC/20s and 65 ºC/ 30 minutes) presented values above the stipulated by the method. Some heat-treated milk samples showed an increase in residual alkaline phosphatase activity over the shelf-life, indicating that the enzyme may have undergone reactivation after heat treatment. It was concluded that the official method did not show a good sensitivity for alkaline phosphatase detection, it was verified that the limit parameter to certify the pasteurization efficiency of the quantitative method must be adjusted for each type of milk: skimmed, semi-skimmed and whole; and the requirement to perform the ALP test immediately after the milk pasteurization process was reinforced, in order to minimize the chances of false-positive results due to possible reactivation of the enzyme.