Studies of Scharer's Original Method for Alkaline Phosphatase in Milk with a Modification Utilizing an Organic Buffer

Abstract

Based on studies of Scharer's original (1938) method for alkaline phosphatase in milk, a modified procedure was developed utilizing two strengths of an organic buffer, 2-amino-2-methyl propanol. This modification, designated the AMP method, has the following advantages: (a) elimination of non-enzymatic substrate hydrolysis resulting from immersion in boiling water; (b) complete reaction of liberated phenol to form indophenol by control of temperature. pH. and volume of 2,6-dibromoquinonechloroimide; and (c) provision of a clear solution for spectrophotometric measurement following protein precipitation and filtration. The method is able to detect less than 0.1% of raw milk in pasteurized milk, and has a precision with a 1-h incubation period equivalent to the Kay and Graham method requiring 24 h incubation. A standard of 1.6 phosphatase activity units is proposed for pasteurized milk on the basis of equivalence to 2.3 Lovibond color units by the Kay and Graham method.