Determining the activity of residual alkaline phosphatase (ALP, EC 3.1.3.1) in milk validates adequate pasteurization and confirms that cross-contamination of raw milk has not occurred, thus ensuring product safety. However, flavored milks have been shown to yield false positive or false negative ALP activities, which inaccurately represents the microbiological safety of the milk sample. The aim of this study was to investigate the effects of 3 common milk-flavoring constituents on ALP activity using the fluorometric assay. Preliminary work determined no significant differences (P > 0.05) in ALP activity as a function of dual-stage homogenization (2,000 psi) or differing pasteurization methods including high temperature, short time (72°C for 15 s) and low temperature, long time (63°C for 30 min); therefore, homogenization and varying pasteurization conditions were not included as experimental treatments in the present study design. Raw milk was standardized to 3.25% milkfat. The batch of milk was divided into 4 treatment groups. Sucrose, pure vanilla extract, or cocoa solids were added at 8, 0.1, and 1.5%, respectively. A control (no ingredients) was included as the fourth group. All 4 treatments were pasteurized at high temperature, short time conditions. Experimentation was independently replicated 3 times, and each analysis was done in duplicate. ALP activity did not significantly differ among treatment groups (P > 0.05) and was adequately inactivated (<350 mU/L). However, the treatment containing cocoa solids had the highest ALP activity at 106.70 mU/L in comparison to the control at 19.32 mU/L, suggesting the primary polyphenols of the ingredient are inducing a conformational change of the enzyme or interacting with the quantified fluorescence produced. With flavored milk sales rising in recent years, ensuring product safety and understanding potential interactions is imperative in the dairy industry.