Milk is a perishable product. The causes of milk breakdown can be caused by various factors, such as pathogenic bacteria. Plantaricin IIA-1A5 is a bacteriocin, antimicrobial substrate that is produced by Lactobacillus plantarum IIA-1A5, a lactic acid bacteria isolated from fresh beef at traditional market in Indonesia. Plantaricin IIA-1A5 has properties to reduce the number of bacteria contamination in food. The purpose of this study was to determine the effect of crude plantaricin on fresh milk from dairy cows in terms of physicochemical, pH, and microbiological aspects. Three types of treatment on milk consisting of untreated (control), plantaricin 3.66 ml, and synthetic antibiotic penicillin 3.66 ml. The three milks were storage at room temperature at 4-time intervals, 1 h, 3 h, 5 h, and 7 h since treatment was given with 3 repetitions. Physicochemical testing was carried out using lactoscan, pH testing using a pH meter, and total microbes. The results showed that there was no significant difference in the physicochemical properties of milk and pH, but there was a significant difference (P = 0.05) in the S. aureus population at the 6th hour after giving plantaricin treatment. Therefore, it can be concluded that plantaricin has the potential as an alternative preservative for cow milk.