Bovine adrenal phenylethanolamine N-methyltransferase is activated by inorganic phosphate. Addition of potassium phosphate to the enzyme assay mixture increased the enzyme activity 5- to 8-fold depending on the purity of the enzyme. Neuraminidase treatment of the enzyme decreased the activation by one half. The activity of the enzyme was also increased by other negatively charged ions, including chloride and sulfate. However, the activation was less than 3-fold, and pretreatment with neuraminidase did not alter the degree of activation. The high degree of activation was specific to bovine adrenal enzyme. Rat adrenal enzyme was activated less than 2-fold by phosphate and not affected at all by chloride and sulfate. These results suggest that bovine adrenal phenylethanolamine N-methyltransferase is a glycosylated protein, containing sialic acid moieties, and that this carbohydrate moiety plays a role in the activation of this enzyme. These results are also consistent with our previous finding [D. H. Park, E. E. Baetge, B. B. Kaplan, V. R. Albert, D. J. Reis and T. H. Joh, J. Neurochem. 38, 410 (1982)] that the difference in characteristics between bovine and rat adrenal phenylethanolamine N-methyltransferase is due to posttranslational modification of the enzyme.