Two kinds of activated carbon (AC)-supported Pt catalysts with similar particle sizes were respectively synthesized using the polyol method and incipient wetness impregnation method, and then tested for glycerol oxidation in a base-free condition. It is revealed that the initial reaction rate on the Pt catalyst prepared by the polyol method (Pt/AC(PO)) is nearly triple that on the Pt catalyst prepared by incipient wetness impregnation method (Pt/ACIWI), and the Pt/ACIWI catalyst favors further oxidation of glyceraldehyde to glyceric acid and more CC bond cleavage, which might mainly arise from the difference in relative coverage of oxygen to glycerol caused by the different electronic properties of Pt as well as the difference in the catalyst surface properties. Moreover, the deactivation mechanisms of both catalysts were also investigated. The results show that strongly adsorbed intermediates contribute significantly to the deactivation of the Pt/AC(PO) catalyst, while both the oxygen poison and the adsorption of intermediates on Pt surfaces cause the deactivation of the Pt/AC(IWI) catalyst.