Crataegus oxycantha L., commonly known as hawthorn, has traditionally been used for its beneficial effect on cardiovascular health, which is related to its flavonoid content. The aim of the present study was to evaluate the antibacterial properties of a fluid extract and a hydro-ethanolic macerate from buds of Crataegus oxycantha against clinical isolates of Ureaplasma urealyticum. The major purified flavonoids present in the extracts were also tested against ATCC strains and clinical isolates. Both the fluid extract and the hydro-ethanolic macerate were active against thirty-due clinical strains of U. urealyticum, with MIC ranges between 15.6 and 250 μg/ml and 15.6 and 62.5 μg/ml, respectively. All pure organic compounds, with the exception of rutin, showed activity against the strains tested, luteolin 3,7-diglucoside being the most active compound (MICs in the range of 0.48 and 1.95 μg/ml), followed by apigenin-7-O-glucoside (MICs in the range of 0.48 and 3.9 μg/ml). The activity of the pure flavonoids was greater against the clinical isolates compared to the ATCC strains. The data presented here demonstrate that flavonoids present in Crataegus oxycantha are effective against clinical isolates of U. urealyticum and could be used in combination with antibiotics in order to combat resistance.