Objective:The purpose of this studywas to optimize the conditions of analyzing oral bacterial end products by using reversed- phase high performance liquid chromatography (RP-HPLC).Methods:Different culture fluid, temperature, solution pH, extract, deposition were used to investigate oral bacterial end productswithHPLC. The optimal chromatographic parameterswere: flowrate of 110 mlPmin and 210 mlPmin, detection UV at 217 nm, 5% methanol in phosphate buffer solution (pH218) as mobile phase, linear gradientwashwith 5% to 40% methanol for10 min, at ambient temperature.Results:Oral bacteria cultured in brain heart infusion broth (BHI); their supernatantwas adjusted to pH117, and then extracted by ether. With the above conditions, an opti- mal result can be obtained by analyzing using HPLC.Conclusion:The results of this paper will facilitate the identification and classification of oral bacteria by analyzing oral bacterial end products.