Tonle Sap lake has been the largest fresh water lake in Southeast Asia, providing a fundamental environment for nurturing more than three million people in the five surrounding provinces, which about 90% lived from the fishery or agriculture. However, the ecosystem and populations living around the lake have experienced environmental pressure, since the lake water, fishes, living organisms are polluted. More than that, sediment is the type of soil which provides beneficial nutrients, and it also has been polluted by discharge of untreated pesticide residues from the agriculture actions. This current study aims to identify the presence of 451 pesticide compounds and quantify the concentration of 23 targeted pesticide residues in sediment at Chhnok Trou community of the Tonle Sap Lake. Furthermore, pesticide residues were extracted by Solid-Phase Extraction (SPE) method and detected by Gas Chromatography-Mass Spectrometry (GC-MS). Different cartridges (C18 and PLS3-AC2) and different ratio of elution solvents were tested to validate the extraction method before the actual sample analysis. The recovery yield of combination of amino and activated carbon cartridges using mixture of methanol (MeOH)/dichloromethane (DCM) 10/90 (v/v) for elution step was higher than those of another two conditions of MeOH/DCM: 5/95 and 50/50 (v/v). For the recovery rate of silica cartridge, using mixture of acetic acid/acetonitrile of 1/99 (v/v) for elution step gave higher recovery rate than those of another two mixing conditions of acetic acid/acetonitrile: 3/97 and 5/95 (v/v). In this study, combination of amino and activated carbon cartridges and 10/90 (v/v) of methanol/dichloromethane as a mixture solvent for elution step were used for pesticide extraction and clean-up for sediment samples. As a result, four pesticide residue compounds were presented in sediment samples; whereas, none of targeted compounds were quantitatively detected. Qualitatively, chlorfenapyr, insecticide and acaricide were found in sample JS2, fluquinconazole and fungicide in JS5, mevinphos and insecticide in JS3, while oxabetrinil and herbicide safener were found in JS4 and JS6.