 A novel hybrid nanocomposite based on de-layered molybdenum disulfide (MoS2) by ionic-liquid (IL, [BMIM]BF4)-assisted exfoliation and graphene oxide (GO) was synthesized via a green, efficient, and high-quality method, which combined liquid-phase stripping method and ion-insertion method. In addition, an electrochemical sensor was developed using the MoS2-IL/GO nanocomposites for the determination of chloramphenicol (CAP). The morphology and structure of these synthetic materials were characterized by scanning electron microscopy, transmission electron microscopy, Raman spectroscopy, and X-ray diffraction and the electrochemical characterization by cyclic voltammetry and electrochemical impedance spectroscopy. Based on this method, the insertion of IL can effectively exfoliate de-layered MoS2, and the MoS2-IL/GO nanocomposite exhibit 3D structure with higher surface area, excellent electrical conductivity, and synergistic catalytic capabilities. Under optimized conditions, the sensor responded linearly to CAP ranging from 0.1 to 400 µmol L−1 and the detection limit of 0.047 µmol L−1. In addition, the sensor showed excellent stability, repeatability, reproducibility, and selectivity, and has been applied to detect CAP in eyedrops, milk, and urine samples.