A simple and sensitive fluorescent assay was developed for the sensitive and selective determination of cefixime based on inner filter effect (IFE). In this sensing platform, fluorescent carbon dots (CDs) were prepared by one-pot synthesis and was directly used as fluorophore in IFE. The method is based on the complexation reaction between cefixime and palladium ion in the presence of acidic buffer solution (pH 4). Production Pd(II)-CEFcomplex induced the absorption band which complementary overlap with the excitation spectra of CDs. Due to the competitive absorption, the excitation of CDs was significantly weakened, resulting in the quenching of CDs. The present IFE-based sensing strategy showed a good linear relationship from 0.2×10-6 M to 8 ×10-6 M (R2 = 0.994) and provided an exciting detection limit of 0.9×10-7 M (3δ/slope). The proposed method has been successfully applied for the determination of cefixime in pharmaceutical preparations and human urine samples.