A batch type chemiluminescence (CL) determination of enoxacin is described. In this work, it was observed that enoxacin could enhance the chemiluminescence (CL) emission Ru(phen)32+–Ce(IV) system and this enhancement effect was dependent on the concentration of enoxacin, based on which, CL system was established for the determination of enoxacin. Under the optimum experimental conditions, the linear range and detection limit are 0.6406–64.06 μg/ml and 0.0210 μg/ml, respectively. The R.S.D. is 1.75%. (n=10). The proposed method has been applied to detect the content of enoxacin in pharmaceutical formulation and human serum with satisfactory results. The possible mechanism of the CL reaction was discussed.