The administration of ceftriaxone may reduce the incidence of postoperative infections in patients undergoing surgical procedures. A simple, sensitive and selective high - performance liquid chromatographic method for analysis of ceftriaxone in perilymph fluid by direct injection without any sample pretreatment was developed. Analysis was carried out on Nucleosil C18 100-3 (125 x 4.6 mm, i.d., 5 mu m) column with a mixture of methanol: citrate buffer (30 mM, pH 3.0) (25:75, v/v) as the mobile phase, at a flow rate of 1.0 mL/min and at 270 nm using ultraviolet detector. Sulfisoxazole was used as an internal standard. The optimized method was validated in terms of linearity, sensitivity, specificity, accuracy, precision, recovery, ruggedness and robustness. The linearity range was found to be 0.05-1.00 mu g/mL. The developed method was successfully applied for quantitative determination of ceftriaxone in spiked perilymph fluid and perilymph fluid of patients' intravenous administered ceftriaxone.