Abstract: There were 20 hens in laying period used for test and each hen was administered in single cage. These hens were given intramuscular injection with a single dose of 5 mg/kg every morning and evening, and the injection lasted 5 d. Other than that, these hens were fed in normal. From the first day of drug injection, the eggs were collected and the concentration of colistin antibiotics in eggs was determined by high performance liquid chromatography tandem mass spectrometric (HPLC-MS/MS). The colistin concentration-time data in eggs were analyzed by Winnonlin program. The main pharmacokinetic parameters of eggs were as follow: Cmax was 1 411.252(578.2536) g/kg, Tmax was 6.425844(0.666114) d, AUC was 1 0647.39(3159.706) g/kgd, K01-HL was 2.173256(1.115728) d, K10-HL was 2.237842(1.072385) d and CL was 0.000503(0.000124) L/d.