Objective To establish an HPLC method for the simultaneous determination of CTX and DDP and to research their pharmacokinetics in rat. Methods The separation was performed on a Hypersil-CTX. Results Retention time of CTX was 5.15 min. The linearity range of calibration curve was 0.5-10 μg/ml (r =0.9999), the average recovery was 97.4 %(RSD＜5 %). The detection limit was 0.1 μg/ml(S/N＞3). For DDP the retention time was 9.26 min and the linearity range of calibration curve was 0.1-2.0 μg/ml (r =0.9998).The average recovery was 98.1%(RSD＜5 %). The detection limit was 0.1 μg/ml (S/N＞3). Conclusion The method is effective for determination of CTX and DDP. It also provides the method eligible for pharmacokinetics of CTX and DDP. Drug combination make eliminating rate decrease and eliminating half life prolong, have effect on each other.