t is a simple, sensitive, rapid and economic chromatographic method has been developed for the determination of cytarabine in rat plasma using standard API. This analytical technique used for development was high performance thin layer chromatography. HPTLC aetron with already coated silica gel plates 60 F 254 (10X10 cm) at 250 nm thickness was used as stationary phase. the mobile phase used consisted of 2-butanone :acetone :water (65:20:15 % v/v/v)[4] the plasma sample were extracted by protein precipitation with methanol concentration ranges of 1000,2000,3000,4000,5000 ng/ml respectively ,were used mixed plasma for the calibration curves .the stability of cytarabine in plasma were confirmed during three freeze-thaw cycles (-20 ºC ) on a bench for 24 hrs and post preparattively for 48 hrs.this method was validated statistically and proved suitable for the determination of cytarabine in rat plasma.