A method of chiral separation and determination of acephate and methamidophos enantiomers in tea matrix was developed and validated.The efficiencies of two different commercial chiral columns to separate enantiomers of these two pesticides were evaluated and the gas chromatography condition of the better one（BGB-176） was optimized.After soaked in 1 mL water,the tea samples were extracted with 10 mL acetonitrile and purified with C18,primary secondary amine（PSA） and graphitized carbon black（GCB）,then determined by gas chromatography with the external method.The results showed that the recoveries of acephate and methamidophos enantiomers at three spiked levels of 0.04,0.08,0.40 mg/kg were in the range of 58.3%-66.4% and 50.8%-57.6%,respectively.The relative standard deviations（n=5） were all less than 7%.The limits of detection of these two pesticides enantiomers ranged from 0.003 mg/kg to 0.01 mg/kg.The method is quick,convenient and reliable,and could meet the requirement of analysis.The result also showed that the degradations of acephate and methamidophos in tea garden were enantioselective.