OBJECTIVE：To investigate th e effects of juglone on brain tissu e of rats and its relationship with the biomarkers related to brain tissue injury. METHODS ：Totally 40 rats were divided into blank group ，juglone high-dose ，medium-dose and low-dose groups （34.832，17.416，8.708 mg/kg），with 10 rats in each group. They were given medicine intragastrically once a day ， for consecutive 4 weeks.After last administration ，the general behavior ，brain index and brain tissue morphology were investigated. UPLC-MRM-MS method was used to determine the serum contents of L-dopa（L-Dopa），L-tyrosine（L-Tyr）and L-tryptophan（L-Trp） in rats. The chromatographic condition included Waters Acquity UPLC BEH C 18 column，mobile phase consisted of ammonium acetate aqueous solution-acetonitrile ，gradient elution ，at the flow rate of 0.3 mL/min，sample size of 5 μL，column temperature of 30 ℃；MS condition include electrospray ion source ，in positive ion mode ，capillary voltage of 3 500 V，desolvent gas flow of 650 L/h，desolvent temperature of 350 ℃，ion source temperature of 110 ℃. RESULTS ：Compared with blank group ，the rats in each dose group showed the behavior of tiredness and weakness of limbs. The brain tissue morphology showed pathological changes ， which contained blood vessel congestion in the cerebral and cerebellar cortex ，partial cell nucleus pyknosis in the pyramidal cell layer，deep staining of nuclei ，irregular shape and unclear boundary and other pathological changes ；the brain index of juglone high-dose group increased significantly （P＜0.05）. The established UPLC-MRM-MS method showed good specificity and the range of L-Dopa，L-Tyr and L-Trp were 31.25-32 000，31.25-32 000，15.625-16 000 ng/mL（r＝0.999 1-0.999 9），respectively. The limits of detection were 6.250，5.625，3.125 ng/mL，respectively. The limits of quantitation were 15.625，18.75，10.00 ng/mL，respectively. RSDs of precision ，accuracy and stability （24 h）tests were all Matrix effects were 95.1％-100.1％ （RSD are not more than 3.25％，n＝3）. Compared with blank group，the contents of L-Dopa were increased significantly injuglone medium-dose and high-dose groups （P＜0.01）. The contents of L-Tyr were increased significantly in juglone lowdose，medium-dose and high-dose gro ups，while the contents of L-Trp were decreased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS ：Juglone has an effect on the general behavior，brain index and brain tissue morphology of rats. It may affect the brain function of rats by increasing the contents of L-Dopa and L-Tyr in serum and decreasing the contents of L-Trp.