[Objective] The aim was to study the chemical constituents of Euphorbia hirta. [Method] The constituents were isolated and purified by various chromatography methods including normal phase and reverse phase silica gel as well as Sephadex LH-20 gel column, and then elucidated by spectroscopic analysis such as MS and NMR etc. Compounds 1, 4, 5 and 6 were evaluated for their immunostimulatory activity against splenic cell of mouse by FCM. [Result] Six compounds were isolated from Euphorbia hirt, and identified as diisobutyl-O-phthalate （compond 1）, diethylhexyl phthalate （compond 2）, hispidulin （compond 3）, acetyl peroxide （compond 4）, quercetein （compond 5） and gallic acid （compond 6）. All compounds were inactive against splenic cell of mouse. [Conclusion] Compounds 1, 2 and 3 were isolated from this plant for the first time, and compound 4 was the first naturally occurring compound. This study provided reference for isolating bioactive substances from Euphorbia hirta.
