In order to reveal the relationship between Lilium male sterile and fertile lines in changes of hormones, some endogenous substances contents and energy metabolism enzyme activities, this study used chromatographytandemmass spectrometry and high performance liquid chromatography to test hormones. Endogenous substances and energy metabolism enzyme activities were measured by colorimetry. The effects of hormone content, hormone ratio, endogenous substance content and energy metabolism enzyme activity on male sterility of Lilium were analyzed. The results show that: （1） during the development period which is from sporogenous cell stage to tetrad stage, changes of endogenous hormone, endogenous substance content and energy metabolic enzyme activity in male sterile line and fertile line were different. The content of IAA in male sterile line was higher than that in fertile line at all stages. The contents of ZR and GA₄ in male sterile and fertile line were similar at pollen mother cell stage, but two contents in male sterile line were lower than those in fertile line at tetrad stage. ABA content in male sterile line accumulated at tetrad stage. JA content in male sterile line was lower than it in the fertile line at the pollen mother cell stage. However, the trend is opposite at the tetrad stage. （2）The contents of soluble sugar, soluble protein and proline in fertile line were higher than those in male sterile line at all stages. However, the content of MDA in male sterile line was significantly higher than that in fertile line at tetrad stage. （3）The activities of COD and ATP in fertile line were higher than those in sterile line at all anther development stages. This study suggest that: from sporogenous cell stage to tetrad stage, high IAA, ABA and low GA₄, ZR contents may lead to the deficiency of soluble sugar, soluble protein, proline contents and lower energy metabolism enzyme activities. These are the possible reasons to cause male sterile phenomenon of Lilium.