Unlike traditional passive optical sensors, hyperspectral LiDAR emits the active, full-waveform and gaussian laser pulse. After interacting with the vegetation leaf surface, the backscattered intensities for different waveforms return to the receiver and are then recorded. Previous research on spectral reflection characteristic have only focused on the circumstance at the incidence angle of 0°, the reflection characteristics at other multiple incidence angles and their impacts and errors on leaf chlorophyll content estimation have seldom been studied. This study used the hyperspectral LiDAR with 32 bands developed by our lab to obtain the leaf reflection spectrum over different incidence angles, and the intricate reflection characteristic was then analyzed at the bands with the high signal-to-noise ratio. After that, spectral indices were chosen to study the impact of leaf bi-directional reflection characteristics measured by hyperspectral LiDAR on leaf chlorophyll content estimation. The results show that, (1) the returned intensity of vegetation leaf measured by hyperspectral LiDAR gradually decreases as the incidence angle increases, but the bi-directional reflectance factor (BRF) does not show the same trait with intensity. There are two different features with the incidence angle in the visible and near-infrared bands. The maximum BRF value for visible bands occur in the incidence angles of 0° to 10°, but 60° for the maximum BRF values for the near-infrared bands. The minimum BRF values for all bands both occur in the incidence angle of 45°, and the difference between the maximum and minimum values is about 0.1. The changing trend of the BRF at the incidence angle of 10° to 60° for the visible and near-infrared bands decreases first and then increase; (2) BRF has a great impact on chlorophyll retrieval accuracy based on the relationship analysis on spectral indices and chlorophyll content at different incidence angles. However, there is not a synchronous decrease or synchronous increase trend for R2 and RMSE. Specifically, R2 decreases first, then increases, and finally decreases to the minimum at the incidence angle of 50°, in which the increase occurs at 60°. RMSE presents the opposite changing trend. For the different spectral indices, R2 owns a 4 times fluctuation with 0.14~0.63. RMSE owns about 1.5 times fluctuation with 0.5~0.8 mg·g-1. The great changes of R2 and RMSE reveal the essential impact of bi-directional reflection characteristics on leaf chlorophyll content retrieval.