Color change of pupil area was investigated in Red, Green, Blue (RGB)and Hue, Saturation, Intensity (HSI)color models from July to November in 2010 and from May to December in 2011to estimate the serum vitamin A level of Japanese black cattle during their vitamin A controlled stage. A 2CCD camera was used to acquire the eye images. The results showed lower vitamin A level cattle usually accompanied with higher red component value, lower saturation in their eye images. An estimation model was built based on red and green component ratio. The estimation error is about 10 IU/dL. The reasonable result shows the feasibility to estimate the vitamin A level by color change of pupil area in Japanese black cattle.