The spectral data were analyzed by using the Partial Least Squares Regression. The result shows that the addition of hemoglobin changes the number of principle component from one into three, while the largest contribution wavelength changes from 459 nm to 432 nm and 433 nm. This indicates that absorption noise of hemoglobin was introduced in the spectral data used for PLS model. Furthermore, compared with pure bilirubin solution, the Root Mean Squared Error of Prediction(RMSEP) of the model for mixed solution increases 24.90%, and the correlation coefficient of the model for mixed solution reduces 0.29%. It means that the addition of hemoglobin decreases the prediction ability of the model. Meantime, a new absorption peak at 408 nm appears, which seems there is a hypochromic effect, due to some weak key combinations between hemoglobin and bilirubin. And the new absorption peak changes the number of principal components and reduces the predictive ability of the model. Overall, the influence of hemoglobin should be reduced in order to improve the transcutaneous detection accuracy of bilirubin.