An impact of nitrogen, phosphorus and potassium fertilizers on the concentration changes of carotenoid pigments in the Indigofera tinctoria plant by using Raman spectroscopic techniques is studied. Three different concentration levels of the fertilizers with a normal supply as control were added to the plants at two stages. The Raman spectra were taken to determine the carotenoid concentration level changes in the plant leaves in vivo. The ring-stretching mode are the Raman spectroscopic signatures for the carotenoid pigment and its magnitude increased significantly (over 170%) for the case of phosphorus and potassium fertilizers. The effect from the nitrogen fertilizer was detected to be about 130% in comparison with the corresponding control plants. This study has a potential application for the increased extraction of the indigotin dye from plants for the medical and textile industries.