This study employed Fourier transform infrared (FTIR) spectroscopy to determine the chemical composition of brain tissues and the changes induced by irisin at doses of 50 mg and 100 mg. Brain tissues were collected from control rats and those administered with irisin, and key vibrational peaks were analyzed. In the 50 mg irisin group, all described vibrations decreased compared to control tissues, while the 100 mg group showed a decrease only in lipid vibrations. Comparatively, the 50 mg group had lower absorbance of phospholipids, amides, and lipid functional groups than the 100 mg group. Lower amounts of these compounds were found in treated tissues compared to controls, with higher levels in the 100 mg group. Ratios between amide peaks revealed significant differences between groups. Principal component analysis (PCA) differentiated control and irisin-treated tissues, primarily using PC1 and PC3. The decision tree model exhibited high classification accuracy, especially in the 800-1800 cm⁻1 range, with high sensitivity and specificity. FTIR spectroscopy effectively highlighted chemical changes in brain tissues due to irisin, demonstrating dose-dependent variations. The combination of PCA, ROC analysis, and decision tree modeling underscored the potential of FTIR spectroscopy for studying the biochemical effects of compounds like irisin.