Many attempts have been made to estimate the post-mortem interval (PMI) using bioanalytical methods based on multiple biological samples. Cartilage tissues could be used as an alternative for this purpose because their rate of degradation is slower than that of other soft tissue or biofluid samples. In this study, we applied Fourier transform infrared (FTIR) spectroscopy to acquire bioinformation from human annular cartilages within 30 days post-mortem. Principal component analysis (PCA) showed that sex and causes of death have almost no impact on the overall spectral variations caused by post-mortem changes. With pre-processing approaches, several predicted models were established using a conventional machine learning method, known as the partial least square (PLS) regression. The best model achieved a satisfactory prediction with a low error of 1.49 days using the second derivative transform of 3-point smoothing and extended multiplicative scatter correction (EMSC), and the spectral regions from proteins and carbohydrates contributed greatly to the PMI prediction. This study demonstrates the feasibility of cartilage-based FTIR analysis for PMI estimation. Further work will introduce advanced algorithms for more accurate and precise PMI prediction.