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The Fourier transform infrared (FTIR) imaging technique was used in a transmission model for the evaluation of twelve oral hyperkeratosis (HK), eleven oral epithelial dysplasia (OED), and eleven oral squamous cell carcinoma (OSCC) biopsy samples in the fingerprint region of 1800–950 cm−1. A series of 100 µm × 100 µm FTIR imaging areas were defined in each sample section in reference to the hematoxylin and eosin staining image of an adjacent section of the same sample. After outlier removal, signal preprocessing, and cluster analysis, a representative spectrum was generated for only the epithelial tissue in each area. Two representative spectra were selected from each sample to reflect intra-sample heterogeneity, which resulted in a total of 68 representative spectra from 34 samples for further analysis. Exploratory analyses using Principal component analysis and hierarchical cluster analysis showed good separation between the HK and OSCC spectra and overlaps of OED spectra with either HK or OSCC spectra. Three machine learning discriminant models based on partial least squares discriminant analysis (PLSDA), support vector machines discriminant analysis (SVMDA), and extreme gradient boosting discriminant analysis (XGBDA) were trained using 46 representative spectra from 12 HK and 11 OSCC samples. The PLSDA model achieved 100% sensitivity and 100% specificity, while both SVM and XGBDA models generated 95% sensitivity and 96% specificity, respectively. The PLSDA discriminant model was further used to classify the 11 OED samples into HK-grade (6), OSCC-grade (4), or borderline case (1) based on their FTIR spectral similarity to either HK or OSCC cases, providing a potential risk stratification strategy for the precancerous OED samples. The results of the current study support the application of the FTIR-machine learning technique in early oral cancer detection. [ABSTRACT FROM AUTHOR]