Glaucoma is one of the major causes of irreversible blindness in the world. Detecting glaucoma as early as possible ensures the best possible treatment and outcome. Since there are currently no glaucoma-specific biomarkers identified and no glaucoma screening method available, we decided to analyze molecular structure of aqueous humor using FTIR spectroscopy which offers a unique opportunity to investigate the composition of substances on a molecular basis. The study included 87 age matched subjects, divided into two groups 1) 55 glaucoma patients (158 aqueous humor samples) and 2) a control group comprising 32 patients with cataracts (93 aqueous humor samples). All aqueous humor samples were collected at the start of the glaucoma or cataract surgery. FTIR spectra, of the samples dried on transparent silicon plates, were recorded and analyzed by statistical methods of STT, PCA and SVM. Our study has shown that FTIR spectroscopy can differentiate media structure between the control group and the glaucoma patient group. A model for spectra recognition attributable to a certain type of pathology was established. Results of FTIR spectroscopy analyzed by statistical methods of STT, PCA and SVM have proven to be a promising method for differentiation of aqueous humor samples between patients with cataract and glaucoma, therefore this method can form the basis for the development of new methods in early diagnostics of glaucoma.