Biodiesel susceptibility to oxidation is one of the major problems concerning its use as an alternative to diesel fuel. Although well-characterized, the oxidation process cannot be completely prevented since it can be affected by a large number of factors, such as fuel composition, storage conditions, contaminants, temperature, and the presence of air and light. In this work, we propose Fourier transform infrared spectroscopy (FTIR) in association with principal components analysis (PCA) and hierarchical cluster analysis (HCA) as a method for monitoring the extent of oxidation degree of biodiesel in the low rate phase, before the end of the induction period, in which the changes in the physical properties such as viscosity, mass density, and chain composition of the sample remain almost undetectable. A detailed investigation of thermo-oxidation of biodiesel is reported for a mixture of 50/50 (%) of soybean oil and animal fat biodiesels. The biodiesel degradation was accelerated when maintaining the temperature of the sample at 110 degrees C under constant air flux for different times. Oil stability index, mass density, viscosity, gross caloric value, esters composition, UV-vis, and FTIR spectra were measured in order to analyze the degree of degradation of each sample. The multivariate analysis in the FTIR spectra clearly shows the discrimination of the samples in the first stage of the degradation process. These results could be useful as a method for monitoring the fuel quality during storage, helping fuel producers, suppliers, and users.