This work provides a systematic and comprehensible lineshape analysis of the Raman spectra from (i) the main esters that compose the soybean biodiesel, namely Palmitate, Stearate, Oleate, Linoleate and Linolenate, (ii) mixtures of these esters, (iii) a soybean biodiesel reference material. Using theoretical spectral simulation as a guide, the experimental spectra are fit considering all the 3N-6 vibrational modes, where N is the number of atoms in the molecule. We demonstrate that while intensity analysis may lead to uncertainty in the definition of the relative ester contents in a mixture, a complete spectral analysis involving all parameters, including peak frequencies, allows spectral determination of esters composition. Finally we define a protocol that can be used for defining the specific composition of unknown, including adulterated biodiesels, important for quality control. (C) 2013 Elsevier Ltd. All rights reserved.