This contribution describes the development of a chromatographic method capable of simultaneously identifying and quantifying the constituents found in oleaginous biomass including algae oil-and in biofuel samples through a single cost-effective analysis. Major constituents of the aforementioned analytes include oxygen- and/or nitrogen-containing compounds along with fuel-like hydrocarbons. A novel simulated distillation gas chromatographic (SimDist) technique including dual detection capabilities-specifically flame ionization (FID) and mass spectrometry (MS)-produced identical chromatograms with perfectly aligned signals. FID chromatograms afforded excellent quantitative data while the corresponding MS spectra enabled accurate and thorough compound identification. Simulated distillation data displayed a remarkably linear relationship between retention time and the boiling point of heteroatom-containing compounds in addition to n-alkanes. Indeed, although standard SimDist data were calibrated using the boiling points of n-alkanes, analyses involving other compounds yielded insights into the effect of additional functionalities on both retention time and response factor. Notably, the method developed proved superior relative to commonly employed techniques in the identification and quantification of polyunsaturated fatty acids in algae oil.