The use of vibrational spectroscopy to probe environmental samples is increasing with the development of new methods, including microspectroscopic probes. In this study, we compare different vibrational methods to interrogate lipids and other compounds found in environmental samples. In particular, we compare the vibrational spectra for different lipids that include fatty acids (protonated and deprotonated forms), fatty alcohols, and fatty esters by utilizing attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy, optical photothermal infrared (O-PTIR) spectroscopy, micro-Raman spectroscopy, and atomic force microscopy infrared (AFM-IR) spectroscopy. We show the utility of infrared methods to clearly delineate the structure of the lipid, i.e., whether it is an acid, alcohol, or ester. In contrast, it is difficult to differentiate these from micro-Raman spectroscopy. Furthermore, in the case of fatty acids, the protonation state can also be determined by infrared methods. In most cases, there is a high correlation between the three different infrared techniques as seen for ATR-FTIR and O-PTIR spectroscopy
however, this is not always true with AFM-IR spectroscopy for samples with low signal-to-noise or in a liquid phase state. Additionally, substrate-deposited aerosols were collected from the Scripps Ocean-Atmosphere Research Simulator (SOARS) and examined with both the O-PTIR and micro-Raman spectroscopy to show how these two vibrational probes together can provide essential chemical insights into environmental samples that are difficult to achieve otherwise.