Mass spectrometry (MS) with low temperature plasma (LTP) as an ionization source is one of the widely used ambient methods in analyzing various bio-chemical samples for their detection, identification, differentiation, etc. While the LTP-MS allows selective analysis of a sample with low-molecular weight without thermal damages, it has difficulties desorbing a target molecule from the sample surface with a low volatility. Hence, for the purpose of enhancing the desorption and ionization efficiencies of the LTP-MS when analyzing the fatty acids, two methods were compared: directly heating the sample surface and increasing the vapor pressure of the fatty acid itself by derivatization. In addition, the fatty acids in Drosophila were directly analyzed with the LTP-MS. As a result, it was found that the LTP-MS detection efficiency was enhanced with the derivatization as much as with the heating condition while there was little synergy in employing both of the heating and derivatization together.