This paper focuses on the development of an enhanced LC/ESI-MS method for the identification and quantification of fatty acids through derivatization. Fatty acids were derivatized with 2-bromo-1-methylpyridinium iodide and 3-carbinol-1-methylpyridinium iodide, forming 3-acyloxymethyl-1-methylpyridinium iodide (AMMP). This process attaches a quaternary amine to analytes and enabled ESI-MS in the positive mode of ionization with common LC mobile phases. Moreover, detection sensitivity was generally 2500-fold higher than in the negative mode of ionization used with underivatized fatty acids. The limits of detection were roughly 1.0-4.0 nM (or 10 pg/injection) for standard fatty acids from C10 to C24 and spanned approximately 2 orders of magnitude in linearity. AMMP derivatives had unique tandem mass spectra characterized by common ions at m/z 107.0, 124.0, and 178.0. Individual fatty acids also had unique fingerprint regions that allowed identification of their carbon skeleton number, number of double bonds, and double bond position. The derivatization method also allowed coding of analytes as a means of recognizing derivatives and enhancing quantification. 2H-Coding was achieved through derivatization with deuterated 3-carbinol-1-methyl-d3-pyridinium iodide. The 2H-coded derivatization reagent, 3-acyloxymethyl-1-methyl-d3-pyridinium iodide, was used in two ways. One was to differentially label equal fractions of a sample such that after being recombined and analyzed by ESI-MS all fatty acids appeared as doublet clusters of ions separated by roughly 3 amu. This greatly facilitated identification of fatty acids in complex mixtures. Another use of stable isotope coding was in comparative quantification. Control and experimental samples were differentially labeled with nondeuterated and deuterated isotopomers of CPM, respectively. After mixing the two samples, they were analyzed by ESI-MS. The abundance of a fatty acid in an experimental sample relative to the control was established by the isotope ratio of the isotopomeric fatty acids. Absolute quantification was achieved by adding differentially labeled fatty acid standards to experimental samples containing unknown quantities of fatty acids. Utility of the method was examined in the analysis of human serum samples.