During pretreatment of corn stover several degradation compounds are produced that may inhibit downstream biological processing. The nature of these degradation products can vary depending on the thermochemical severity of pretreatment. Ammonia Fiber Expansion (AFEX) pretreatment produces a set of nitrogen-containing compounds including phenolic amides, sugar amides, and amines. These compounds are specific to AFEX over other pretreatment methods. Degradation products normally derived directly from plant biomass, such as ferulic acid, p-coumaric acid, and acetic acid, have amide analogs detected in AFEX-pretreated samples. It is important to determine the types and quantities of these nitrogenous compounds in pretreated biomass. The loss of ammonia through undesirable reactions to various inhibitory compounds is a potential hurdle to making cellulosic ethanol economically feasible using AFEX. On the other hand, the process may offer access to valuable co-products derived from these reactions. Described here are the GC-MS and LC-MS methods developed for identification and quantification of some important nitrogenous compounds. AFEX pretreated biomass was also characterized by NMR to study the incorporation of nitrogen to lignin, phenolics and reducing sugar aldehydic groups. Some of these methods were subsequently used to examine differences in degradation product concentration for untreated, AFEX-pretreated, and acid-pretreated corn stover. The effect of AFEX pretreatment severity on the formation of various nitrogenous compounds is also investigated.