Understanding the migration and transformation of nitrogen during biomass pyrolysis and the influence of cellulose components on the nitrogen-containing components of biomass is important for the clean and efficient utilization of biomass. In this study, we performed pyrolysis-photoionization time-of-flight mass spectrometry (Py-PI-TOF-MS), pyrolysis–gas chromatography/mass spectrometry (Py-GC/MS), and closed U-tube pyrolysis to analyze phenylalanine, glutamic acid, and their mixtures, the product composition of the primary and secondary reactions of amino acids, and the effect of glucose on the pyrolysis products of the two amino acids. The experimental results were combined with density functional theory calculations to reveal the pyrolysis path of the two amino acids and the mechanism of the effect of glucose on amino acid pyrolysis. The results revealed that phenylalanine undergoes deamination and decarboxylation reactions during pyrolysis. The hydrogen on the carboxyl group is transferred during this process to release CO₂ and generate phenethylamine. Phenylalanine generates a large amount of N-phenethylpropane-2-imine through a secondary reaction. Glucose may play a role in the hydrogen supply and promote the deamination of phenethylamine to produce styrene. The primary reaction product of glutamic acid is a nitrogen-containing heterocyclic ring. The pyrolysis product, l-pyroglutamic acid, generates 2-pyrrolidinone through a decarboxylation reaction. Glucose can reduce the reaction energy barrier for the formation of 2-pyrrolidinone.