One of the most studied mechanisms proposed to explain plant growth promotion due to Azospirillum inoculation is the production of bacterial phytohormones, plant growth regulators, and other related metabolites. Nowadays, we know that some beneficial effects of inoculation with rhizobacteria, particularly Azospirillum, can be correlated with the production and metabolism of chemically defined compounds with capacity to modify plant physiology. These include plant growth regulators such as auxins, cytokinins, gibberellins, abscisic acid, and nitric oxide, as well as cell structural molecules such as lipopolysaccharides, exopolysaccharides, and flagellin. In this chapter, we highlight the analytical and biological methods used to identify and quantify the most relevant phytohormones, plant growth regulators, and other related molecules produced by Azospirillum.