Stoichiometric relationships for biological reactions involving intermediate formation are developed from microbial reaction fundamentals and thermodynamic principles. Biological reactions proceed through intermediates, which sequester carbon and electrons whenever their degradation is relatively slow. Modeling intermediate formation and subsequent utilization requires evaluation of the distribution of electrons, energy, and macronutrients (C and N) between energy-generating pathways and cell-synthesis pathways for each step in the mineralization of the primary electron-donor substrate. We describe how energy and electron ba lances a re utilized to predict the stoichiometry for each step of a multi-step degradation process. Each stoichiometric relationship developed predicts substrate utilization, cell growth, and the formation of other products (e.g., H2CO3 or H+) for one step in the pathway to full mineralization. A modeling example demonstrates how different kinetics for each step in the degradation of nitrilotriacetic acid (NTA) leads to observed patterns in experimental results, such as a delay in the release of H2CO3 after NTA is removed from solution. (C) 2000 John Wiley & Sons, Inc.