A study was made to define the validity of stoichiometric equations for the overall reaction of bacterial metabolism, using mixed microbial populations and soluble carbonaceous organics as substrates. In the study, simultaneous measurements of oxygen utilization, substrate removal, synthesized cell materials and COD of the mixed-liquor suspension were made. Oxygen uptake was measured by the Warburg manometric technique; substrate and mixed-liquor COD by the dichromate procedure; synthesized cell materials by the membrane filtration technique. During the reaction, multiple carbon balances, expressed in percent of the total biological oxygen demand, were carried out and are presented graphically. Within the experimental error of the determinations involved, the stoichiometric equations written for glucose, glutamic acid and sodium acetate metabolism are valid throughout the progression of bacterial growth. The experimental results illustrate the usefulness of the total biological oxygen demand concept in the field of environmental engineering. Carbon balances can be employed in the investigation of the biological degradability of complex compounds, and delineation of the kinetics involved in the system.