A mixed population of heterotrophic bacteria is kept in the dark at 20 degrees for five days in a 300 ml sample of wastewater diluted with phosphate buffer (pH 7, 2). Inorganic growth elements, acclimated microorganisms and oxygen can be added to facilitate the BOD test. For the determination of carbonaceous BOD a nitrification inhibitor can be added. The amount of oxygen consumed by the microbes is measured at time 0 and then after five days of incubation, using chemical procedures, an oxygen electrode, or a BOD apparatus. BOD is calculated using the equation: BOD (mg/L) = (D1-D5 )/ P where D1= initial dissolved oxygen (DO), D5 = DO at day 5, and P= decimal volume fraction of wastewater used. The BOD5 test is the most commonly used. In addition to determining the amount of oxygen required for biological treatment of organic content in wastewater, this method provides information on the required size of the treatment plant, efficiency and compliance with wastewater permits