In the current steady state design and kinetic simulation models for activated sludge systems, the heterotrophic active biomass (XH) is a key parameter. However, this parameter remains hypothetical within the structure of the models; it has not been measured directly, primarily due to the lack of suitable simple experimental techniques. In this paper a simple batch test procedure is used to quantify XH in mixed liquor samples drawn from a well-defined parent anoxic/aerobic activated sludge system operated at sludge ages 12d and 20d. The measured XH concentrations are in close agreement with those calculated theoretically for the parent system at 12d sludge age, but are about 1/2 the theoretical values for the system at 20d sludge age. While the good correspondence at 12d sludge age provides substantive direct evidence supporting both the models and the experimental method, reasons for the poor correspondence at 20d sludge age need to be found.

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