Microbial community structure is one of the important factors controlling the pollutant-degrading capacity of ecosystems. The analysis of microbial quinones has gained increased recognition as a simple and useful approach for studying microbial structure in environmental samples. The analytical precision of quinone characterization using high performance liquid chromatograph (HPLC) with a UV-detector was studied in this study. Activated sludge was used as a typical mixed culture. The coefficient of variation of quinone content was lower than 6%, and that of microbial diversity calculated from the composition of quinones was as low as 3%. Statistical analyses on the analytical precision of quinones demonstrated that the critical value of dissimilarity between two quinone profiles of activated sludge, which is used to make a judgement whether the two quinone profiles are different or not, is 0.1 for the analytical method used in this study. The values of minimum biomass required for quinone analysis to have a reliable analytical result of microbial quinones were 2 mg-dry-cell for activated sludge.