The accumulation of perfluorooctanoic acid (PFOA) has been detected in wildlife, soil, and water. Further, 8:2 fluorotelomer alcohol (8:2 FTOH) is used for the industrial synthesis of other fluorotelomer compounds, surfactants, and polymeric materials; however, it was recently found to be a potential source of PFOA contamination in the environment. 1H,1H,2H,2H,8H,8H-perfluorododecanol (degradable telomer fluoroalcohol (DTFA)), which is a newly developed fluorotelomer, contains the -CH2- group in the fluorinated carbon backbone, making it potentially degradable through biological reactions. In this study, we investigated the biodegradation of DTFA in a mixed bacterial culture obtained from activated sludge. Optimized quantitative liquid chromatography-mass spectrometry analysis of the predicted metabolites generated in the culture revealed accumulations of the transformation products from DTFA to 2H,2H,8H,8H-PFDoA and 2H,8H,8H-2-PFUDoA via multiple processes. Furthermore, the production of short fluorinated compounds, perfluorobutanoic acid, perfluoropentanoic acid, and perfluoropentanedioic acid, which are believed to have lower accumulation potential and toxicity toward organisms than PFOA, was determined.