Doxycycline, an antibiotic, is largely used in human and veterinary medicine. The conventional treatment with activated sludge is not very efficient. Laccase appeared to be the main enzyme secreted essentially by white rot fungi as Trametes versicolor and Phlebia fascicularia on the degradation of xenobiotic compounds from the pharmaceutical industry. The main purpose of this study was to enhance the biodegradation of doxycycline through activated sludge combined with addition of glucose as a carbon co-substrate to improve the growth of the microbial population present in the activated sludge, phenol as a laccase mediator, copper as a cofactor and inductor for laccase production. The enhancement of the biodegradation of doxycycline was 50, 90, 68 and 83% greater respectively with the addition of glucose, copper, phenol and with a mixture of the compounds after 14 days of treatment at 25°C. Compared with the biotic control (activated sludge alone), a 30% increase for the test with the addition of phenol was observed.