Microbial transformation of propenylbenzenes may offer a cleaner and cheaper alternative to natural production of flavors and fragrances. In the present study, the biotransformation of trans-anethole using cells of a Colombian strain of the fungus Colletotrichum acutatum was investigated. Initially, fungitoxicity of this compound against C. acutatum was evaluated; trans-anethole displayed a relatively weak toxicity against the microorganism (<70%, at 200 μg/mL and after 48 hours) and apparently a detoxification mechanism was present. Then, the microorganism was incubated with the substrate at room conditions, using three different culture media (Czapek-Dox, Sabouraud and PDB). Results show that trans-anethole is mainly degraded through an epoxide-diol pathway (trans-anethole to anethole-epoxide, then to syn- and anti-anethole-diol, p-anisaldehyde, p-anisic acid and p-anisic alcohol). However, other minor metabolites [e.g. 3-(4-methoxyphenyl)-1-propanol, 1-(4-methoxyphenyl)-2-propanol, ethyl ester of anisic acid], possibly proceeding from other metabolic pathways were also found. Additionally, it was demonstrated that the concentration of metabolic products is dependent on culture medium used, being anethole-diol the major product obtained in all media used. Interestingly, some of the compounds generated in the biotransformation have been utilized as flavors and fragrances. Based on the identified metabolites, a possible metabolic pathway of the biotransformation of trans-anethole by C. acutatum was proposed.