Aims: This research aims to investigate the efficiency of two lipolytic enzymes - fungal cutinase and yeast esterase - upon the biodegradation of dihexyl phthalate (DHP). Method and Results: During the enzymatic degradation of DHP dissolved in methanol, several degradation products were detected and their time-course changes were monitored using GC/MS. The DHP-degradation rate of cutinase was surprisingly high; i.e. almost 70% of the initial DHP (500 mg l(-1)) was decomposed within 4.5 h. Although the same amount of esterase was employed, more than 85% of the DHP remained after 3 days. Almost all the DHP was converted by cutinase into 1,3-isobenzofurandione (IBF), whereas hexyl methyl phthalate and IBF were abundantly produced by esterase. In addition, the toxicities of the DHP-degraded products by esterase were evaluated using various recombinant bioluminescent bacteria, which caused oxidative and protein damage, whereas the hydrolysis products from cutinase never caused any cellular damage in the methanol-containing reaction system. Conclusions: Cutinase starts to act as a DHP-degrader much earlier and faster than esterase, with high stability in ester-hydrolytic activity, therefore a plausible approach to the practical application of cutinase for DHP degradation in the DHP-contaminated environments may be possible. Significance and Impact of the Study: This study describes the enhanced degradation and detoxification of DHP using Fusarium oxysporum f. sp. pisi cutinase.