Taxol is a curatively effective but rare anti-tumor agent extracted from Taxus (yew) barks; however, the high cost and low production of the extraction method have limited its widespread use. In this study, fungi isolated from Taxus chinensis var. mairei were tested for their ability to produce taxol. High performance liquid chromatography combined with mass spectrometry confirmed that Aspergillus aculeatinus Tax-6, one of the endophytic fungi, could produce taxol in potato dextrose agar liquid medium. NaOAc, Cu2+, and salicylic acid were introduced into the medium to enhance taxol production of strain Tax-6 because NaOAc is an important precursor of taxol, Cu2+ may enhance the activity of oxidase and catalyze the formation of taxol, and salicylic acid could be an elicitor signal. Application of response surface methodology to optimize the culture led to the addition of CuSO4, salicylic acid and sodium acetate at 0.1 mg L-1, 10 mg L-1 and 8 g L-1 to improve taxol yield from 334.92 to 1337.56 μg L-1. Overall, the results of this study confirmed that fungal taxol has the potential to be broadly applied by optimizing the culture conditions.