Objective To investigate the inhibitory effect of the butyl alcohol extract of Phellinus igniarius decoction (BAEP) on the biofilm formation of Candida albicans standard strain SC5314 in vitro. Methods The double-dilution method was used to determine the minimal inhibitory concentration (MIC) and sessile minimal inhibitory concentration (SMIC50) of BAEP for C. albicans. XTT assay was used to analyze the influence of BAEP on the metabolic activity of C. albicans SC5314 biofilm. The solid medium was used to observe the effect of BAEP on the colonial morphology of C. albicans. An inverted microscope and a fluorescence microscope were used to investigate the effect of BAEP on the morphology and activity of C. albicans SC5314 biofilm, and a scanning electron microscope was used to observe the effect of BAEP on the morphological structure of C. albicans biofilm. Quantitative real-time PCR (qRT-PCR) was used to observe the changes in the transcriptional levels of biofilm-associated genes ALS1 and HWP1. Results The SMIC50 of BAEP was 1 024 μg/mL for C. albicans biofilm. The results of colonial morphology showed that BAEP 1 024 μg/mL affected the colony of C. albicans. BAEP 1 024 μg/mL inhibited the morphology and activity of C. albicans biofilm under the inverted microscope and the fluorescence microscope. The scanning electron microscope showed that BAEP 1 024 μg/mL had a marked inhibitory effect on C. albicans biofilm. The results of qRT-PCR showed that BAEP significantly reduced the expression of HWP1 and increased the expression of ALS1. Conclusion BAEP exerts a certain inhibitory effect on the biofilm formation of C. albicans SC5314.