Invasive fungal infections (IFIs) are emerging as serious infectious diseases worldwide, and due to the lack of effective antifungal agents and serious drug resistance, the limited efficacy of existing drugs has led to high morbidity and mortality in patients. We optimized the lead compound 7 by conformational restriction strategy to obtain a series of 3-thiophene phenyl compounds, of which compound 21b showed excellent inhibitory activity against pathogenic and drug-resistant fungi. In addition, the preferred compound 21b could prevent the formation of fungal biofilms and displayed satisfactory fungicidal activity. Furthermore, compound 21b was almost non-toxic to mammalian THLE2 and RAW264.7 cells and did not pose a risk of drug-drug interactions. These results strongly suggested that compound 21b is worthy of further study as a potential azole inhibitor.