Fungal pathogens can invade not only the fruit peel but also the outer part of the fruit mesocarp, limiting the efficacy of fungicides. In this study, the relationships between fungicide structure, diffusion capacity and in vivo efficacy were evaluated for the first time. The diffusion capacity from pear peel to mesocarp of 11 antifungal compounds, including p-aminobenzoic acid, carbendazim, difenoconazole, dipicolinic acid, flusilazole, gentamicin, kojic acid, prochloraz, quinolinic acid, thiophanate methyl and thiram was screened. The obtained results indicated that size and especially polarity were negatively correlated with the diffusion capacity. Although some antifungal compounds, such as prochloraz and carbendazim, were completely degraded after a few days in peel and mesocarp, other compounds, such as p-aminobenzoic acid and kojic acid, showed high stability. When applying the antifungal compounds at the EC50 concentrations, it was observed that the compounds with high diffusion capacity showed higher in vivo antifungal activity against Alternaria alternata than compounds with low diffusion capacity. In contrast, there was no relationship between stability and in vivo efficacy. Collectively, the obtained results indicated that the diffusion capacity plays an important role in the efficacy of fungicides for the control of pear fruit diseases.