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Biological adhesive pads of some reptiles and insects, such as tree frogs and grasshoppers, are covered with planar microstructures and have strong and stable adhesive ability on both wet and dry substrates. These adhesion forces do not mainly come from van-der-Waals force but wet adhesion. In this study, the influence of substrates' surface roughness on the wet adhesion of man-made adhesive pads inspired by tree frog toe pads is investigated experimentally. Biomimetic polydimethylsiloxane adhesive pads with planar hexagon microstructures with a microchannels width of 10 μm are fabricated by combining electroforming with soft lithography. Experiments of wet adhesive force between the pads and sandpaper slices with different average surface roughness are carried out at various preloads. Results show that the rougher surface leads to the decrement of wet adhesion force. It is also observed that if the microcosmic profile height of the substrates is near or less than the width of microchannels in the biomimetic adhesive pads, the microstructures and preloads can increase significantly the wet adhesive force, otherwise the microstructures and preloads do not contribute indistinctively. The experimental results can be explained by analysing the relation between the solid contact area and the area with a liquid bridge.