AbstractBiased Brownian movement, a key process of biological molecular motors, was achieved by the neutralization of a rotaxane consisting of a secondary ammonium salt, in which the ammonium group is located next to the terminal group, and crown ether. Active transport occurred kinetically upon fixation of the transient pseudorotaxane by the rapid acylation of the amino group. The bulky terminal group enhanced the biased Brownian movement, although it suppressed the formation of the pseudorotaxane. Efficient active transport was possible when a large excess of acylation agent and triethylamine were used at 0 °C with 1.5 equivalents of crown ether. The efficiency of active transport increased as the electrophilicity of the acylation agent increased. The highest efficiency of active transport was observed with the cyclopentyl terminal group with one methylene spacer. A simple, two‐step, unidirectional transport was carried out using the active transport system based on the cyclopentyl terminal group.