Self-assembled monolayers (SAMs) on metal surfaces offer a convenient approach for fabricating molecularly tailored interfaces with well-defined compositions, structures, and thicknesses. This work uses molecular dynamics (MD) simulations to investigate the effect of temperature on the structure and barrier properties of n-alkanethiolate [CH3(CH2)n-1S] SAMs on gold, with chain lengths varying from n = 8 to 30. Our MD simulation system consisted of 90 flexible n-alkanethiolate chains held at a fixed surface density and periodically replicated in the plane of the gold surface, and and the transport of a molecular species through the SAMs was simulated using the “z-constraint algorithm”. Structurally, the chain ends became highly disordered at approximately 325 K for all investigated chain lengths, and further increases in temperature caused the middle portion of the monolayer to become progressively less crystalline. MD simulations on these SAMs also showed that the through-film diffusion coefficient for oxygen decreases with increasing temperature from 200 K to 400 K as the monolayers became less crystalline and contained a lower free volume alignment in the z direction. Above 400 K, the through-film diffusion coefficients increased with temperature as the kinetic energy for the diffusing molecule increased and there was little change in the free volume alignment within the SAM. The maximum free energy barrier against transport of molecular species for these monolayers remained almost constant with the increasing temperature possibly due to at least some part of the middle region of SAMs remaining relatively ordered at these temperatures, at least up to 400 K.