There is a considerable industrial interest in both the scale-up and optimization of chromatographic operations used in the purification of fine chemicals and biomolecules. One of the major factors affecting the adsorption operation is the adsorption isotherm. Reversed-phase high-performance liquid chromatography (RP-HPLC) was used to measure the adsorption isotherm of phenol and a-cresol. From the experimental results, the retention times were decreased with increasing sample sizes, and the front of the peak was very stiff, so Langmuir adsorption isotherm was applied. Also Early-eluting component, phenol, in a mixture of the two components elutes faster than that in a pure component. Pulsed-injection method (PIM) was used to determine the two parameters of the Langmuir isotherm. The resulting parameters, a and b, were used to calculate the elution profiles of phenol and o-cresol in pure and mixed state. The agreement between the experimental data and calculated elution profiles was fairly good in a mixture as well as a pure component.