The effect of alkyl chain length on adsorbent pore volume and void volume of HPLC columns is described. The results provide evidence that alkyl chains attached on silica surface are densely packed. A correlation of a decrease of pore volume with an increase of the alkyl modifier chain length was found. Effective molecular volume of bonded chains was found to be similar to the molecular volume of corresponding liquid alkanes. An absence of noticeable penetration of acetonitrile, methanol, or tetrahydrofuran molecules between bonded chains at any water-organic eluent composition was found.