A molecular comparative study and a conformational analysis of a new 16-crown-5 ether were conducted using X-ray crystallography and NMR spectroscopy. The resulting X-ray crystal structure of the cyanoethyl sym-dibenzo-16-crown-5 ether showed three important conformational isomers, which are stable conformations not commonly seen in X-ray crystallography. The B3LYP/6-31G* method of the Gaussian 98 package program was also applied to study the new cyanoethyl sym-dibenzo-16-crown-5 ether. Theoretical conformation analysis was conducted using molecular mechanics and density functions to determine the relative stabilities of the three isomers of the cyanoethyl sym-dibenzo-16-crown-5 ether. The optimal geometric structure of the compound was determined. Reasonable agreement between the calculated values of the molecule mechanics and those obtained from the X-ray crystal structure was found. The ionization potentials, higest occupied molecular orbital and lowest unoccupied molecular orbital energies, energy gaps, heat of formation, atomization energies, and vibration frequencies of the compound were also calculated. The results of this study allow researchers to better understand the structure and properties of the new cyanoethyl sym-dibenzo-16-crown-5 ether.