Cycloenantiomeric hexahomooxacalix[3]arenes with different substituents on the three upper rims were synthesized for the first time by fixing their conformation into a cone. A cycloenantiomeric hexahomooxacalix[3]arene 6 was resolved into both enantiomeric forms and chiroptically characterized. Preliminary 1H-NMR studies indicated that the optically resolved cycloenantiomer 6 could discriminate the enantiomers of hydrochloride of phenylalanine ethyl ester.