The molecular packing in triacylglycerols having different acyl chains has been examined by differential scanning calorimetry, powder X-ray diffraction and vibrational spectroscopy (infrared and Raman) techniques. In the triacylglycerols examined, the acyl chain length, unsaturation or the position of substitution on the glycerol were changed systematically to observe their influence on the molecular packing in different polymorphic forms. Variation in the 3-acyl chain length of 1,2-dipalmitoy1-3-acyl-sn-glycerols (PPX) influenced the molecular packing along the long axis in the stable polymorphic forms. Three different modes of packing were observed. If X < 4, the compounds packed in a bilayer structure similar to diacylglycerols, or if X > 10 and < 16 the compounds packed in a bilayer structure but similar to mono acid triacylglycerols. However for intermediate 3-acyl chain lengths, as in PP6 and PP8 the stable packing can occured only through chain segregation resulting in a trilayer structure. In the triacylglycerols containing unsaturated acyl chains, 1,2-dioleoy1-3-acyl-sn-glycerols (00X) and 1,3-dioleoy1-2-acyl-sn-glycerols (0X0) the stable polymorphic forms packed in a trilayer structure where the odd acyl chains segregated and formed a middle layer. In a metastable hexagonal packing α-phase) the long range ordering is minimal. Because of this lack of specific chain-chain interaction the 3-short acyl chain compounds of PPX packed in a unimolecular length structure (except PP2) whereas the 3-long acyl chain compounds packed in a bilayer structure. In orthorhombic perpendicular and triclinic parallel packing where the specific chain-chain interaction is increased, the end plane methyl packing and the glycerol conformation played important roles in the formation of bi-, tri- and hexalayer structures. The driving force in the formation of these different structures is to minimize the crystal defects created by the odd acyl chains and to enhance the specific chain-chain interactions. The presence of an odd acyl chain influenced the lateral chain packing as well, e.g., the stability of the orthorhombic perpendicular packing is enhanced by the presence of an odd acyl chain and even in some cases it is favored over the triclinic parallel packing. The odd acyl chain at the 1- or 3-position of -sn-glycerol stabilized the orthorhombic perpendicular packing. This indicates the glycerol conformation is probably perpendicular to the layer plane and thus is different from the monoacid triacylglycerols.