Hevea brasiliensis was introduced to Thailand for the production of natural rubber. RRIT251 and RRIM600 are high yield varieties while PB350 produces a high yield of latex and resistance to diseases. The cultivar identity must be ascertained prior to plantation establishment because it is important to relate the genetic background with the production of certain secondary metabolites. In this study, the internal transcribed spacer region was used to evaluate the varieties among these 3 cultivars. Callus induction was initiated with 2 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) and 2 mg/L 6-benzylaminopurine (BAP). Hexadecanoic acid, methyl ester and 9-octadecanoic acid (Z), methyl ester were detected in RRIM600 calli. 2-Furancarboxaldehyde-5-(hydroxymethyl) was dominant in the RRIT251 and PB350 calli. These secondary metabolites are relevant in various high performance commercial sectors.