The molecular composition of fossil resin from the Eocene Ameki Formation, southern Nigeria has been analyzed by infrared spectroscopy, pyrolysis-gas chromatography-mass spectrometry and thermochemolysis gas chromatography-mass spectrometry to determine the structural class and botanical source of the resin. The pyrolysis products were dominated by bicyclic products derived from regular labdatriene structure and lacked succinic acid indicating Class Ib type amber. The biomarker compositions in the pyrolysates are dominated by sesquiterpenoids of the cadinane and bisabolane classes that are common constituents of higher plants and labdane type diterpenoids. The exclusive presence of labdane type diterpenoids and the absence of higher plant triterpenoids strongly suggest that the amber was derived from a gymnosperm (conifer) botanical source. This observation contrasts with the angiosperm source previously suggested for the resin based on the palaeobotanical studies of the host sediments. The specific family source within the conifers could not be ascertained based on the molecular composition of the resin. However, the presence of methyl-16,17-dinor-callitrisate pointed to the Cupressaceae family although other Cupressaceae biomarker indicators were not detected in the resin.