Caffeine inheritance was investigated in F2 and BC1F1 generations between Coffea arabica var. Bourbon Vermelho (BV) and Coffea canephora var. Robusta 4x (R4x). The caffeine content of seeds and leaves was determined during 2004 and 2005. Microsatellite loci-markers were used to deduce the meiotic pattern of chromosome pairing of tetraploid interspecific hybrids. Genetic analysis indicated that caffeine content in seeds was quantitatively inherited and controlled by genes with additive effects. The estimates of broad-sense heritability of caffeine content in seeds were high for both generations. In coffee leaves, the caffeine content (BSH) from the same populations showed transgressive segregants with enhanced levels and high BSH. Segregation of loci-markers in BC1F1 populations showed that the ratios of the gametes genotype did not differ significantly from those expected assuming random associations and tetrasomic inheritance. The results confirm the existence of distinct mechanisms controlling the caffeine content in seeds and leaves, the gene exchange between the C. arabica BV and C. canephora R4x genomes and favorable conditions for improving caffeine content in this coffee population.