To examine whether the rate of nucleotide substitution is affected by generation time of the organism, I attempted to estimate an accurate rate of synonymous (silent) substitution in Drosophila lineages, using alcohol dehydrogenase (Adh) and heat shock protein 82 (hsp82) genes. The results obtained suggest that the rate of synonymous substitution in Drosophila lineages is roughly 10^-8 per site per year. This rate is approximately two times higher than that of rodents and ten times greater than higher primates. The higher rate in Drosophila may be explained by the shorter generation times of the Drosophila species, though the possibility that the mutation mechanism in Drosophila may differ from that in mammals cannot be excluded.