Callus tissue culture of Coffea arabica L. cv Hybrido de Timor prepared from apical portions of orthotropic branches produced 49 to 92 times as much caffeine per unit weight of tissue as did the original explant. Cell-free extracts made from 42 to 54-day-old callus cultures in which active biosynthesis was occurring exhibited N-methyl-N (9)-nucleoside hydrolase and N-methyltransferase enzyme activities. Similar cell-free extracts exhibited selective biodegradative activity in forming urea from xanthine. Biosynthetic substrate specificities are similar to those of the enzyme obtained from green coffee fruit and tea leaves, suggesting that callus cultures of C. arabica form caffeine in the same way as the coffee fruit and tea leaves.