BISBENZYLISOQUINOLINE BIOSYNTHESIS IN BERBERIS STOLONIFERA CELL CUL TURES

Abstract

Bisbenzylisoquinoline alkaloid-producing cell cultures of Berberis stolonifera were fed with 14C-Iabelled tyrosine, tyramine, and several chiral l-benzyl-l,2,3,4-tetrahydroisoquinolines. Berbamunine subsequently isolated from these cuItures showed significan tiy higher incorporation than 2-norberbamunine, berbamine, aromoline or isotetrandrine, suggesting that it is the first dimer formed in these cells. L-Tyrosine labelled the isoquinoline and benzyl moieties of berbamunine is ea equal ratios (1.2: 1). Tyramine was almost exclusively incorporated into the isoquinoline portion of this dimer. (R)-N-Methylcoclaurine gave the highest relative incorporations into bisbenzylisoquinolines (29% into berbamunine), followed by (R)-coclaurine (10%), (S)-coclaurine (5.2%), and (S)-Nmethylcoclaurine (2.4%). As both (S)-coclaurine and (S)-N-methylcoclaurine are well incorporated into the protoberberine fraction (21 and 54%, respectively), these results indicate that the dimerization pathway and the C-3'hydroxylation leading to reticuline are competing for these substrates. Feeding experiments with (1-13C)-(R)- and (S)coclaurine and NMR studies of the resulting alkaloids confirmed the biosynthetic route to berbamunine, and also showed that the (R) isomer is incorporated in similar isotopic excess into both halves of the (R, R) dimer guattegaumerine which had not been found previously in Berberis species. No racemization of (S)- to (R)-coclaurine (or its derivatives) or vice versa occurs in this tissue.