The Biosynthesis of Tropane Alkaloids in <i>Datura </i><i>stramonium:</i> The Identity of the Intermediates between <i>N</i>-Methylpyrrolinium Salt and Tropinone

Abstract

The biosynthesis of the tropane alkaloids in transformed root cultures of Datura stramonium has been studied using sodium [1,2- 13 C 2 ]acetate, ( R,S )-[2,3- 13 C 2 ]-1-(1-methyl-2-pyrrolidinyl)propan-2-one {( R,S )-[2‘,3‘- 13 C 2 ]hygrine}, ethyl ( R,S )-[1,2- 13 C 2,2- 14 C]-2-(1-methyl-2-pyrrolidinyl)acetate, and ethyl ( R,S )-[2,3- 13 C 2,3- 14 C]-4-(1-methyl-2-pyrrolidinyl)-3-oxobutanoate. The incorporation of ( R,S )-[2‘,3‘- 13 C 2 ]hygrine into cuscohygrine and several other condensation products was high (15−40% specific incorporation), but label was not recovered in either tropine or tropine esters (hyoscyamine; 0.0 ± 0.5% specific incorporation). None of the recovered alkaloids was labeled when ethyl ( R,S )-[1,2- 13 C 2,2- 14 C]-2-(1-methyl-2-pyrrolidinyl)acetate was fed to the cultures. In contrast, sodium [1,2- 13 C 2 ]acetate and ethyl ( R,S )-[2,3- 13 C 2,3- 14 C]-4-(1-methyl-2-pyrrolidinyl)-3-oxobutanoate were incorporated into hyoscyamine (9 and 2% specific incorporation, respectively) and a number of other tropane alkaloids (up to 12% specific incorporation). These data provide further evidence that hygrine is not a direct precursor of tropane alkaloids. 13 C-Label from acetate was incorporated symmetrically into the C-2 and C-4 positions of (−)-hyoscyamine. The evidence supports a pathway in which acetoacetate reacts via its C-4 position with N- methyl-Δ 1 -pyrrolinium salt to give 4-(1-methyl-2-pyrrolidinyl)-3-oxobutanoate. This intermediate favors cyclization to give 2-carboxytropinone, tropinone being formed by decarboxylation.