Biotransformations Of (S)-N, N-dimethyl-3-keto-3-(2-thienyl)-1-prapanamine

Duloxetine, trade name'Cymbalta'and chemical name (S)-N-methyl-3-(1-naphthalenyloxy)-3-(2-thienyl)-1-prapanamine, is used as an antidepressant by inhibiting the reuptake of norepinephrine and serotonin.(S)-N,N-dimethyl-3-hydroxy-(2-thienyl)-1-propanamine [(S)-DHTP] is a key intermediate during the synthesis of Duloxetine. There were many reports related to the synthesis of (S)-DHTP by chemical and chemoenzymatic methods, while few was involved in the biotransformation of (S)-DHTP from its precursor.This research aimed to screen out the fungus capable of reducing N, N-dimethyl-3-keto-3- (2-thienyl)-1-propanamine (DKTP) enantioselectively to (S)-DHTP, then, to identify it by amplification and sequence analysis of its 26S rDNA D1/D2 region and to determine its growth conditions by culture it in different media. Finally, in order to obtain a more economical and reasonable synthetic route for (S)-alcohols, the transformation conditions of the selected fungus were optimized. The results were as followed:Firstly, a new fungus strain, which can asymmetrically reduce DKTP to (S)-DHTP with high yield (more than 90%) and enantiometric excess (more than 98%), was isolated by the using of two screening methods, one reported by others, another improved by ourselves. It was proven that, compared to the reported method, the improved method was more convenient and efficient.Secondly, the newly isolated strain was identified as one of the Rhodotorula Harrison through morphological and DNA test method. So it was named as Rhodotorula sp. 507.Thirdly, it was found that the optimal growth condition of Rhodotorula sp. 507 was 30℃, pH 7.0 and 48h after cultured. 1 g·L-1 glucose and 1.2 g·L-1 peptone were its optimal carbon and nitrogen source, respectively.Lastly, the results of optimization showed that the highest conversion rate of (S)-DHTP could be obtained when Rhodotorula sp. 507 was cultured for 16h, at the concentrations of the inducer (acetophenone), substrate and the glucose added later were 0.24 g·L-1, 1 g·L-1and 1 g·L-1, respectively. At this condition, it could yield (S)-alcohol at 95% conversion and more than 99% enantiometric excess.So, it could be concluded that, because of its high specific ketone reductase, Rhodotorula sp. 507 could transfer DKTP into (S)-DHTP under the culture condition established in this study efficiently. The application of this fungus would make it easy to get enough material for the synthesis of Duloxetine and would be very benefit to the therapy of depression. Furthermore, the purification and modification of the ketone reductase involved in this process would be very important to the manufacture of other chrial drugs.