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Microbial conversion of EM574 and EM523, gastrointestinal motor stimulating agents
Y Funabashi1, S Hakoda, N Inatomi
1Discovery Research Division, Takeda Chemical Industries, Ltd., Japan.
Abstract:
EM574 exerts gastrointestinal motor stimulating (GMS) activity even after being converted to its metabolites P1 and P2 in dogs. These metabolites were isolated from dog liver using a series of chromatographic procedures. Their structures were determined to be the 15- and 14-hydroxyl derivatives of EM574, respectively, by spectral analysis. Large scale preparation by microbial transformation was investigated for further evaluation of the metabolites, because the amounts obtained by oxidation with dog liver homogenate were limited. Three strains of actinomycetes, Amycolatopsis tolypophorus IFO 13151, Dactylosporangium variesporum IFO 14104 and Nocardia capreola IFO 12847, were found to have the aiming oxidative potency. HPLC analysis of the crude extracts from these three cultures showed that the bioactive metabolites, EM574 P1 and P2 were produced. They were isolated from the culture broth with the other bioactive products EM574 P3 and P4. These bioactive products were prepared by large scale cultivation. EM574 P3 and P4 showed GMS activity comparable to that of EM574 P1 and P2. The structures of EM574 P3 and P4 were elucidated by spectral analysis and found to be the 3"-O-demethyl derivatives of EM574 P2 and EM574, respectively. Moreover, the absolute configuration at the C14 position of P2 was determined to be R by spectral analysis of the 6-membered cyclic carbonate of EM574 P2.
Insights
EM574 and its metabolites P1-P4 exhibit gastrointestinal motor stimulating activity. Microbial transformation using actinomycetes efficiently produced these active compounds for further study.
Area of Science:
- Pharmacology
- Microbiology
- Organic Chemistry
Background:
- EM574 demonstrates gastrointestinal motor stimulating (GMS) activity.
- This activity persists even after EM574 is metabolized into P1 and P2 in vivo.
Purpose of the Study:
- To isolate and characterize EM574 metabolites P1 and P2.
- To investigate microbial transformation for large-scale production of EM574 metabolites.
- To evaluate the GMS activity of newly identified metabolites P3 and P4.
Main Methods:
- Isolation of metabolites P1 and P2 from dog liver using chromatography.
- Structure elucidation of P1 and P2 via spectral analysis.
- Screening of actinomycetes strains for oxidative transformation of EM574.
- Large-scale cultivation and isolation of bioactive metabolites (P1-P4).
- Structure elucidation of P3 and P4 via spectral analysis.
- Determination of the absolute configuration of P2.
Main Results:
- Metabolites P1 and P2 were identified as 15- and 14-hydroxyl derivatives of EM574.
- Three actinomycetes strains (Amycolatopsis tolypophorus, Dactylosporangium variesporum, Nocardia capreola) showed potent oxidative activity.
- Bioactive metabolites EM574 P1, P2, P3, and P4 were produced and isolated.
- EM574 P3 and P4 exhibited GMS activity comparable to P1 and P2.
- P3 and P4 were identified as 3"-O-demethyl derivatives of EM574 P2 and EM574, respectively.
- The absolute configuration at C14 of P2 was determined as R.
Conclusions:
- Microbial transformation is a viable method for large-scale production of EM574 metabolites.
- The identified metabolites (P1-P4) retain significant GMS activity.
- Further investigation into these metabolites is warranted for their pharmacological potential.