Synthesis, antimicrobial and antiproliferative properties of epi-oligomycin A, the (33 S )-diastereomer of oligomycin A

: We describe the synthesis of epi-oligomycin A, a (33 S )-diastereomer of the antibiotic oligomycin A. The structure of (33 S )-oligomycin A was determined by elemental analysis, spectroscopic studies including 1D and 2D NMR spectroscopy, and mass spectrometry. Isomerization of C33 hydroxyl group led to minor changes in the potency against Aspergillus niger , Candida spp. , and filamentous fungi whereas the activity against Streptomyces fradiae decreased by approximately 20-fold compared to oligomycin A. We observed that 33-epi-oligomycin A had the same activity on the human leukemia cell line K562 as oligomycin A but was more potent for the multidrug resistant subline K562/4. Non-malignant cells were less sensitive to both oligomycin isomers. Finally, our results pointed at the dependence of the cytotoxicity of oligomycins on oxygen supply.

Fermentation was performed for 8 days at 28 °C in liquid medium. Isolation and purification were performed by extraction with acetone-hexane mixture followed by separation of the complex by HPLC and crystallization (Danilenko et al. 2012).
High-resolution electrospray mass spectra (HR-MS ESI) were recorded on a Bruker «micrOTOF-Q II» -MS instrument (Bruker Daltonics GmbH, Bremen, Germany). The samples were dissolved in methanol (0.10 mg/ml) and the solutions of the samples were injected directly into the ESI source by a syringe at a flow rate 3 μl/min. The mass spectrometer was operated under the following conditions: an endplate offset of -500 V, nebulizer pressure of 0.4 Bar, a drying gas flow rate of 4 l/min at 180°C and a capillary voltage of -4.5 kV and 4 kV in the positive and negative ionization mode, respectively. The instrument was calibrated with a Fluka electrospray calibration solution (Sigma-Aldrich, Buchi, Switzerland) that was 100 times diluted with MeCN. The accuracy was better than 0.43 ppm in a mass range between m/z 118.0862 и 2721.8948. All solvents used were purchased in best LC-MS qualities.
Optical rotations were measured on an Optical Activity AA-55 polarimeter (Optical Activity Ltd., Cambridgeshire, UK). NMR spectra were recorded on a Bruker Avance 600 spectrometer (Bruker, Germany) with proton resonance frequency 600 MHz. For the substance one-dimensional 1 H and 13 C NMR spectra were registered, as well as a series of two-dimensional spectra, namely 1 H-1 H scalar correlation COSY, 1 H-1 H space correlation ROESY and heteronuclear correlations 1 H-13 C-HSQC and 1 H-13 C-HMBC. About 10 mg of the substance was dissolved in 550μl of DMSO-d 6 .
The spectra were recorded at 298K. Chemical shifts were measured, in 1 H and 13 C spectra, relative to the signals of the solvent (DMSO-d 6 ). Two-dimensional spectra were recorded on TXI (Triple Resonance Inverse) probe with the pulsed field gradient. The COSY experiment was realized with double quantum filter (DQF). The gradient coherence selection with detection by the method of echo-antiecho was used for HSQC spectrum. The ROESY experiment was recorded with a mixing time 300 ms.

Cell culture and antiproliferative activity
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